Pollen Germination and Tube Growth

See discussions, stats, and author profiles for this publication at: https://www.researchgate.
net/publication/226003062
Pollen Germination and Tube Growth
Chapter · October 2010

DOI: 10.1007/978-3-642-02301-9_13
CITATIONS READS
206 13,358
2 authors, including:
Weicai Yang
Institute of Genetics and Developmental Biology, CAS
141 PUBLICATIONS 7,220 CITATIONS
SEE PROFILE
All content following this page was uploaded by Weicai Yang on 05 June 2015.
The user has requested enhancement of the downloaded file.

Chapter 13
D.-Q. Shi and W.-C. Yang
Plant Developmental Biology -

Biotechnological Perspectives: Volume 1,
Page 245-282
Pua, Eng Chong; Davey, Michael R. (Eds.)
Springer-Verlag Berlin Heidelberg 2010
Chapter 13
13.1 Introduction
In higher plants, the sexual organs, e.g. anther and pistil, are separated spatially
either within a flower in hermaphrodite plants or in different flowers as in monoe-
cious plants. Successful fertilization requires temporally and spatially coordinated
development of male and female organs and gametophytes. Mechanisms have also
been evolved to ensure pollen survival in harsh environments and to deliver the
male gametes to the female gametophyte enclosed within the ovule, which in turn is
embedded in the pistil. Furthermore, crosstalk between the male and female
gametophytes ensures the success of fertilization that occurs at high efficiency
and in a controlled manner.
The advent of multicellular gametophyte and siphonogamy in higher plants is
instrumental in improving efficiency of sexual reproduction. In species with tricel-
lular pollen grains, like Arabidopsis and rice, mature pollen grains are composed of
two small sperm cells and a large vegetative cell. However, in tobacco, maize and
other species with bicellular pollen grains, mature pollen grains contain a large
vegetative cell and a small generative cell that will precede the last mitosis to
produce two sperm cells in the pollen tube. The pollen tube, a tubular structure that
originates from the vegetative cell, will deliver the two sperms to the female
gametophyte, a phenomenon called siphonogamy (Fig. 13.1). In addition to the
cells in pollen grains, specialized pollen walls have been developed to protect
the male gametes and to meet the requirement for dispersal and interaction with
the pistil. With respect to the female gametophyte, it is composed typically of seven
cells, namely an egg cell, two synergids, a central cell and three antipodal cells that
often degenerate before fertilization (Yang and Sundaresan 2000). The egg,

The CAS Key Laboratory of Molecular and Developmental Biology, Institute of Genetics and
Developmental Biology, Chinese Academy of Sciences, Beijing 100101, China
e-mail: [email protected]
E.C. Pua and M.R. Davey (eds.), 245

Plant Developmental Biology – Biotechnological Perspectives: Volume 1,
DOI 10.1007/978-3-642-02301-9_13, # Springer-Verlag Berlin Heidelberg 2010
246 D. Shi and W. Yang
Fig. 13.1 Overview of pollen tubes in Arabidopsis. a Micrograph showing a germinating pollen
(left) and pollen tube developing in vitro; b the same micrograph as in a showing sperm (arrow-
heads) and the vegetative nucleus as revealed by DAPI staining; c scanning electron micrograph
showing a germinated pollen grain (Pg) and pollen tube (*) on a papillar cell (Pc). Scale bars in a, b
or c are 10 mm
together with central cell and synergids, forms a functional female germ unit (FGU)
to fulfil the task of pollen tube attraction, sperm release and transportation, and
double fertilization. This chapter reviews our current understanding of pollen
germination and tube growth, with a focus on Arabidopsis and other model species.
13.2 Mature Pollen Grains
As mentioned above, a tricellular pollen grain consists of two small sperm cells
enclosed within a large vegetative cell, and together they form a functional assem-
blage, the male germ unit (MGU), which plays a potential role in the transport and
delivery of sperm. The sperm and the vegetative cell can be easily distinguished by
their size and nuclear morphology when pollen grains are released from the anther.
The vegetative nucleus is highly lobed, while the sperm nucleus is elongated. Pollen
grains undergo a maturation process prior to anther dehiscence to prepare them-
selves for survival, crosstalk with the female sexual organ, germination and tube
growth. This process includes the elaboration of the pollen wall, dehydration and
accumulation of storage materials.
13 Pollen Germination and Tube Growth 247
13.2.1 Pollen Wall
The pollen wall is more complex than other cell walls in plants. It is composed of
several layers, with a highly organized ultrastructure of each layer. The pollen wall
is usually divided into three principle strata at maturity, namely the intine, exine
and pollen coat, with the relative amount of each varying between species. The
intine is the innermost wall made primarily of multilayered cellulose. It shares
many of the chemical and structural features with the primary wall of a somatic cell.
Immediately outside the intine is the reticulated exine that is composed of the
chemically resistant biopolymer sporopollenin and interrupted by openings called
apertures. The exine can be further divided into an outer sculptured layer, the
sexine, and a simple inner layer, the nexine. Outside the reticulate exine layer
resides the pollen coat that contains lipid, protein and other material deposited from
the tapetum of the anther.
As the most specialized cell wall layer, the exine, provides the distinctive and
characteristic features of pollen grains and spores. The exine pattern of the reticu-
late pollen wall is a unique feature of a species and can be divided into three major
classes, namely baculate, granulate and spinulate. It has been important in palynol-
ogy since the early 1970s because of its vast morphological diversity and composi-
tion of sporopollenin. Prior to the formation of the intine layer, exine formation is
initiated from the tetrad stage during microsporogenesis. The glycocalyx-like
fibrillar polysaccharide material, primexine, is deposited originally onto the micro-
spore surface between the plasma membrane and the b-1,3-glucan (callose) wall.
Subsequently, the precursors of sporopollenin, for example, the long-chain fatty
acids, aromatic compounds, phenolics and carotenoid (Guilford et al. 1988) that are
synthesized in tapetal cells, are polymerized and accumulated to form sporopollenin
at raised sites of the plasma membrane, leading to the formation of the probacula
within the primexine matrix. The prebacula elongates until it reaches the callose wall
to form the bacula structure. At the end of the tetrad stage, as callose starts to
dissolve, a structure called the tectum appears on top of the bacula, and simulta-
neously a foot layer is formed at the basal bacula (Paxson-Sowders et al. 1997,
2001). The primexine patterning requires coordination between the extracellular
callose layer, the plasma membrane, and the underlying cytoskeleton, vesicles and
endoplasmic reticulum. Callose deposition is fundamental to the exine patterning
and formation. For example, mutation of a male-specific b-1,3-glucan synthase
gene, CalS5, disrupts the exine organization (Nishikawa et al. 2005). It has been
proposed that callose can increase the local concentration of primexine subunits by
trapping these elements and preventing them from diffusing into other locules. In
addition, callose can function as a physical support during primexine assembly or
interacts directly with primexine nucleation sites.
Sporopollenin is a biopolymer that is resistant to non-oxidative physical,
biological and chemical degradation processes (Blackmore 2007). Thus, as sporo-
pollenin polymerizes, the exine becomes less elastic and increasingly resistant to
acids, bases and enzymes. The spines or other characteristic structures that are
attributed to wide variety of taxa are formed at the surface of microspores. As exine
patterning commencing from the tetrad stage continues, the aperture is set where
the exine is much reduced or virtually absent between the plasma membrane and the
adjacent callose wall. Apertures of exine are an important feature for taxonomic
classification; they differ in number, distribution and architecture within families,
species or even a single plant. Apertures are the sites for pollen tube exit; they also
function in water uptake, transfer of recognition substances and accommodation of
volume changes (Furness 2007). Despite morphological description, molecular
mechanisms and factors controlling sporopollenin accumulation, aperture position-
ing and exine patterning remain to be clarified (Blackmore 2007).
The pollen coat refers to lipoidal substances covering mature pollen grains of
entomophilous plants. The major substructure of the pollen coat is pollenkitt or
tryphine, which refers to the extracellular lipidic substance that originates from
plastids of tapetal cells and that covers the outer surface of the exine. Pollenkitt and
tryphine may appear identical, although they differ in ontogenesis (Pacini 1997).
The pollen coat is composed of lipids, proteins, pigments and aromatic compounds
that fill the sculptured cavities of the pollen exine. A large proportion of the coat
lipids are very long-chain lipids (C29 and C30) that account for about 70% of the
coat lipids (Mayfield and Preuss 2000). In Arabidopsis, lipases and oleosin-like
proteins amount to >90% of the detectable pollen coat proteins larger than 10 kD
(Mayfield et al. 2001). Together, pollen coat substances account for 10–15% of
total pollen mass (Piffanelli et al. 1997). Besides the protective function against
environmental attack, the lipids and proteins of the pollen coat mediate the hydra-
tion of pollen grains on stigma cells and the recognition between pollen and the
stigma. Proteins in the pollen coat may also act as ligands and receptors in this
recognition system and/or components needed for efficient pollination. The cysteine-
rich protein (SCR) residing in the pollen coat was discovered as the male
recognition factor in self-incompatibility (SI) in Brassicaceae (Schopfer et al.
1999; Shiba et al. 2001). In Arabidopsis pollen, the major coat protein is an
oleosin-like coat protein, GRP17-1. Knockout of GRP17-1 causes delay in pollen
hydration and reduced transmission, suggesting that GRP17-1 is essential for
initiation of pollen hydration on the stigma and for pollen fitness. However, the
molecular mechanism of GRP17-1 function in pollen hydration is not clear. Oleo-
sin, a major component of the oil body in seeds of oil crops, plays a role in
preventing oil body fusion during seed dehydration. It is not clear whether the
oleosin-like proteins play a similar role for the pollen coat. They may either
communicate directly with the stigma or mediate the biophysical properties of the
pollen coat (Mayfield and Preuss 2000).
13.2.2 Pollen Maturation
Before anther dehiscence, pollen undergoes a maturation process, which prepares

the grain for independent living and subsequent activities for sexual reproduction.
The maturation processes include pollen dehydration and accumulation of storage

materials.
13.2.2.1 Pollen Dehydration
Mature pollen grains represent an arrested developmental stage in the life cycle of
higher plants because of dehydration before dispersal. Dehydration is the loss of a
certain amount of water from fresh grains. Pollens of most plant species (70%)
undergo dehydration before anthesis. Pollen grains vary considerably in their
degree of dehydration at dispersal, depending on their habitat. Dehydration causes
physical changes, such as in size and shape, influencing speed of pollen transport, as
well as harmomegathic changes in the appearance of the pollen wall. More impor-
tantly, pollen dehydration maintains pollen grains in a “dormant” or “quiescent”
state, because the water content is approximately 15–35% that of active metabo-
lism, and therefore increases the ability to withstand changes in environmental
conditions after dispersal. Dehydration is thus highly related with pollen viability
and composition of reserves within the grains. In Cucurbita pepo and several
species of Gramineae, pollen grains are dehisced in a hydrated state and remain
metabolically active. Thus, they are short-lived (Nepi and Pacini 1993). Water is
tied up with membrane components during dehydration and reaches a threshold
point at the end of dehydration. Consequently, the phospholipid material of the
membrane system is converted from a liquid crystalline form to a gel state. In
addition, the degree of dehydration is highly correlated with contents of carbohy-
drates and lipids in mature pollen grains.
13.2.2.2 Reserves in Mature Pollen Grains
The reserves in mature pollen grains are mainly carbohydrates, which are used as
nutrient storage for energy resource or synthesis of pollen tube wall precursors.
Proteins are usually absent in the reserves or their concentrations are lower than that
in seeds. Pollen carbohydrate reserves at maturity may exist in different forms: (1)
polysaccharides such as starch in amyloplasts or polysaccharides in cytoplasmic
vesicles, (2) disaccharides such as sucrose and (3) monosaccharides such as glucose
and fructose (Pacini 1996). Starch serves as an energy supply or is converted to
other substances during pollen development and maturation (Kuang and Musgrave
1996). In most members of the Graminaceae, pollen grains contain a large amount
of starch, with only a small quantity of soluble sugars, and are only slightly
dehydrated at anthesis. The pollen viability decreases sharply after dispersal. In
Chamaerops humilis, Malus domestica, Mercurialis annua, Prunus avium, Ricinus
communis, Trachycarpus excelsa and Typha latifolia, pollen dehydrates before
anthesis and is often of small or medium size. This type of pollen is characterized
by long duration of pollen survival after dispersal. It contains high concentrations of
soluble sugars and cytoplasmic polysaccharides with little or no starch. In Lilium,
Magnolia grandiflora, Passiflora caerulea and Pinus halepensis, pollen grains

contain intermediate quantities of starch, cytoplasmic polysaccharides and soluble
sugars, which are associated with intermediate longevity (Speranza et al. 1997).
In addition to the lipidic matrix of pollen walls and intracellular membrane
systems, large amounts of oil bodies are also found in the cytoplasm of pollen
grains in some species, such as Brassica napus (Charzynska et al. 1989), Gossy-
pium hirsutum (Wetzel and Jensen 1992) and Arabidopsis thaliana (Kuang and
Musgrave 1996). In most cases, lipid bodies are present in the cytoplasm of
vegetative cells. However, lipid bodies have also been observed in generative
cells of lily pollen or only in the generative cell cytoplasm in Polystachia pub-
escens. The distribution and appearance of lipid bodies have been recognized as
cytological markers for the monitoring of pollen cell fate (Park and Twell 2001).
Lipids are reserves in pollen in a manner that is analogous to the accumulation of
storage oil bodies in many seeds. Both the membrane and storage lipids of pollen
grains provide the substrates and energy reserves for pollen germination and
subsequent elongation of the pollen tube. In Arabidopsis, considerable amounts
of lipid droplets are present throughout the vegetative cytoplasm in mature pollen
grains (Kuang and Musgrave 1996).
13.2.2.3 Transcriptome and Proteome of Pollen Grains
Pollen grains of most species are metabolically quiescent. However, they can
respond rapidly upon pollination by interacting with stigma cells, and germinating
pollen tubes undergo rapid polar growth. Undoubtedly, the readiness of pollen
grains depends on the “pre-stored” substances, especially transcripts and proteins.
Recent molecular analysis on mature pollen grains of representative species, in-
cluding Arabidopsis, maize, lily, tobacco and Plumbago zelanica, indicated that
mature pollen expresses a comparatively reduced and unique set of genes that are
tightly associated with their main task of delivering sperm to the embryo sac
(Honys and Twell 2003; Lee and Lee 2003; Becker et al. 2003; da Costa-Nunes
and Grossniklaus 2004). Transcriptome profiles of Arabidopsis pollen show that,
although the total number of expressed genes decline as male gametogenesis
progresses, the expression of genes that function in the cytoskeleton, vesicle
trafficking, cell wall biosynthesis and regulation, ion dynamics, signal transduction
and other aspects related to pollen germination and tube growth is up-regulated
significantly. For example, expression of gene clusters related to cell proliferation
declines as pollen matures, while genes involved in pollen maturation or even post-
pollination events increase. Consistently, the repression of constitutive genes and
activation of male gametophyte-specific genes associated with specialization of
mature pollen in preparation for pollen germination and pollen tube growth are
observed in pollen transcriptomes. Furthermore, genes involved in small RNA
biogenesis are down-regulated in Arabidopsis pollen. Fifteen genes of small RNA
pathways are absent in mature pollen, although they are active in vegetative tissues.
These findings indicate that small RNA pathways may be inactivated in mature
pollen of Arabidopsis. The observations also support the hypothesis that the male
gametophyte reduces the complexity of its transcriptome, compared with the
sporophyte, and maintains a limited amount of transcripts essential for pollen
germination and the later delivery of sperm (Honys and Twell 2003; Becker et al.
2003; Pina et al. 2005).
Proteome profiles of rice pollen indicated that mature pollen grains pre-synthesize
proteins needed for pollen function. These analyses showed that proteins involved
in cell wall metabolism, protein synthesis and degradation, cytoskeleton dynamics
and carbohydrate/energy metabolism are abundant in germinating rice pollen. In
addition, multiple protein isoforms are present in mature pollen, suggesting that
posttranslational modifications are common in mature pollen. Since transcripts of
genes involved in transcription and protein synthesis are in low abundance in
mature pollen grains, as shown by transcriptome profiles, it is apparent that a
large amount of proteins are stored (Dai et al. 2006, 2007). These results suggest
that the stored mRNAs and proteins are critical for subsequent pollen activity.
13.3 Pollen-Stigma Interaction
The pistil is a female sexual organ, an elongated structure located in a central

position in a flower. A fully developed pistil is composed of three parts, namely the
uppermost stigma, basal ovary that contains ovules and a style that connects the
stigma and ovary. Pollen grains released from the anther are deposited on the stigma
surface either directly in selfing plants or indirectly by wind or insects in cross-
pollinating plants. After germination at the compatible stigma, the pollen tube
subsequently penetrates through the stylar transmitting tissues of the ovary, exits
the placenta and travels along the funicular surface. It finally enters an ovule and
then the embryo sac, in which the pollen tube tip bursts in the synergid to release the
two sperm cells for double fertilization. The nonmotile sperms are delivered
directly to the embryo sac by the pollen tube, which allows flowering plants to
carry out sexual reproduction on land without the need for water (Hiscock and
Allen 2008).
13.3.1 The Stigma
The stigma surface is the place where pollen grains adhere, hydrate and germinate.
In flowering plants, the interaction between pollen and stigma is the first event that
initiates the process of fertilization. Successful pollen-stigma communication and
coordination are essential to the subsequent processes of pollination. Stigmas are
classified as wet or dry, according to the amount of matrices secreted by the
specialized papillar cells of the stigma surface. For example, grasses such as rice,
and crucifers such as Brassica species and Arabidopsis possess dry stigmas with
an epidermis of many large, hairy papillae cells interacting directly with pollen
grains, to accept compatible pollen and reject pollen that is incompatible or from
foreign species. The surface of papillar cells is covered with an interrupted layer
of waxy cuticle overlaid with a distinct proteinaceous pellicle of poorly defined
molecular composition (Ciampolini et al. 2001; Hiscock et al. 2002). However,
wet stigmas often contain three distinct zones, namely an epidermis with papillae,
a subepidermal secretory zone and a parenchymatous ground tissue. Legumes and
members of the Orchidaceae and Solanaceae families are typical wet stigma-type
plants. Most exudates are produced by the cells of the secretory zone, and less
secretion is derived from the stigmatic papillae of wet stigmas at pistil maturity
(Kandasamy and Kristen 1987). Moreover, pollen and stigma structures are
largely coevoluted, since plants of trinucleate pollen species usually have dry
stigmas, while the binucleate pollen species are concomitant with wet stigmas
(Edlund et al. 2004).
Transcriptome profiling of rice stigma indicates that stigma-specific genes share
conserved roles in plants. The majority of genes that are specifically or preferen-
tially expressed in stigmas of Arabidopsis and rice belong to the cell wall-related
and signal transduction groups, indicating that these two classes of genes play
conserved roles in the stigma. Other highly expressed genes in the rice stigma are
related with auxin signalling pathways, transport functions and stress responses (Li
et al. 2007). In Arabidopsis, stigma-specific genes have functions related to the
development of the stigma epidermis, in pollen recognition, or in the promotion of
adhesion, hydration and germination of pollen grains (Tung et al. 2005).
13.3.2 Pollen Recognition
The pollen coat, as the first barrier for recognition, contains molecules involved in
the initial interaction with the stigma (Mayfield et al. 2001). Proteins and lipids in
the pollen coat act as ligands for receptors in the stigma, as seen in the self-
incompatibility (SI) system exemplified in Brassica. The interaction between the
pollen surface and the stigma differs from species to species, due to diversities in
morphology and composition of the pollen coat and in the extracellular matrix
(ECM) of the stigma. The interactions between pollen and dry stigmas are highly
regulated and usually contain multiple steps of recognition, adhesion, rehydration
and germination. The dry stigma of some species, such as crucifers, constitutes an
important barrier for the rehydration of pollen grains. This barrier serves as an early
discrimination for the male-female recognition system during pollination. To date,
the best-characterized pollen-pistil interaction is the SI responses on dry stigmas in
Brassica and on wet stigmas in Antirrhinum and petunia of the Scrophulariaceae.
In the Brassica SI system, the male determinant, a pollen-secreted small cysteine-
rich (SCR) protein, interacts with the female determinants, a cell wall S-locus
glycoprotein (SLG) and plasma membrane receptor kinase (SRK) of the stigma
epidermal cells, to initiate an SI reaction (Schopfer et al. 1999; Nasrallah 2000). In

contrast, in the wet stigmas, a wide range of proteins, and the lipidic exudate or
carbohydrate-rich material that is produced by the cells of the secretory zone are
secreted at pistil maturity. Also, a thin layer of water in crystal form surrounds the
cells of the secretory zone underneath the exudates. Thus, the pollen grains on the
stigma establish direct contact with the pistil through the exudates, while the
hydration of pollen is facilitated by the carbohydrate- and lipid-rich secretions.
Stigma secretion also plays a key role in pollen capture and adhesion. As a result,
pollen is trapped rapidly and immersed within the secretion when contacting a wet
stigma. It appears that the first interaction of pollen-stigma in wet stigmas is passive
and indiscriminate (Swanson et al. 2004). Therefore, the inhibition of incompatible
pollen may occur at later stages.
13.3.3 Pollen Adherence and Hydration
The pollen-stigma signalling machinery is critical for pollen recognition, and cell
adhesion is likely required to stabilize pollen-stigma epidermal contacts. Adhesion
of pollen grains onto the stigma surface is a key step, especially for plants with dry
stigmas. Adhesion occurs prior to pollen hydration and germination, while the
recognition may exist in one or through all the phases of adhesion, rehydration,
pollen germination or even tube growth.
In Arabidopsis, pollen-stigma adhesion is rapid and highly selective. The bind-
ing force between pollen grains and stigma cells averages about 5.010–7 N. If, by
analogy, this adhesive is spread onto an area of 0.1–0.5 m2, it would be sufficiently
strong to suspend a 100-kg object. Furthermore, the magnitude of pollen adhesion
increases during pollination, and rapid remodelling of the stigma surface occurs
simultaneously on adhesion. The stigma surface in contact with pollen grains
protrudes into the space and the cell wall thickness changes. However, initial
pollen-stigma adhesion is independent of the pollen coat. The adhesion molecules
may exist in the exine, since the purified exine alone can bind to the stigma with
high affinity (Zinkl et al. 1999).
After landing on the stigmatic surface of the pistil, the dehydrated pollen grains
rehydrate, mobilize water from the papillar surface, activate their metabolism, and
then germinate if the pistil is receptive. Adhesion and hydration are also complex
and highly regulated in species with dry stigmas (Preuss et al. 1993; Dickinson
1995). When pollen grains fall on a compatible stigma, the exine coating changes
dramatically in response to contact with the stigma surface and, subsequently, the
coating flows out to form a “foot” to glue the pollen to the stigma (Elleman and
Dickinson 1986). The “foot” differs from other parts of the pollen coat in some
physical and chemical properties, particularly its insolubility in cyclohexane
(Elleman et al. 1989). Furthermore, the hydration is dependent on pollen-pistil
recognition and interaction. Compatible and incompatible pollen grains behave
differently if they are present on the same stigmatic papilla of Brassica. The
compatible grains hydrate within 90 min, while the incompatible grains swell
slightly by retaining their fusiform shape for up to 24 h. Only the sole hydration of
the grain adjacent to the stigmatic papilla takes place whether the pollen grains are
present on the stigma in chains or the “compound” pollinations are made with grains
from other closely related species (Sarker et al. 1988). This implies that hydration
occurs only between stigmatic papilla and pollen, but it cannot be passed amongst
pollen grains. Post-pollination events occur rapidly in Arabidopsis. Apparent cyto-
logical changes take place within 5 min after pollen alights on the stigma. The
hydration and regain of cytoplasm polarity in pollen grains and emergence of pollen
tubes occur within 15 min after pollination. At about 20 min after pollination, pollen
tubes can be visibly detected invading the expanded papillar cell wall and, after
40–50 min, pollen tubes have digested their way through both the separated outer
and inner layers of the papillar cell wall and have reached the intercellular matrix
separating the papillar cells from the subepidermal cells of the stigma (Kandasamy
et al. 1994). However, in species with wet stigmas, the hydration of pollen grains
is passive and autonomous. The stigma exudates are essential for pollen hydration,
as they provide pollen grains with a medium containing not only water but also
other materials for metabolism. Consistently, mutations eliminating exudate
formation from stigmas result in female sterility in tobacco (Goldman et al. 1994;
Wolters-Arts et al. 1998).
To address the signals controlling pollen adhesion and hydration, recent studies
in Arabidopsis suggested that lipids might be the key signalling molecule. When
triacylglycerides are added, pollen germination is restored on stigma-less pistils and
the surface of other vegetative tissues, including leaves (Wolters-Arts et al. 1998).
It has been reported that different lipids define the timing of pollen hydration,
suggesting that lipids may mediate the permeability of the pollen grain surface to
water (Wolters-Arts et al. 2002). Studies on eceriferum (cer) mutants of Arabidopsis
confirmed the importance of pollen coat lipids for pollen hydration (Preuss et al.
1993; Fiebig et al. 2000). In Arabidopsis, CER genes form a large family involved in
the biosynthesis of long-chain lipids that play a role in forming a hydrophobic
waxy cuticle critical for resistance to water loss, pathogen attack or UV irradiation
(Post-Beittenmiller 1996). All cer mutants (cer1, cer2, cer3, cer6/pop1, cer8 and
cer10) are unable to synthesize long-chain lipids and, consequently, result in male
sterility due to lack of long-chain lipids in the pollen coat (Koornneef et al. 1989;
Preuss et al. 1993; Hannoufa et al. 1996; Millar et al. 1999; Fiebig et al. 2000).
Further studies show that the lack of lipid in the pollen coat interferes with pollen
hydration but not adhesion. Nevertheless, the male fertility of cer mutants can be
restored with high humidity, exogenous application of various triacylglycerides,
mineral oil and a nonspecific alkane (Hernandez-Pinzon et al. 1999). Similarly,
mutation in FACELESS POLLEN-1 (FTP1), a gene encoding an aldehyde decarbo-
nylase involved in wax biosynthesis, resulted in a phenotype similar to cer mutations
(Ariizumi et al. 2003). These results suggest that lipid of the pollen coat is vital for
pollen hydration.
Apart from pollen coat lipids, proteins in the pollen coat are also critical for
pollen hydration. Knockout of the GRP17 gene, which encodes an oleosin-like
protein present in the pollen coat, causes the delay of pollen hydration and substan-
tial reduction of pollen fitness. This implies that GRP17 plays a role in pollen
hydration (Mayfield and Preuss 2000; Mayfield et al. 2001). Other proteins, such as
aquaporins, also play an important role in regulating pollen hydration. Aquaporins
are a class of proteins that specifically facilitate the passive movement of water and/
or small neutral solutes (e.g. urea, boric acid and silicic acid) or gases (e.g.
ammonia and carbon dioxide) through membranes (Maurel et al. 2008). It has
been reported that an aquaporin-like plasma membrane protein encoded by MIP-
MOP (major intrinsic protein associated with the MOD locus) is abundant in
papillar cells of the stigma, and may act as a major regulator of hydration by
controlling water flow from the stigma to the pollen grains (Dixit et al. 2001).
During pollen hydration, the papillar cell is thought to actively transport water,
calcium, boron and other components essential for pollen germination into the
pollen grains. As pollen grains hydrate, extracellular Ca2+ flows into the grains.
This influx appears to trigger the rapid activation of the stored RNA, protein
and bioactive small molecules that allow rapid germination and tube growth
(Mascarenhas 1993).
13.4 Pollen Germination and Tube Growth
Hydration causes changes in the water content and volume of the pollen grain,
which may act as the initial signal to trigger pollen germination. As hydration
occurs, Ca2+ influx takes place and initiates the reorganization of the cytoskeleton
and also polarizes the cytoplasm of the vegetative cell in the pollen grain. Subse-
quently, cytoplasmic polarity of the vegetative cell is established, as manifested by
local accumulation of vesicles and a cytoplasmic gradient of Ca2+ beneath the pore
near the site of adhesion (Kandasamy et al. 1994; Franklin-Tong 1999a, b).
Consequently, the vegetative cell germinates to produce a pollen tube. The aperture
is often the site where the pollen tube emerges. However, for inaperturated pollen
grains, the pollen tube may exit anywhere.
The pollen tube wall is an extension of the intine, and tube elongation is the rapid
tip growth of a single cell growing directionally from one end. Establishment and
maintenance of the polarity of the tube are important to attain rapid polar tip
growth. Consistently, gene products involved in cell rescue, transcription, subcel-
lular localization, metabolism, proteins with binding function, and cellular transport
are overrepresented during pollen germination, except for the “pre-stored” tran-
scripts and proteins in pollen. Similarly, these genes are also up-regulated during
pollen tube growth (Wang et al. 2008). Also, it has been well documented that
calcium, the cytoskeleton, small GTPases and other factors regulating polar growth
are critical for pollen germination and tube growth.
13.4.1 Calcium Signalling in Pollen Germination

and Tube Growth
As a universal signalling molecule, Ca2+ is essential for pollen tube growth. The
role of calcium has been demonstrated in several studies. Firstly, accumulation of
cytoplasmic Ca2+ ([Ca2+]cyt) at the germinal aperture, where the pollen tube
emerges, occurs soon after hydration and calcium influx takes place at the pollen
tube tip (Franklin-Tong et al. 2002; Lazzaro et al. 2005; Bushart and Roux 2007).
Secondly, if no [Ca2+]cyt gradient is established in the pollen grains, there is no
protrusion following hydration, and germination is inhibited. Inhibition of calcium
channels can also block pollen germination in vitro (Franklin-Tong et al. 2002;
Wang et al. 2004; Bushart and Roux 2007). Thirdly, pollen tube growth is arrested
when Ca2+ uptake is inhibited (Jaffe et al. 1975; Bednarska 1989; Franklin-Tong
1999a). The germination and reorientation of pollen tubes have been shown to
parallel calcium concentration. Disruption or modification of the Ca2+ gradient at
the tube apex interrupts tube growth (Miller et al. 1992) or induces bending of the
growth axis towards the zone of higher [Ca2+]cyt (Malhó and Trewavas 1996).
These findings clearly show that calcium is essential for pollen germination and
tube growth in vitro. Interestingly, concomitant with a [Ca2+]cyt increase in the
pollen grain, a [Ca2+]cyt gradient of the papillar cell can also be induced by
pollination (Iwano et al. 2004), suggesting that interaction between pollen
grain and papillar cells during pollination is mediated through calcium-dependent
signalling.
How does Ca2+ exert its control over tip growth? The regulation of [Ca2+]cyt
dynamics is a combined effect of ion pumps, antiporters and uniporters. The
dynamics of the Ca2+ signal is controlled by influx (through channels) and efflux
(through pumps and antiporters; Schiøtt et al. 2004). Influx of Ca2+ is limited to a
small region at the tube apex (Pierson et al. 1996; Holdaway-Clarke et al. 1997),
and this tip-localized entry appears largely responsible for the formation of the Ca2+
gradient. Also, Ca2+ signalling is changed when efflux is blocked by the disruption
of the Ca2+ pump or by antiporters (Schiøtt et al. 2004). Besides the formation of a
calcium gradient along the growing tip, [Ca2+]cyt oscillation may likely play a role,
as fluctuations in [Ca2+]cyt have been observed to correlate with pulses in tube
growth (Holdaway-Clarke and Hepler 2003; Malhó et al. 2006). It has also been
reported that [Ca2+]cyt ranges from 2 to 10 mM at the apex, decreasing sharply to 20
to 200 nM within 20 mm down the apex (Obermeyer and Weisenseel 1991; Rathore
et al. 1991; Miller et al. 1992; Malhó et al. 1994; Pierson et al. 1994; Franklin-Tong
et al. 1997). Therefore, it has been speculated that both the concentration gradient
and oscillation of [Ca2+]cyt are important signals, which may function via the Ca2+
sensor calmodulin (CaM) and its downstream effectors, such as CaM-binding
proteins in the pollen tube. In Arabidopsis, mutations in NO POLLEN GERMINA-
TION1 (NPG1) and NPG1-related genes, coding for CaM-binding proteins, abolish
pollen germination (Golovkin and Reddy 2003), confirming a role of Ca2+ signal-
ling in pollen germination.
How the Ca2+ signalling leads to polar tube growth remains to be elucidated,
although a large number of proteins involved in Ca2+ signalling have been detected
in the pollen transcriptome (Honys and Twell 2004; Pina et al. 2005; Becker and
Feijó 2007). Recent studies showed that calcium-dependent protein kinases
(CDPKs), such as the calmodulin-like domain containing protein kinases, are likely
the candidate modulators of Ca2+ signalling (Yoon et al. 2006). Pollen-specific
CDPKs have been identified from maize and petunia, and they may serve as Ca2+
regulators in pollen tube growth (Estruch et al. 1994; Moutinho et al. 1998; Yoon
et al. 2006). Inhibition of CDPK activity with kinase inhibitors or calmodulin
(CaM) antagonists has been shown to impair pollen germination and tube growth.
CaM has been considered as a primary sensor of the Ca2+ signal regulating the
activity of many proteins, such as enzymes, cytoskeletal and structural proteins, via
its Ca2+-binding capacity. Consistent with the polar distribution of Ca2+, CaM
accumulates at the germinal aperture, and a tip-base CaM gradient exists in
elongating pollen tubes (Hauber et al. 1984; Tirlapur et al. 1994). In tobacco,
CaM accumulates in the cytoplasm close to the three germinal apertures of the
pollen grains, particularly near the aperture from which the pollen tube emerges.
Furthermore, directional migration of CaM has also been observed during pollen
hydration and germination, suggesting that the dynamic patterns of CaM distribu-
tion may play a crucial role in the establishment of polarity during pollen germina-
tion (Tao et al. 2004). In the growing pollen tube, the binding capacity of CaM
appears more intense in the subapical region, indicating a higher concentration of
CaM targeting molecules, such as cytoskeletal elements, in this region. Microfila-
ments resembling the V-shaped collar reported for CaM binding are present at
the growing tip (Miller et al. 1996; Moutinho et al. 1998), suggesting a close link
between the actin cytoskeleton and calcium signalling during pollen germination
and tube growth. In addition, CaM may crosstalk with the cAMP signalling
pathway, as CaM binds cyclic nucleotide-gated channels (CNGC), the mutation
of which affects polarized tip growth of pollen tubes in Arabidopsis (Frietsch
et al. 2007). Expression of CNGC18 in Escherchia coli resulted in a time- and
concentration-dependent Ca2+ accumulation, indicating that CNGC may transduce
the cyclic nucleotide signal into a Ca2+ influx, thereby regulating polar tip growth.
In yeast and animals, it has been well established that CaM controls the actin
cytoskeleton via regulation of phosophatidylinositol 4,5-bis-phosphate (PIP3) syn-
thesis (Desrivieres et al. 2002), but a similar link in plants remains to be elucidated.
13.4.2 The Cytoskeleton
Cytoskeleton dynamics regulate many important cellular processes, such as organ-

elle movement, chromosome segregation, flagellar movement and pollen tube
elongation. A hallmark pollen tube feature is its cytoskeleton, which comprises
actin filaments and microtubules (MTs). Microfilaments (MFs) and MTs have long
been recognized as important structures for pollen germination and tube growth.
However, the ability to dissect the distribution and dynamics of the cytoskeleton in
pollen tubes growth either in vivo or in vitro became feasible only after the
availability of immunocytochemical and in vivo GFP-labelling methods in plant
research. Based on cytoskeleton organization, the pollen tube can be divided into
three main regions, namely the shank, subapical and apical regions. Observations of
chemically fixed and living pollen tubes reveal the elaborate cytoskeleton consti-
tuting an extensive matrix of actin and microtubule bundles along the shank of the
tube, reaching the subapical region, but hardly at the apical dome (Kost et al. 1998;
Fu et al. 2001; Chen et al. 2002; Lovy-Wheeler et al. 2005; Cheung and Wu 2008).
As a distinct structural and functional domain in elongating pollen tubes, the
subapical region is characterized as a network of interdigitating actin cables, and
a dramatic dynamics of organelles is associated with this actin mesh in this region
(Chen et al. 2002; Cheung and Wu 2004). Although mitochondria and Golgi bodies
have been observed throughout the pollen tube, the highest concentration is found
in the subapical region. Tubular endoplasmic reticulum is concentrated mostly in
the apical region, whereas rough endoplasmic reticulum is abundant, starting
behind the inverted cone region and throughout the rest of the cytoplasm (Cheung
and Wu 2007). The apical cytoplasm of the tube is the so-called clear zone,
which lacks light-reflecting organelles (Steer and Steer 1989; Pierson et al. 1990).
This zone is packed with secretory vesicles and recycled endocytosed membrane
(Camacho and Malhó 2003; Parton et al. 2003). Many of the vesicles in the clear
zone fuse with the apical membrane, thereby adding new membrane, membrane
proteins, cell wall materials, secretory proteins and other signal molecules for the
rapid growth of the tube (Luu et al. 2000; de Graaf et al. 2005).
13.4.2.1 Actin Cytoskeleton
As adaptive to the tip-growing character of the pollen tube, the cytoskeleton of the
polar cell develops special features for rapid polar growth and the delivery of sperm
cells in the tube, including specific cytoskeleton-binding proteins and polar config-
uration. The polarization of actin filaments and microtubules are initiated as
hydration and germination of pollen grains occur. Subsequently, MFs and MTs
are organized as longer bundles and enter the emerging tube (Tiwari and Polito
1990). In the elongating pollen tubes, actin filaments and MTs are structured in
bundles that extend along the longitudinal axis reaching the subapical region. These
occur both in the cortical and central cytoplasm, as well as in the apical dome of the
pollen tube. A dynamic form of tip-localized short actin bundles (SABs) and a
striking cortical fringe of F-actin have been observed in tobacco and lily pollen
tubes with the aid of GFP-tagged actin-binding domain of mouse talin (Fu et al.
2001; Lovy-Wheeler et al. 2005). In lily pollen tubes, the actin fringe starts a few
microns back from the tip and extends basally for an additional 5 to 10 mm. It seems
that this fringe is fragile and difficult to preserve. However, the key structure is
likely to perform rapid remodelling and polymerization and, subsequently, plays a
central role in establishing cell/cytoplasmic polarity and in controlling rapid,
oscillatory growth in lily pollen tubes (Cárdenas et al. 2008). Consistently, the
dynamics of actin both in the apical and subapical region correlates closely with the
tip growth of pollen tubes (Cheung and Wu 2004). The prevention of actin
polymerization with low concentrations of latrunculin B or enhancing actin poly-
merization profoundly affects pollen tube growth (Cárdenas et al. 2005; T. Chen
et al. 2007). Furthermore, the strictly regulated actin dynamics and the level of
nascent actin filament production seem critical to maintain tip growth (Vidali et al.
2001; Chen et al. 2002; Cheung and Wu 2004).
In tip-growing cells, such as pollen tubes and root hairs, cytoplasmic streaming,
a process dependent on the actin cytoskeleton, is critical for rapid growth. For
example, cultured tobacco pollen tubes possess a diameter of 10 mm, but the tubes
can elongate at an average rate of 5 mm min–1 and grow to a length of up to 15 mm,
which is 1,500 times their diameter (Parton et al. 2003; Kost 2008). The rapid tip
growth depends on a continuous supply of secretory vesicles containing cell wall
material, which fuse with the plasma membrane specifically at the apex, manifested
by a constant flow of organelles towards the apex and vice versa (Derksen et al.
1995; Campanoni and Blatt 2007). A bidirectional, “reverse fountain” pattern of
cytoplasmic streaming is a hallmark of growing pollen tubes in angiosperms.
During the streaming, the forward lanes flow along the edge of the cell but, in the
apex, they undergo a reversal in direction, and stream basally through the pollen
tube interior. The cytoplasmic streaming allows the transport of cargo particles or
secretory vesicles, such as Golgi vesicles, carrying pectin and many other cell wall
components to the rapidly growing cell (Cai et al. 1997; Franklin-Tong 1999a). The
transport is thought to be driven by myosin on the vesicle surface that moves along
the actin microfilaments (Miller et al. 1995; Yokota et al. 1995). Actin MFs in the
shank are organized into bundles with uniform polarity and serve as tracks for the
tip-ward streaming. However, in the subapical region, F-actins are less organized
into fine filament bundles and form a collar-like zone that contributes to the
reversed streaming and, consequently, gives rise to a reverse-fountain pattern
(Hepler et al. 2001; Li et al. 2001; Ren and Xiang 2007). The actin MFs also
play a critical role in the reversal of streaming in the pollen tube apex (Cárdenas
et al. 2005).
The dynamics of the actin cytoskeleton is tightly regulated in growing pollen
tubes, especially in the apical and subapical regions. The formation of F-actin
arrays depends on the biochemical interactions of actin monomers and actin-
binding proteins (ABPs). The elaborate architecture of the actin cytoskeleton is a
result of the fine spatial and temporal incorporation of ABPs. Therefore, ABPs
regulate the depolymerization and polymerization of actin filaments, equilibrate the
globular actin (G-actin) and F-actin pools, and perceive or transduce various signals
that affect the cytoskeleton, such as Ca2+, pH, phosphoinositides and phosphoryla-
tion. ABPs are divided into several groups according to their function and structure.
These include profilin, actin-depolymerizing factor (ADF) or cofilin, villin, gelsolin
or fragmin, actin nucleators and heterodimeric capping proteins (Hussey et al.
2006; Ren and Xiang 2007; Xiang et al. 2007; Cheung and Wu 2008; Qualmann
and Kessels 2008). Profilin is distributed throughout the pollen tube cytoplasm.
It binds monomeric (G-) actin, modulates actin nucleation and enhances actin
polymerization. Subject to the high Ca2+ concentrations in the apical region,
more actin-profilin complexes may form at the tip (Kovar et al. 2000; Vidali
et al. 2001; Snowman et al. 2002). ADF/cofilin binds to both the monomeric and
F-actin to enhance actin depolymerization by increasing the off-rate of actin
monomer from the pointed ends, and inducing filament severing (Chen 2002). Its
binding activity is regulated by phosphorylation (Allwood et al. 2001), pH and
phosphatidylinositol 4,5-bisphosphate (PIP2) or phosphatidylinositol 4-monopho-
sphate (PIP; Gungabissoon et al. 1998). ADF serves as actin filaments at high pH
(8.0) and binds to F-actin at a lower pH of 6.0 (Gungabissoon et al. 2001; Yeoh
et al. 2002). Villin has been shown to assemble actin filaments into bundles in a
unipolar manner. It is also one of the Ca2+-regulated ABPs, as it is negatively
regulated by a Ca2+-calmodulin complex (Yokota and Shimmen 2000). In the tube
apex, as the Ca2+ concentrations are greater than in other regions, the villin activity
is inhibited and actin bundles are not detected. This finding is consistent with the
results of previous studies showing that only a mesh of short actin filaments, but not
actin bundles, is found in the pollen tube apex (Miller et al. 1996; Kost et al. 1998;
Yokota and Shimmen 2000). Gelsolin, fragmin and severin are members of the
gelsolin family that perform severing, capping and nucleating activity in a Ca2+-
dependent fashion (Staiger and Hussey 2004). Gelsolin-like proteins have been
identified in maize, lily and poppy pollen. In poppy pollen, PrABP80, a Ca2+-
regulated gelsolin-like ABP, can nucleate actin polymerization from monomers and
block the assembly of the profilin-actin complex onto actin filament ends, thereby
enhancing profilin-mediated actin depolymerization (Huang et al. 2004). It has also
been observed that the Arabidopsis heterodimeric capping protein (AtCP) prevents
the addition of profilin actin to barbed ends, being sensitive to phosphatidylinositol
4,5-bisphosphate. In the presence of phosphatidic acid (PA), AtCP is unable to block
the barbed or rapidly growing and shrinking end of actin filaments. Inhibition of CP
activity in cells by elevated PA results in the stimulation of actin polymerization
from a large pool of profilin-actin, suggesting that capping proteins may imply a link
to lipid-regulated polar cell growth (Huang et al. 2006). The roles of ABPs in pollen
tube growth require further investigation.
13.4.2.2 Microtubule Cytoskeleton
While the role of the MT cytoskeleton in pollen tube growth is controversial, it has
been reported that MT-depolymerizing chemicals (such as colchicine) do not
inhibit organelle movement in the pollen tube (Heslop-Harrison et al. 1988).
However, MT perturbation has been shown to cause the loss of cytoplasmic
organization (Joos et al. 1994), and the pulsed growth rate of tobacco pollen
tubes is modified when MT is depolymerized (Geitmann et al. 1995). The ability
to maintain the growth direction of the pollen tube can be affected by MT degrada-
tion, while the integrity of the MT cytoskeleton depends on the presence of actin
filaments. However, the actin filaments appear to be independent of the MT
configuration (Gossot and Geitmann 2007). Both the MT and the actin cytoskeleton
are involved in organelle movement. The mitochondria and the Golgi apparatus
have been shown to move slower along MTs than is the case for actin MFs
(Romagnoli et al. 2007), indicating that MTs play a role in the positioning of
organelles. It has been proposed that MTs mediate the migration of the male
germ unit and the trafficking of vesicles and organelles either independently
(Astrom et al. 1995; Romagnoli et al. 2003) or through interaction with actin
filaments (Cai et al. 1997, 2000; Gossot and Geitmann 2007). Recently, the role
of MTs in polar growth was demonstrated in fission yeast, Schizosaccharomyces
pombe. MTs were shown to mediate the recruitment or accumulation of polar
factors to the growing tip, leading to a polarity alteration of cell growth in the
physically bent fission yeast cells (Minc et al. 2009). It remains to be investigated
whether MTs have a similar role in pollen tube growth.
13.4.3 Crosstalk Between Calcium Signalling and Cytoskeleton

in the Pollen Tube
The polar growth of the pollen tube requires the continuous delivery of new
materials carried by secretory vesicles to the apex of the tube, while pollen tubes
exhibit oscillatory growth. In Arabidopsis pollen tubes, the pollen tube growth rate
increases following the actin-dependent and oscillatory motion of the endoplasmic
reticulum (ER). It has been well documented that actin dynamics underlies the
mechanism that initiates the surge in growth (Fu et al. 2001; Lovy-Wheeler et al.
2007). As discussed above, the configuration of actin filaments is controlled by a
variety of ABPs, most of which are regulated by the Ca2+ signal. Ca2+ is a central
factor controlling the transition from G-actin (or short actin mesh) in the tube apex
to the F-actin cables in the shank. Furthermore, Ca2+ controls the fusion of vesicles
and dynamics of actin filaments through its diverse activities and interactions with
ABPs involved in actin bundling, severing, capping and nucleating. It has been
reported that myosin-mediated organelle movement along actin bundles is inhibited
by high Ca2+ concentrations at the apex, facilitated in the base region of the pollen
tube by lower Ca2+ concentrations. Besides the regulation of ABPs by Ca2+ con-
centrations, Ca2+ channels, together with channel-mediated Ca2+ influxes across the
plasma membrane of pollen and pollen tubes, are closely associated with actin MFs.
In Arabidopsis, Ca2+ channel blockers, La3+ and Gd3+, and F-actin depolymeriza-
tion reagents significantly inhibit pollen germination and tube growth (Wang et al.
2004). The inhibition of pollen germination and tube growth by cytochalasins is
enhanced by an increase in external Ca2+. In contrast, Lat-B inhibition on pollen
tube growth is coupled closely with a decline in the apical Ca2+ gradient (Cárdenas
et al. 2008). Similarly, pollen tubes exhibiting non-oscillating growth display a
reduced and non-oscillating Ca2+ gradient. This reflects either the different modes
of action amongst the inhibitors or the pleiotropic effects of Lat-B. Lat-B can cause
a number of reversible phenotypes, including elimination of the acidic domain at

the extreme tube apex, a shift of the alkaline band towards the tip, degradation of
the cortical fringe, disorganization of the microfilaments in the shank, and oblitera-
tion of the clear zone with the organelles invading into and through the extreme
apex of the tube. These results suggest that there may be crosstalk between the
actin cytoskeleton and Ca2+ signalling in controlling pollen tube growth (Cárdenas
et al. 2008).
13.4.4 Small GTPases and Pollen Tube Growth
Small GTP-binding proteins are molecular switches that can cycle intrinsically
between an “active” GTP-bound form and an “inactive” GDP-bound form, depend-
ing on their substrate. This reversible reaction is modulated by proteins that affect
the rate of either GTP hydrolysis or GDP release, thereby operating the switch.
These small G proteins represent a conserved signalling pathway in eukaryotes, and
control a variety of cellular processes, including polar cell growth. Guanine nucle-
otide exchange factors (GEFs) activate membrane-associated GTPases by exchang-
ing GDP with GTP, while GTPase-activating proteins (GAPs) promote GTP
hydrolysis to inactivate GTPases. Guanine nucleotide-dissociation inhibitors
(GDIs) have been shown to inhibit the activation of GTPases by suppressing
nucleotide exchange and sequestering GTPases into the cytosol. The small GTPases
in the Ras-like small GTPase superfamily are classified into five distinct families,
according to their structure and function. These are Ras, Rab, Rho, Arf and Ran.
Ras GTPases regulate cell proliferation in yeast and mammalian systems, but they
are evolutionally lost from some lineages, including plants. Rho GTPases are
involved in actin dynamics and signal transduction pathways involving MAP
kinases (Vernoud et al. 2003). Plants have a novel and unique family of Rho-like
GTPases (ROPs), represented by a large number of proteins (Zheng and Yang 2000;
Valster et al. 2000). Since plant ROPs are similar to animal Rac proteins in primary
sequence, and interact with proteins bearing the “CRIB” (Cdc42/Rac-interactive
binding) domain, they are also referred to as RAC proteins, Rac-like GTPases or
Rac-Rop GTPases (Vernoud et al. 2003; Kost 2008). ROPs regulate actin dynamics
and modulate H2O2 production in polarly growing cells and defence responses
(Yang 2002; Morel et al. 2004). The Rab and Arf (ADP-ribosylation factors)
GTPase families participate in endosomal vesicle trafficking, and Ras-related
nuclear protein (Ran) GTPases function in nucleo-cytoplasmic transport of proteins
and RNAs (Vernoud et al. 2003). Small GTPases are often lipid-modified and
membrane-localized, with the exception of Ran GTPases (Molendijk et al. 2004).
Hence, among the proteins associated with GTPases, there are docking/scaffolding
proteins that target GTPases to specific membranes or membrane domains, or as
GDI-displacement factors (GDFs) to reinforce the re-association of GTPases with
the membrane (Fauré and Dagher 2001; DerMardirossian and Bokoch 2005).
Here we focus on the role of GTPases, especially Rab GTPases and ROPs, in pollen
tube growth in plants.
13.4.4.1 Rab GTPases
In pollen tubes, small GTPases have been implicated in at least two vital functions,
namely regulation of membrane vesicle trafficking and regulation of actin micro-
filaments (Krichevsky et al. 2007). Rab GTPases function crucially in membrane
fusion processes in the pollen tube (Molendijk et al. 2004; Cole and Fowler 2006).
As discussed above, rapid and polarized pollen tube growth demands an active
secretory system, which can provide sufficient material for the rapid elongation of
the tube. Hence, exocytosis responsible for the transport of secretory vesicles, small
signal molecules or enzymes is essential for pollen tube growth. Also, endocytosis
that may contribute to recycling and retrieval of the excess cellular material also
occurs at the growing tip of pollen tubes (Carroll et al. 1998; Krichevsky et al.
2007). There is increasing evidence showing that Rab GTPases play a pivotal role
in intracellular membrane trafficking, including endomembrane trafficking, exo/
endocytosis and membrane recycling (Surpin and Raikhel 2004; Molendijk et al.
2004). It is likely that Rab GTPases interact with SNARE proteins present in
organelle membranes and vesicles, in guiding these to their correct destiny and
providing specificity for membrane fusion (Stenmark and Olkkonen 2001; Zerial
and McBride 2001; Pfeffer and Aivazian 2004). Individual members of Rab
GTPases are localized to different cellular membrane compartments. These mem-
bers organize intracellular membrane trafficking at three consecutive stages, namely
membrane budding and vesicle formation, tethering vesicles to target areas and
promoting fusion of vesicles with target membranes (Tamm et al. 2003; Seabra and
Wasmeier 2004).
In gaining insight into the Rab GTPase function in pollen tube growth, genetic
approaches are greatly hampered by the overlapping functions of members of the
Rab family. Instead, a dominant-negative approach is employed. This approach
involves changes in Rab GTPases induced by introducing amino acid mutations
either stabilizing the GTP-bound activated state and up-regulating the regulatory
activity of Rab GTPases, referred to as constitutively active (CA), or locking the
GDP-bound inactivated form as dominant-negative (DN). Studies based on CA or
DN mutations of Rab GTPases in Arabidopsis and tobacco have revealed the
functions and regulatory pathways of Rab proteins, which are highly conserved
among yeast, animals and plants (Batoko et al. 2000; Grebe et al. 2003; Preuss et al.
2004; Ueda et al. 2004; de Graaf et al. 2005). A pollen-predominant Rab2 from
tobacco, NtRab2, has been shown to function in vesicle trafficking between the ER
and Golgi bodies, transporting cell membrane and secretory proteins. DN mutations
in NtRab2 inhibit the transport of pollen proteins entering the secretory pathway,
thereby suppressing pollen tube elongation. This implies that protein secretion in
pollen tubes relies on Rab2 GTPase that may function specifically in ER-to-Golgi
trafficking in highly secretory cell types (Cheung et al. 2002). Similarly, NtRab11b,
a member of the Rab11 subfamily of tobacco, is localized predominantly to the

transport vesicle-occupied apex of elongating pollen tubes, suggesting that it may
be involved in the apical accumulation of transport vesicles (Derksen et al. 1995;
Hepler et al. 2001). DN or CA expression of NtRab11b inhibits pollen tube growth,
disrupts growth directionality and causes male fertility. In addition, F-actin organi-
zation in the apical and subapical regions of pollen tubes is also affected when
NtRab11b activity is misregulated.
Unlike yeast and animals, only a few Rab GTPase effectors have been identified
in plants. A recent study has shown that PI-4 Kb1, an effector of RABA4b in
Arabidopsis, plays a role in the polarized expansion of root hair cells (Preuss et al.
2006). However, the role of PI-4 Kb1 in pollen tube growth remains to be eluci-
dated.
13.4.4.2 Rho-Like GTPases
In addition to Rabs, the Rho GTPase family has also been reported to be involved in
pollen tube growth. Rho GTPases are Ras-related small guanine nucleotide-binding
proteins that bind GTP and GDP with high affinity, but hydrolyze GTP inefficiently
(Yalovsky et al. 2008). Similarly to the GTPases of other families, the activity and
physiological status of Rhos are also modulated by the accessory binding proteins.
As key nodes of signalling networks and central players of multiple pathways, Rho
GTPases receive input signals from upstream regulators and then transduce to
downstream effectors, thereby orchestrating a wide range of intracellular signalling
networks. Like Rabs, Rho GTPases are also involved in regulating a variety of
cellular processes, such as cell polarity, endocytosis and vesicle trafficking (Berken
2006; Nibau et al. 2006; Yang and Fu 2007). The most fundamental effects of Rho
GTPases are on the actin cytoskeleton and dynamics. Rho signalling is often
restricted to a specific domain of the plasma membrane, where vesicle exocytosis
takes place (Fu et al. 2002; Gu et al. 2006; Hwang et al. 2008; Lee et al. 2008).
Rho family proteins in animals are functionally divided into five subfamilies,
namely the Rho-like, Rac-like, Cdc42-like, Rnd and RhoBTB subfamilies, which
have emerged as key regulators of the actin cytoskeleton and its dynamics
(Burridge and Wennerberg 2004). However, the ROP family is a single, unique
Rho-like GTPase from plants. The ROP family is known to regulate tip growth
and polar cell expansion in plant cells. In Arabidopsis, there are 11 different ROP
genes, seven of which are abundant or preferentially expressed in mature pollen
(Vernoud et al. 2003; Honys and Twell 2004; Pina et al. 2005). More ROPs and
their interacting proteins have been identified from results of studies on gain- or
loss-of-function mutants (Gu et al. 2004; Hwang and Yang 2006). In particular,
the pollen-specific AtROP1, which is localized to the plasma membrane in the
apical region of the tube, has been shown to be a central switch and essential
regulator of pollen tube tip growth. Blocking of ROP signalling by DN-Atrop1
mutation inhibits pollen tube elongation, whereas CA-Atrop1 mutation induced
isotropic growth (Kost et al. 1999; Li et al. 1999). The dynamics of short actin
bundles (SABs) in the apex of tobacco pollen tubes is also regulated by AtROP1.
Over-expression of AtROP1 causes depolarized growth of the pollen tube, which
can be rescued by over-expression of GDI, RopGAP1, or LatB treatment. Recently,
it was shown that REN1 (ROP ENHANCER 1), a Rho GAP that localized to the
apical cap and exocytic vesicles in the pollen tube tip, controls a negative-
feedback-based global inhibition of ROP1 and regulates the dynamics of ROP1
through the restriction of active ROP1 to the apical cap (Hwang et al. 2008). These
results provide direct evidence that Rho GTPase and actin organization are closely
linked in cell polarity and tip growth in plants (Fu et al. 2001). Similarly, over-
expression of a tobacco ROP/Rac protein, NtRac1, induces the actin cables to
extend to the apical region, and causes tremendous disorganization of actin cables
in the expanded tips (Chen et al. 2002). These findings suggest a central role of
ROP in pollen tube growth.
PtdIns 4,5-P2 is a downstream effector of ROP and a ligand for a large number
of regulatory proteins, including actin-binding proteins and proteins with pleck-
strin homology (PH-) or C2 domains. In tobacco pollen tubes, the ROP/RAC
GTPase interacts physically with phosphatidylinositol monophosphate kinase
(PtdIns P-K) and colocalizes with its product, PtdIns 4,5-P2, in the apical mem-
brane (Kost et al. 1999). PtdIns 4,5-P2 might function both downstream and
upstream of RAC/ROP activation. As an effector of Rac-Rop GTPases, PtdIns
4,5-P2 promotes the fusion of secretory vesicles with the plasma membrane; on the
other hand, PtdIns 4,5-P2 might function as a positive regulator to stimulate the
actin cytoskeleton and membrane trafficking by regulating the activity of ABPs
and calcium homeostasis during tip growth (Janmey 1994; Kost et al. 1999; Kost
2008). In addition, PtdIns 4,5-P2 is regulated by a pollen tube phosphoinositide-
specific phospholipase C (PLC), which maintains the polar distribution of PtdIns
4,5-P2 and Rho signalling. Studies on petunia PLC1 and tobacco NtPLC3 indicate
a major function of PLC activity in the regulation of PtdIns 4,5-P2 in the apical
plasma membrane of elongating pollen tubes. It is likely that PLC activity main-
tains the spatially restricted RAC/ROP signalling and polarized cell expansion by
preventing the lateral spreading of PtdIns 4,5-P2 (Dowd et al. 2006; Kost 2008).
Diacylglycerol (DAG) and the Ca2+-mobilizing messenger InsP3 are the two other
cellular regulators released by PLC-mediated hydrolysis of PtdIns 4,5-P2. While
InsP3 is involved in Ca2+ release from internal stores in plant cells, DAG is
detected specifically in the plasma membrane at the tip of elongating pollen
tubes (Helling et al. 2006). Furthermore, accumulation of DAG at the apex of
pollen tubes is prevented in the presence of PLC inhibitors, suggesting that DAG
accumulation may play an important role in RAC/ROP-dependent signalling, and
that it may be linked to membrane trafficking in growing pollen tubes. However,
evidence is needed to substantiate this, as signalling-dependent accumulation of
DAG has not been detected in plant cells other than tobacco pollen tubes (Helling
et al. 2006; Kost 2008).
ROP GTPases can transduce signals to downstream effectors, such as RICs
(ROP-interactive CRIB motif-containing proteins), to implement cellular processes
through divergent pathways. In pollen tubes, ROP GTPases can regulate actin
dynamics and tip growth via two counteracting downstream pathways. AtROP1
appears to operate downstream networks through two different RICs, RIC3 and
RIC4. RIC3 participates in Ca2+ regulation, whereas RIC4 modulates the actin
cytoskeleton. AtROP1 not only activates RIC3 to induce a high Ca2+ concentration
in the apical region and actin disassembly, but also promotes RIC4 to stimulate
actin polymerization and dynamics in the pollen tube (Gu et al. 2005; Lee et al.
2008). The apical accumulation of exocytotic vesicles oscillates in phase with, but
slightly behind, apical actin assembly, and is enhanced by overexpression of RIC4,
indicating a close link among RICs, exocytosis and actin dynamics in pollen tubes.
Furthermore, RIC4 overexpression inhibits exocytosis. This inhibition can be
suppressed by Lat-B or RIC3, implying that polar vesicle accumulation depends
on RIC4-mediated actin assembly, whereas the RIC3-dependent Ca2+ signal
reduces vesicle accumulation at the tip, resulting in actin disassembly (Lee et al.
2008). It is apparent that the ability of ROPs to integrate counteracting pathways
facilitates the fine tuning of actin dynamics during tip growth (Gu et al. 2005; Nibau
et al. 2006).
In addition to the downstream effectors, ROP/RAC GTPases can also activate a
signalling pathway that acts directly on the actin cytoskeleton by regulating the
activity of ABPs (Bamburg 1999; Lawler 1999). Among the multiple mechanisms
regulating actin binding and depolymerizing activity of ADFs/coffilins, Rho
GTPases play an important role. For example, phosphorylation of NtADF1 at the
conserved Ser-6 position is mediated by either NtRac1 in tobacco pollen tubes
(Chen and Citovsky 2003) or a Ca2+-dependent protein (Allwood et al. 2001).
These results indicate that the interplay among ROP, Ca2+ signalling and the
cytoskeleton is a complex network in pollen tube growth.
Compared to ROP downstream signalling pathways, knowledge of signalling
upstream of the ROP small GTPases has been limited. Recent work in tomato
suggests that a membrane receptor-mediated signalling may be upstream of the
ROP pathway. LePRK1 and LePRK2 are leucine repeat (LRR)-containing mem-
brane receptor-like kinases present in tomato pollen tubes. The extracelular LRR
domain of LePRK2 can interact with the pollen protein LAT52 prior to germina-
tion, and switches to interact with a stigma-secreted protein, LeSTIG1, during and
after germination (Tang et al. 2002, 2004). Pollen with reduced expression of
LAT52 failed to hydrate and germinate in vitro, and formed aberrant pollen tubes
in the pistil (Muschietti et al. 1994). Knockdown of LePRK2 has been shown to
reduce pollen germination and tube growth rate (Zhang et al. 2008), suggesting that
LePRK2 positively affects pollen germination and tube growth. The intracellular
kinase domain interacts with KPP, a RopGEF, which plays a role in the polarity of
the tube (Kaothien et al. 2005). These observations appear to be in line with the
results showing that co-expressing AtRopGEF12 and AtPRK2a (a LePRK2 homo-
log in Arabidopsis) resulted in wider pollen tube tips (Zhang and McCormick
2007). Pollen tubes with reduced LePRK2 expression display abnormal vacuole
behaviour and impaired Ca2+ response (Zhang et al. 2008). These results indicate
that the membrane receptor-like kinase LePRK2 most likely acts upstream of ROP
signalling in controlling polar growth of the pollen tube.
13.4.5 Pectin Methyltransferase and Pectin Modification
The plant cell wall is a refined network of hemicelluloses and pectin, with cellulose
microfibrils embedded in this matrix (Denès et al. 2000). In most plant species, the
pollen tube cell wall consists of two distinct layers. The inner layer, composed of
callose and cellulose, is a secondary wall that is first visible some distance behind
the tip (15 mm in tobacco pollen tubes), while the outer wall layer contains mainly
pectin and lacks cellulose (Ferguson et al. 1998). Unlike other plant cell walls, the
components of the pollen tube tip comprise the outer wall layer, a single layer of
pectin but not including callose or cellulose. For rapidly elongating pollen tubes,
pectin is an extension-controlling cell wall molecule. Hence, the metabolism and
modification of pectin are critical for pollen tube extension in the pistil. The pectin
network is thought to be established chemically with three covalently linked
components, namely homogalacturonan (HGA), rhamnogalacturonan-I and rham-
nogalacturonan II (O’Neill et al. 1990). HGAs are linear polymers composed of
1,4-a-D-galacturonic acid (GalA) residues. GalAs represent the major component
of pectin. These polysaccharides are synthesized, polymerized and methylesterified
with side chains within the Golgi apparatus and, subsequently, transported via the
Golgi-derived vesicles to the pollen tip and secreted into the wall in a highly
methylesterified state (Sterling et al. 2001). Following vesicle discharge, HGAs
are gradually de-esterified by wall-associated pectin methylesterases (PMEs;
EC 3.1.1.11). During the process of de-esterification, the methoxyl groups of the
polygalacturonic acid chain are converted into carboxyl groups. Thus, the acidic
residues are exposed and cross-linked by Ca2+, generating a new layer of pectin
(Catoire et al. 1998; Limberg et al. 2000). The cell wall tends to stiffen, as a result
of PME action and cross-linking by calcium. However, the activity of PMEs is
influenced by the surrounding conditions. The localized reduction in pH, caused
by protons produced in a de-esterification reaction, can inhibit some PME isoforms.
It also stimulates the activity of cell wall hydrolases, and results in cell wall
loosening, promoting wall yielding and cell expansion (Wen et al. 1999; Ren
and Kermode 2000). Also, polar growth of the pollen tube tip is fine-tuned by
the equilibrium of the interplay between these counteracting effects (Röckel
et al. 2008).
In plants, PMEs represent a large family of enzymes that are involved in cell
wall extension, cell adhesion, fruit maturation and senescence, cambial cell differ-
entiation, and resistance to biotic attack (Tieman and Handa 1994; Wen et al. 1999;
Micheli et al. 2000; Chen and Citovsky 2003). The Arabidopsis genome contains 67
PME-related genes, among which at least 18 isoforms are highly expressed in
pollen (Pina et al. 2005), underlying their importance during pollen germination
and/or pollen tube growth. PMEs are encoded mostly as pre-proproteins with a
catalytic domain and an inhibitor domain (PMEI; Markovic and Jornvall 1992; Tian
et al. 2006). The pre-region is required for protein targeting to the endoplasmic
reticulum (ER), while the PMEI domain, also known as pro-region, is a large
peptide of about 250 amino acid residues (Giovane et al. 2004). This peptide
regulates the correct folding of PME or inhibition of the enzyme activity to prevent
premature demethylesterification of pectin (Micheli 2001).
Pollen-specific PMEs have been identified from a number of plant species,
including willow (Futamura et al. 2000), tobacco (Bosch et al. 2005), maize
(Wakeley et al. 1998) and Arabidopsis (Bosch et al. 2005; Jiang et al. 2005;
Bosch and Hepler 2006; Tian et al. 2006). The loss of function of the pollen PME
encoded by VANGARD1 (VGD1) resulted in male sterility. Although vgd1 mutant
pollen grains are capable of germinating, pollen tube growth in the transmitting
tract of the style is greatly retarded. Pollen tubes of vgd1 frequently burst when
germinated and grown in vitro. Further analysis indicates that the total pectin
demethylesterification activity of PMEs in the vgd1 pollen grains is decreased to
82% of that of the wild-type pollen. These results suggest that the vgd1 mutation
reduces PME activity in pollen grains, thereby altering the mechanical properties
of the cell wall (Jiang et al. 2005). The second PME, AtPPM1 (A. thaliana pollen-
specific PME1), is also highly and specifically expressed in Arabidopsis pollen
grains. Compared with the wild-type pollen, overall PME activity is reduced by
20% in atppm1 pollen grains. Mutation of atppm1 has no apparent effect on pollen
morphology, including the cell wall pattern. However, the pollen tubes exhibited
curved, irregular morphology and are markedly stunted 6 h after germination, and
the mutant pollen tubes remain short 24 h after germination (Tian et al. 2006).
These results demonstrate the indispensable functions of PMEs in tip growth of
pollen tubes. Consistently, application of exogenous PME causes a dramatic
decrease in pollen germination rate and growth rate of pollen tubes in tobacco
and lily. PME-treated pollen tubes of both species display a significant increase in
cell wall thickness, especially at the apex (Bosch et al. 2005), suggesting that
excessive PME is also detrimental to the pollen tube. It is therefore speculated that
PME activity may be modulated by its inhibitors for pollen tube growth in vivo.
PEMIs, one of the most potent inhibitors of PME activity in vitro or in planta,
have been identified (Wolf et al. 2003; Giovane et al. 2004; Raiola et al. 2004;
Lionetti et al. 2007; Pelloux et al. 2007). Homology search reveals that there are at
least 14 genes in Arabidopsis that may encode either PMEIs or invertase inhibitors
(Rausch and Greiner 2004). Inhibition occurs through the formation of a reversible
1:1 complex of PME and PMEI (D’Avino et al. 2003). In the complex, PMEI
almost completely covers the pectin-binding cleft that contains the active site of the
PME (Johansson et al. 2002). The model of interaction between PME and PMEI
may reflect the intramolecular binding of the PMEI pro-region to the PME domain
(Hothorn et al. 2004). AtPMEI1 and AtPMEI2 are two proteinaceous inhibitors of
PME specifically expressed in Arabidopsis pollen. They interact physically with
each other, and AtPMEI2 inactivates AtPPME1 in vitro. While AtPMEI2 shows the
growth-promoting effect, AtPPME1 exerts the growth-inhibiting effect on pollen
tubes. The antagonistic effects of AtPMEI2 and AtPPME1 may correlate with their
location in pollen tubes. In tobacco, AtPPME1-YFP is uniformly distributed
throughout the cell wall of pollen tubes, including the tip region, but AtPMEI2-
YFP is detected exclusively at the cell wall of pollen tube tips. Experimental
evidence shows that AtPMEI2, but not AtPPME1, undergoes endocytic trafficking.
AtPMEI2 may be constantly and selectively internalized in the region 5–15 mm

behind the tip, and accumulates exclusively in the apex to restrain the premature
activity of freshly secreted PMEs from the Golgi apparatus (Röckel et al. 2008).
These results show that polar distribution of PMEI isoforms in pollen tubes
regulates cell wall stability via interaction with PMEs. However, it is not clear as
to what mediates the formation and dissociation of the PME-PMEI complex,
although the stability of the complex is influenced by pH (Giovane et al. 2004),
and how this complex functions during tip growth of pollen tubes remains to be
elucidated.
13.4.6 Pollen Tube Guidance
Unlike other polar growing cells, pollen tube growth is a guided process. Following
germination on the stigma surface, pollen tubes penetrate the style and navigate
through the transmitting tract, emerge from the septum, grow along the funicular
surface and target precisely the embryo sac embedded in the ovule. Communication
between the male and female is essential during this process. The whole process of
pollen tube guidance is often divided into two phases, namely the sporophytic
phase, which includes the stage when the pollen tube elongates through the trans-
mitting tract of the pistil, and the gametophytic phase that includes the pollen tube
emerging from the transmitting tract, growing along the funiculus, until its entry
into the micropyle of the ovule (Higashiyama 2002; Y.H. Chen et al. 2007). How
the pollen tube finds its way to the ovule is a question that has fascinated biologists
for centuries (Lord 2003). However, the mechanisms underlying the pollen tube
guidance to the ovule are being revealed. Chemoattractants for the pollen tube have
been proposed. The presumed attractant is expected to be able to diffuse and form a
concentration gradient that can lead to the directional growth of the pollen tube.
Multiple signals have been implicated to play a role in the sporophytic phase of
pollen tube guidance. On the stigma surface, several potential cues have been
proposed to direct pollen growth. These include lipids, water, ions and proteins.
In the family Solanaceae, pollen germination and directional growth require either
triglycerides in the stigma exudates, or a gradient of water established by the
stigmatic lipids over an aqueous component of the exudates (Wolters-Arts et al.
1998). In the transmitting tract, several extracellular matrix proteins have been
implicated in pollen tube guidance. The major extracellular arabinogalactans in-
clude TTS1 and TTS2 proteins in tobacco (Wang et al. 1993; Cheung et al. 1995;
Wu et al. 2000), a 9-kd stigma-style cysteine-rich adhesion (SCA) protein
(Park et al. 2000) and stylar pectic polysaccharides (Mollet et al. 2000) in lily
and g-amino butyric acid (GABA; Palanivelu et al. 2003). These substances provide
nutritional and guidance cues, and support directional pollen tube growth in the
transmitting tract of the female reproductive organ.
Guided pollen tube growth requires the presence of directional cues in the stylar
tissue of the pistil. A simple model to achieve the directional growth is to form a
gradient of a diffusible compound that acts as a chemotactic attractant for the pollen
tube. This model requires a gradient of attracting signal in the female sporophyte
and a sensing system in the pollen tube. It has been suggested that TTS glycopro-
teins may serve as guiding cues in the pistil (Cheung et al. 1995; Wu et al. 1995,
2000). These glycoproteins bind to the pollen tube surface and are incorporated into
the pollen tube wall in the process of tube elongation (Wu et al. 2000). Also, TTS
proteins possess features of positive chemotropic signals, for example, attracting
pollen tubes, promoting tube elongation when applied in vitro, slowing the pollen
tube growth in vivo by reduction of TTS levels, and the ability to form a gradient of
glycosylation levels in the transmitting tissue (Cheung et al. 1995; Wu et al. 1995,
2000). In addition, a haptotactic model has been proposed by analogy to neuron
guidance in animals, where a large extracellular matrix molecule (laminin) forming
a trail, and a small signalling peptide (netrin) sensed by a neuronal receptor act
together to guide neuron growth. In agreement with such a model, it has been shown
that a large matrix molecule (pectin) and a small peptide (SCA) together are
responsible for pollen tube adhesion in the lily style (Lord 2003). This finding
suggests that both haptotactic and chemotactic mechanisms might be involved in
pollen tube guidance in the style. However, it is not clear whether haptotactic
mechanisms are unique for open styles and chemotactic mechanisms for solid
styles, as in Arabidopsis.
Studies on GABA action in pollen tube guidance support the view that the
gradient of GABA is critical for normal growth and guidance of pollen tubes in
female tissues (Palanivelu et al. 2003). GABA is a four-carbon o-amino acid
previously known as an inhibitory neurotransmitter, and plays roles in developing
nervous systems in animals (Petroff 2002). In Arabidopsis, the formation of an
increasing gradient of GABA from stigma to the micropyle of the ovule by
POLLEN PISTIL IINTERACTION 2 (POP2) has been proposed. POP2 encodes
a transaminase that degrades GABA. Compared to wild-type tissues, the recessive
pop2 mutation results in more than a 100-fold increase in GABA levels in mutant
tissues. The increase in GABA concentrations and elimination of a gradient offer an
explanation to growth arrest and loss of micropylar pollen tube guidance observed
in the pop2 mutant. However, the pop2 mutant is male and female fertile when
crossed with the wild type. Furthermore, pollen tubes of the pop2 mutant, but not
the wild type, fail to target the pop2 mutant ovule, suggesting that POP2 tubes may
turn over GABA efficiently. However, pop2 mutant tubes are hypersensitive to
exogenous GABA, presumably because they lack the enzyme that degrades it
(Palanivelu et al. 2003). In animals, GABA interacts with its receptors through
different mechanisms to function as an inhibitory neurotransmitter in the central
nervous system (Pinal and Tobin 1998; Satin and Kinard 1998; Brice et al. 2002).
Similar receptors have not been identified in plants. In addition, pollen tube
guidance is a species-specific feature. It remains to be seen how GABA, as an
attracting signal, fulfils such a requirement.
Another possible attracting signal is the maize EGG APPARATUS 1 (ZmEA1)

protein (Márton et al. 2005). Knockdown of ZmEA1 expression by RNAi technique
abolishes micropylar pollen tube guidance in maize. The ZmEA1 gene encodes a
small secreted protein and is expressed in the egg apparatus of the embryo sac.
Bioinformatics analysis has identified ZmEA1 homologues in other plant species,
including Arabidopsis and rice (Gray-Mitsumune and Matton 2006). The EA1
proteins share a conserved 27–29 amino acid motif (EA box) near the C terminus
and are divergent outside the box. This satisfies the requirement for species
specificity as a signalling molecule. However, it is not clear whether EA1-like
proteins can also function in pollen tube guidance in other species.
Genetic and laser ablation studies demonstrate an essential role of the gameto-
phytic cells in pollen tube guidance. Laser ablation of the synergid cell, but not the
egg and central cell, abolishes micropylar pollen tube guidance in Torenia, which is
characterized by the embryo sac that protrudes from the ovule integuments and is
accessible to experimental manipulation (Higashiyama et al. 2001). The two syner-
gid cells act synergistically, since ablation of one synergid reduces the efficiency of
pollen tube guidance. In the Arabidopsis mutant eostre, although the gametophytic
cells of the embryo sac are mis-specified, resulting in two egg cells and only one
synergid cell, pollen tube attraction takes place normally and sperm cells are
released correctly in some mutant embryo sacs (Pagnussat et al. 2007). In most
species, the pollen tube enters the synergid cell by extending through the filiform
apparatus, a thickened cell wall extension of the synergid cell at the micropyle.
Loss-of-function mutation of the synergid-expressed gene AtMYB98 in Arabidopsis
causes a defect in the filiform apparatus and pollen tube guidance (Kasahara et al.
2005). As discussed above, knockdown of the egg/synergid-expressed ZmEA1 in
maize also leads to defective pollen tube guidance (Márton et al. 2005). Taken
together, these results suggest an essential role of the synergid cell in pollen tube
guidance.
In addition to the synergid, other gametophytic cells of the embryo sac, such as
the egg and central cell, may also play a role in pollen tube guidance. In Arabi-
dopsis, mutation in the central cell-expressed gene AtCCG (CENTRAL CELL
GUIDANCE) abolishes micropylar pollen tube guidance, demonstrating a critical
role of the central cell in guidance (Y.H. Chen et al. 2007). CCG encodes a nuclear
protein that most likely acts as a transcription regulator in Arabidopsis. Pollen tubes
cease to grow near the micropyle, or turn away or bypass the micropyle of the
mutant embryo sacs, indicating that the mutant embryo sacs are unable to produce
or emit the attracting signal. Furthermore, mutation of the egg-expressed gene
GEX3 also abolishes pollen tube guidance in the ovule (Alandete-Saez et al.
2008). GEX3 is expressed in the egg cell and pollen grains, and it encodes a plasma
membrane-localized protein of unknown function. These findings indicate that
pollen tube guidance is the integrative effects of multiple signalling pathways and
combined contribution of individual gametophytic cells. Compared to the female
guidance during the tube elongation, little is known about the mechanism of how
pollen tubes perceive and respond to the female attracting signals.
13.5 Conclusions
As a tricellular free-living male germ unit, pollen possesses many unique features,
such as its specialized cell wall, and has been the focus of developmental genetics
research in recent years. The pollen tube has been used as a model for understand-
ing polar cell growth for many years. Pollen germination and tube growth are the
results of orchestrated interactions among diverse signalling pathways (Ca2+ and
others), small GTPases and the cytoskeleton. Although a large volume of informa-
tion has been gained, especially in pollen tube cell biology in past decades,
molecular mechanisms governing pollen germination and tube growth remain to
be elucidated in the context of modern developmental genetics. More recently,
pollen transcriptome and proteome analyses have generated tremendous amounts of
data. If combined with large-scale genetic and biochemical analyses, these data
may help to provide insights into the regulatory mechanisms of pollen germination
and tube growth.
References
Alandete-Saez M, Ron M, McCormick S (2008) GEX3, expressed in the male gametophyte and in
the egg cell of Arabidopsis thaliana, is essential for micropylar pollen tube guidance and plays
a role during early embryogenesis. Mol Plant 1:586–598
Allwood EG, Smertenko AP, Hussey PJ (2001) Phosphorylation of plant actin depolymerising
factor by calmodulin-like domain protein kinase. FEBS Lett 499:97–100
Ariizumi T, Hatakeyama K, Hinata K, Sato S, Kato T, Tabata S, Toriyama K (2003) A novel male-
sterile mutant of Arabidopsis thaliana, faceless pollen-1, produces pollen with a smooth
surface and an acetolysis-sensitive exine. Plant Mol Biol 53:107–181
Astrom H, Sorri O, Raudaskoski M (1995) Role of microtubules in the movement of the vegetative
nucleus and generative cell in tobacco pollen tubes. Sex Plant Reprod 8:61–69
Bamburg JR (1999) Proteins of the ADF/cofilin family: essential regulators of actin dynamics.
Annu Rev Cell Biol 15:185–230
Batoko H, Zheng HQ, Hawes C, Moore I (2000) A Rab1 GTPase is required for transport between
the endoplasmic reticulum and Golgi apparatus and for normal Golgi movement in plants.
Plant Cell 12:2201–2218
Becker JD, Feijó JA (2007) How many genes are needed to make a pollen tube? Lessons from
transcriptomics. Ann Bot 100:1117–1123
Becker JD, Boavida LC, Carneiro J, Haury M, Feijó JA (2003) Transcriptional profiling of
Arabidopsis tissues reveals the unique characteristics of the pollen transcriptome. Plant Physiol
133:713–725
Bednarska E (1989) The effect of exogenous Ca2+ ions on pollen grain germination and pollen
tube growth-investigations with the use of 45Ca2+, verapamil, La3+ and ruthenium red. Sex
Plant Reprod 2:53–58
Berken A (2006) ROPs in the spotlight of plant signal transduction. Cell Mol Life Sci
63:2446–2459
Blackmore S (2007) Pollen and spores: microscopic keys to understanding the earth’s biodiversity.
Plant Syst Evol 263:3–12
Bosch M, Hepler PK (2006) Silencing of the tobacco pollen pectin methylesterase NtPPME1
results in retarded in vivo pollen tube growth. Planta 223:736–745
Bosch M, Cheung AY, Hepler PK (2005) Pectin methylesterase, a regulator of pollen tube growth.
Plant Physiol 138:1334–1346
Brice NL, Varadi A, Ashcroft SJH, Molnar E (2002) Metabotropic glutamate and GABA B
receptors contribute to the modulation of glucose-stimulated insulin secretion in pancreatic
beta cells. Diabetologia 45:242–252
Burridge K, Wennerberg K (2004) Rho and Rac take center stage. Cell 116:167–179
Bushart TJ, Roux SJ (2007) Conserved features of germination and polarized cell growth: a few
insights from a pollen-fern spore comparison. Ann Bot 99:9–17
Cai G, Moscatelli A, Cresti M (1997) Cytoskeletal organization and pollen tube growth. Trends
Plant Sci 2:86–91
Cai G, Romagnoli S, Moscatelli A, Ovidi E, Gambellini G, Tiezzi A, Cresti M (2000) Identifica-
tion and characterization of a novel microtubule-based motor associated with membranous
organelles in tobacco pollen tubes. Plant Cell 12:1719–1736
Camacho L, Malhó R (2003) Endo/exocytosis in the pollen tube apex is differentially regulated by
Ca2+ and GTPases. J Exp Bot 54:83–92
Campanoni P, Blatt MR (2007) Membrane trafficking and polar growth in root hairs and pollen
tubes. J Exp Bot 58:65–74
Cárdenas L, Lovy-Wheeler A, Wilsen KL, Hepler PK (2005) Actin polymerization promotes the
reversal of streaming in the apex of pollen tubes. Cell Motil Cytoskel 61:112–127
Cárdenas L, Lovy-Wheeler A, Kunkel JG, Hepler PK (2008) Pollen tube growth oscillations and
intracellular calcium levels are reversibly modulated by actin polymerization. Plant Physiol
146:1611–1621
Carroll AD, Moyen C, Van Kesteren P, Tooke F, Battey NH, Brownlee C (1998) Ca2+, annexins,
and GTP modulate exocytosis from maize root cap protoplasts. Plant Cell 10:1267–1276
Catoire L, Pierron M, Morvan C, du Penhoat CH, Goldberg R (1998) Investigation of the action
patterns of pectin methylesterase isoforms through kinetic analyses and NMR spectroscopy –
implications in cell wall expansion. J Biol Chem 273:33150–33156
Charzynska M, Murgia M, Cresti M (1989) Ultrastructure of the vegetative cell of Brassica napus
pollen with particular reference to microbodies. Protoplasma 152:22–28
Chen CY (2002) Functional analysis of actin depolymerizing factors (ADFs) in Rac-mediated
pollen tube growth. PhD Dissertation, University of Massachusetts, Amherst, MA
Chen MH, Citovsky V (2003) Systemic movement of a tobamovirus requires host cell pectin
methylesterase. Plant J 35:386–392
Chen CY, Wong EI, Vidali L, Estavillo A, Hepler PK, Wu HM, Cheung AY (2002) The regulation
of actin organization by actin depolymerizing factor (ADF) in elongating pollen tubes. Plant
Cell 14:2175–2190
Chen T, Teng N, Wu X, Wang Y, Tang W, Sămaj J, Baluška F, Lin J (2007) Disruption of actin
filaments by latrunculin B affects cell wall construction in Picea meyeri pollen tube by
disturbing vesicle trafficking. Plant Cell Physiol 48:19–30
Chen YH, Li HJ, Shi DQ, Yuan L, Liu J, Sreenivasan R, Baskar R, Grossniklaus U, Yang WC
(2007) The central cell plays a critical role for pollen tube guidance in Arabidopsis. Plant Cell
19:3563–3577
Cheung AY, Wu HM (2004) Overexpression of an Arabidopsis formin stimulates supernumerary
actin cable formation from pollen tube cell membrane. Plant Cell 16:257–269
Cheung AY, Wu HM (2007) Structural and functional compartmentalization in pollen tubes. J Exp
Bot 58:75–82
Cheung AY, Wu HM (2008) Structural and signaling networks for the polar cell growth machinery
in pollen tubes. Annu Rev Plant Biol 59:547–572
Cheung AY, Wang H, Wu HM (1995) A floral transmitting tissue-specific glycoprotein attracts
pollen tubes and stimulates their growth. Cell 82:383–393
Cheung AY, Chen CY-h, Glaven RH, de Graaf BHJ, Vidali L, Hepler PK, Wu HM (2002) Rab2
GTPase regulates vesicle trafficking between the endoplasmic reticulum and the Golgi bodies
and is important to pollen tube growth. Plant Cell 14:945–962
Ciampolini F, Shivanna KR, Cresti M (2001) Organization of the stigma and transmitting tissue of
rice, Oryza sativa (L.). Plant Biol 3:149–155
Cole RA, Fowler JE (2006) Polarized growth: maintaining focus on the tip. Curr Opin Plant Biol
9:579–588
da Costa-Nunes JA, Grossniklaus U (2004) Unveiling the gene-expression profile of pollen.
Genome Biol 5:205.1–205.3
Dai S, Li L, Chen T, Chong K, Xue Y, Wang T (2006) Proteomic analyses of Oryza sativa mature
pollen reveal novel proteins associated with pollen germination and tube growth. Proteomics
6:2504–2529
Dai S, Chen T, Chong K, Xue Y, Liu S, Wang T (2007) Proteomics identification of differentially
expressed proteins associated with pollen germination and tube growth reveals characteristics
of germinated Oryza sativa pollen. Mol Cell Proteomics 6:207–230
D’Avino R, Camardella L, Christensen TM, Giovane A, Servillo L (2003) Tomato pectin
methylesterase: modeling, fluorescence, and inhibitor interaction studies - comparison with
the bacterial (Erwinia chrysanthemi) enzyme. Proteins 53:830–839
de Graaf BH, Cheung AY, Andreyeva T, Levasseur K, Kieliszewski M, Wu HM (2005) Rab11
GTPase-regulated membrane trafficking is crucial for tip-focused pollen tube growth in
tobacco. Plant Cell 17:2564–2579
Denès JM, Baron A, Renard CM, Péan C, Drilleau JF (2000) Different action patterns for apple
pectin methylesterase at pH 7.0 and 4.5. Carbohydrate Res 327:385–393
Derksen J, Rutten T, Lichtscheidl IK, de Win AHN, Pierson ES, Rongen G (1995) Quantitative
analysis of the distribution of organelles in tobacco pollen tubes: implication for exocytosis and
endocytosis. Protoplasma 188:267–276
DerMardirossian C, Bokoch GM (2005) GDIs: central regulatory molecules in Rho GTPase
activation. Trends Cell Biol 15:356–363
Desrivieres S, Cooke FT, Morales-Johansson H, Parker PJ, Hall MN (2002) Calmodulin controls
organization of the actin cytoskeleton via regulation of phosphatidylinositol (4,5)-bisphosphate
synthesis in Saccharomyces cerevisiae. Biochem J 366:945–951
Dickinson HG (1995) Dry stigmas, water and self-incompatibility in Brassica. Sex Plant Reprod
8:1–10
Dixit R, Rizzo C, Nasrallah ME, Nasrallah JB (2001) The Brassica MIP-MOD gene encodes a
functional water channel that is expressed in the stigma epidermis. Plant Mol Biol 45:51–62
Dowd PE, Coursol S, Skirpan AL, Kao T-h, Gilroy S (2006) Petunia phospholipase C1 is involved
in pollen tube growth. Plant Cell 18:1438–1453
Edlund AF, Swanson R, Preuss D (2004) Pollen and stigma structure and function: the role of
diversity in pollination. Plant Cell 16:s84–s97
Elleman CJ, Dickinson HG (1986) Pollen-stigma interactions in Brassica. IV. Structural reorgani-
zation in the pollen grains during hydration. J Cell Sci 80:141–157
Elleman CJ, Sarker RH, Aivalakis G, Slade H, Dickinson HG (1989) Molecular physiology of the
pollen stigma interaction in Brassica. In: Lord E, Bernier G (eds) Plant reproduction – from
floral induction to pollination. American Society of Plant Physiologists, Rockville, MD, Am
Soc Plant Physiol Symp Series, pp 136–145
Estruch JJ, Kadwell S, Merlin E, Crossland L (1994) Cloning and characterization of a maize
pollen-specific calcium dependent calmodulin-independent protein kinase. Proc Natl Acad Sci
USA 91:8837–8841
Fauré J, Dagher MC (2001) Interactions between Rho GTPases and Rho GDP dissociation
inhibitor (Rho-GDI). Biochimie 83:409–414
Ferguson C, Teeri TT, Siika-aho M, Read SM, Bacic A (1998) Location of cellulose and callose in
pollen tubes and grains of Nicotiana tabacum. Planta 206:452–460
Fiebig A, Mayfield JA, Miley NL, Chau S, Fischer RL, Preuss D (2000) Alterations in CER6, a
gene identical to CUT1, differentially affect long-chain lipid content on the surface of pollen
and stems. Plant J 12:2001–2008
Franklin-Tong VE (1999a) Signaling and the modulation of pollen tube growth. Plant Cell
11:727–738
Franklin-Tong VE (1999b) Signaling in pollination. Curr Opin Plant Biol 2:490–495
Franklin-Tong VE, Hackett G, Hepler PK (1997) Ratio imaging of Ca2+ in the self-incompatibility
response in pollen tubes of Papaver rhoeas. Plant J 12:1375–1386
Franklin-Tong VE, Holdaway-Clarke TL, Straatman KR, Kunkel JG, Hepler PK (2002) Involve-
ment of extracellular calcium influx in the self-incompatibility incompatibility response of
Papaver rhoeas. Plant J 29:333–345
Frietsch S, Wang YF, Sladek C, Poulsen LR, Romanowsky SM, Schroeder JI, Harper JF (2007) A
cyclic nucleotide-gated channel is essential for polarized tip growth of pollen. Proc Natl Acad
Sci USA 104:14531–14536
Fu Y, Wu G, Yang Z (2001) Rop GTPase-dependent dynamics of tip-localized f-actin controls tip
growth in pollen tubes. J Cell Biol 152:1019–1032
Fu Y, Li H, Yang Z (2002) The ROP2 GTPase controls the formation of cortical fine F-actin and
the early phase of directional cell expansion during Arabidopsis organogenesis. Plant Cell
14:777–794
Furness CA (2007) Why does some pollen lack apertures? A review of inaperturate pollen in
eudicots. Bot J Linn Soc 155:29–48
Futamura N, Mori H, Kouchi H, Shinohara K (2000) Male flower-specific expression of genes for
polygalacturonase, pectin methylesterase and b-1,3-glucanase in a dioecious willow (Salix
gilgiana Seemen). Plant Cell Physiol 41:16–26
Geitmann A, Li YQ, Cresti M (1995) The role of cytoskeleton and dictyosome activity in
the pulsatory growth of Nicotiana tabacum and Petunia hybrida pollen tubes. Bot Acta
109:102–109
Giovane A, Servillo L, Balestrieri C, Raiola A, D’Avino R, Tamburrini M, Clardiello MA,
Camardella L (2004) Pectin methylesterase inhibitor. BBA-Proteins Proteomics 1696:245–252
Goldman MH, Goldberg RB, Mariani C (1994) Female sterile tobacco plants are produced by
stigma-specific cell ablation. EMBO J 13:2976–2984
Golovkin M, Reddy ASN (2003) A calmodulin-binding protein from Arabidopsis has an essential
role in pollen germination. Proc Natl Acad Sci USA 100:10558–10563
Gossot O, Geitmann A (2007) Pollen tube growth: coping with mechanical obstacles involves the
cytoskeleton. Planta 226:405–416
Gray-Mitsumune M, Matton DP (2006) The EGG APPARATUS 1 gene from maize is a member of
a large gene family found in both monocots and dicots. Planta 223:618–625
Grebe M, Xu J, Möbius W, Ueda T, Nakano A, Geuze HJ, Rook MB, Scheres B (2003)
Arabidopsis sterol endocytosis involves actin-mediated trafficking via ARA6-positive early
endosomes. Curr Biol 13:1378–1387
Gu Y, Wang Z, Yang Z (2004) ROP/RAC GTPase: an old new master regulator for plant signaling.
Curr Opin Plant Biol 7:527–536
Gu Y, Zheng Y, Williams DA (2005) RhoH GTPase: a key regulator of hematopoietic cell
proliferation and apoptosis? Cell Cycle 4:201–202
Gu Y, Li S, Lord EM, Yang Z (2006) Members of a novel class of Arabidopsis Rho guanine
nucleotide exchange factors control Rho GTPase-dependent polar growth. Plant Cell 18:
366–381
Guilford WA, Schneider DM, Labovitz J, Opella SJ (1988) High-resolution solid state 13C-NMR
spectroscopy of sporopollenins from different plant taxa. Plant Physiol 86:134–136
Gungabissoon RA, Jiang CJ, Drøbak BK, Maciver SK, Hussey PJ (1998) Interaction of maize
actin-depolymerising factor with actin and phosphoinositides and its inhibition of plant
phospholipase C. Plant J 16:689–696
Gungabissoon RA, Khan S, Hussey PJ (2001) Interaction of elongation factor 1 alpha from Zea
mays (ZmEF-1 alpha) with F-actin and interplay with the maize actin severing protein,
ZmADF3. Cell Motil Cytoskel 49:104–111
Hannoufa A, Negruk V, Eisner G, Lemieux B (1996) The CER3 gene of Arabidopsis thaliana is
expressed in leaves, stems, roots, flowers and apical meristems. Plant J 10:459–467
Hauber I, Herth W, Reiss HD (1984) Calmodulin in tip growing plant cells, visualised by
fluorescing calmodulin binding phenothiazines. Planta 162:33–39
Helling D, Possart A, Cottier S, Klahre U, Kost B (2006) Pollen tube tip growth depends on plasma
membrane polarization mediated by tobacco PLC3 activity and endocytic membrane recy-
cling. Plant Cell 18:3519–3534
Hepler PK, Vidali L, Cheung AY (2001) Polarized cell growth in higher plants. Annu Rev Cell
Dev Biol 17:159–187
Hernández-Pinzón I, Ross JHE, Barnes KA, Damant AP, Murphy DJ (1999) Composition and role
of tapetal lipid bodies in the biogenesis of the pollen coat of Brassica napus. Planta 208:
588–598
Heslop-Harrison J, Heslop-Harrison Y, Cresti M, Tiezzi A, Moscatelli A (1988) Cytoskeletal
elements, cell shaping and movement in the angiosperm pollen tube. J Cell Sci 91:49–60
Higashiyama T (2002) The synergid cell: attractor and acceptor of the pollen tube for double
fertilization. J Plant Res 115:149–160
Higashiyama T, Yabe S, Sasaki N, Nishimura Y, Miyagishima S, Kuroiwa H, Kuroiwa T (2001)
Pollen tube attraction by the synergid cell. Science 293:1480–1483
Hiscock SJ, Allen AM (2008) Diverse cell signalling pathways regulate pollen-stigma interac-
tions: the search for consensus New Phytol 179:286–317
Hiscock SJ, Bown D, Gurr SJ, Dickinson HG (2002) Serine esterases are required for pollen tube
penetration of the stigma in Brassica. Sex Plant Reprod 15:65–74
Holdaway-Clarke TL, Hepler PK (2003) Control of pollen tube growth: role of ion gradients and
fluxes. New Phytol 159:539–563
Holdaway-Clarke TL, Feijó JA, Hackett GR, Kunkel JG, Hepler PK (1997) Pollen tube growth and
the intracellular cytosolic calcium gradient oscillate in phase while extracellular calcium influx
is delayed. Plant Cell 9:1999–2010
Honys D, Twell D (2003) Comparative analysis of the Arabidopsis pollen transcriptome. Plant
Physiol 132:640–652
Honys D, Twell D (2004) Transcriptome analysis of haploid male gametophyte development in
Arabidopsis. Genome Biol 5:R85
Hothorn M, D’Angelo I, Marquez JA, Greiner S, Scheffzek K (2004) The invertase inhibitor Nt-
CIF from tobacco: a highly thermostable four-helix bundle with an unusual N-terminal
extension. J Mol Biol 335:987–995
Huang S, Blanchoin L, Chaudhry F, Franklin-Tong VE, Staiger CJ (2004) A gelsolin-like protein
from Papaver rhoeas pollen (PrABP80) stimulates calcium-regulated severing and depolymer-
ization of actin filaments. J Biol Chem 279:23364–23375
Huang S, Gao L, Blanchoin L, Staiger CJ (2006) Heterodimeric capping protein from Arabidopsis
is regulated by phosphatidic acid. Mol Biol Cell 17:1946–1958
Hussey PJ, Ketelaar T, Deeks MJ (2006) Control of the actin cytoskeleton in plant cell growth.
Annu Rev Plant Biol 57:109–125
Hwang JU, Yang Z (2006) Small GTPases and spatiotemporal regulation of pollen tube growth.
Plant Cell Monogr 3:95–116
Hwang JU, Vernoud V, Szumlanski A, Nielsen E, Yang Z (2008) A tip-localized RhoGAP
controls cell polarity by globally inhibiting Rho GTPase at the cell apex. Curr Biol 18:1907–
1916
Iwano M, Shiba H, Miwa T, Che FS, Takayama S, Nagai T, Miyawaki A, Isogai A (2004) Ca2+
dynamics in a pollen grain and papilla cell during pollination of Arabidopsis. Plant Physiol
136:3562–3571
Jaffe LA, Weisenseel MH, Jaffe LF (1975) Calcium accumulations within the growing tips of
pollen tubes. J Cell Biol 67:488–492
Janmey PA (1994) Phosphoinositides and calcium as regulators of cellular actin assembly and
disassembly. Annu Rev Physiol 56:169–191
Jiang L, Yang SL, Xie LF, Puah CS, Zhang XQ, Yang WC, Sundaresan V, Ye D (2005)
VANGUARD1 encodes a pectin methylesterase that enhances pollen tube growth in the
Arabidopsis style and transmitting tract. Plant Cell 17:584–596
Johansson K, El-Ahmad M, Friemann R, Jornvall H, Markovic O, Eklund H (2002) Crystal
structure of plant pectin methylesterase. FEBS Lett 514:243–249
Joos U, van Aken J, Kristen U (1994) Microtubules are involved in maintaining the cellular
polarity in pollen tubes of Nicotiana sylvestris. Protoplasma 179:5–15
Kandasamy M, Kristen U (1987) Developmental aspects of ultrastructure, histochemistry and
receptivity of the stigma of Nicotiana sylvevestis. Am J Bot 60:427–437
Kandasamy MK, Nasrallah JB, Nasrallah ME (1994) Pollen-pistil interactions and developmental
regulation of pollen tube growth in Arabidopsis. Development 120:3405–3418
Kaothien P, Ok SH, Shuai B, Wengier D, Cotter R, Kelley D, Kiriakopolos S, Muschietti J,
McCormick S (2005) Kinase partner protein interacts with the LePRK1 and LePRK2 receptor
kinases and plays a role in polarized pollen tube growth. Plant J 42:492–503
Kasahara RD, Portereiko MF, Sandaklie-Nikolova L, Rabiger DS, Drews GN (2005) MYB98 is
required for pollen tube guidance and synergid cell differentiation in Arabidopsis. Plant Cell
17:2981–2992
Koornneef M, Hanhart CJ, Thiel F (1989) A genetic and phenotypic description of eceriferum
(cer) mutants in Arabidopsis thaliana. J Hered 80:118–122
Kost B (2008) Spatial control of Rho (Rac-Rop) signaling in tip-growing plant cells. Trends Cell
Biol 18:119–127
Kost B, Spielhofer P, Chua NH (1998) A GFP-mouse talin fusion protein labels plant actin filaments
in vivo and visualizes the actin cytoskeleton in growing pollen tubes. Plant J 16:393–401
Kost B, Mathur J, Chua NH (1999) Cytoskeleton in plant development. Curr Opin Plant Biol
2:462–470
Kovar DR, Drobak BK, Staiger CJ (2000) Maize profilin isoforms are functionally distinct. Plant
Cell 12:583–598
Krichevsky A, Kozlovsky SV, Tian GW, Chen MH, Zaltsman A, Citovsky V (2007) How pollen
tubes grow. Dev Biol 303:405–420
Kuang A, Musgrave ME (1996) Dynamics of vegetative cytoplasm during generative cell forma-
tion and pollen maturation in Arabidopsis thaliana. Protoplasma 194:81–90
Lawler S (1999) Regulation of actin dynamics: the LIM kinase connection. Curr Biol
9:R800–R802
Lazzaro MD, Cárdenas L, Bhatt AP, Justus CD, Phillips MS, Holdaway-Clarke TL, Hepler PK
(2005) Calcium gradients in conifer pollen tubes: dynamic properties differ from those seen in
angiosperms. J Exp Bot 56:2619–2628
Lee JY, Lee DH (2003) Use of serial analysis of gene expression technology to reveal changes in
gene expression in arabidopsis pollen undergoing cold stress. Plant Physiol 132:517–529
Lee YJ, Szumlanski A, Nielsen E, Yang Z (2008) Rho-GTPase-dependent filamentous actin
dynamics coordinate vesicle targeting and exocytosis during tip growth. J Cell Biol
181:1155–1168
Li H, Lin Y, Heath RM, Zhu MX, Yang Z (1999) Control of pollen tube tip growth by a Rop
GTPase-dependent pathway that leads to tip-localized calcium influx. Plant Cell 11:1731–1742
Li H, Shen JJ, Zheng ZL, Lin Y, Yang Z (2001) The Rop GTPase switch controls multiple
developmental processes in Arabidopsis. Plant Physiol 126:670–684
Li M, Xu W, Yang W, Kong Z, Xue Y (2007) Genome-wide gene expression profiling reveals
conserved and novel molecular functions of the stigma in rice. Plant Physiol 144:1797–1812
Limberg G, Korner R, Buchholt HC, Christensen T, Roepstorff P, Mikkelsen JD (2000) Analysis
of different de-esterification mechanisms for pectin by enzymatic fingerprinting using endo-
pectin lyase and endopolygalacturonase II from A. niger. Carbohydrate Res 327:293–307
Lionetti V, Raiola A, Camardella L, Giovane A, Obel N, Pauly M, Favaron F, Cervone F,
Bellincampi D (2007) Overexpression of pectin methylesterase inhibitors in Arabidopsis
restricts fungal infection by Botrytis cinerea. Plant Physiol 143:1871–1880
Lord EM (2003) Adhesion and guidance in compatible pollination. J Exp Bot 54:47–54
Lovy-Wheeler A, Wilsen KL, Baskin TI, Hepler PK (2005) Enhanced fixation reveals the apical
cortical fringe of actin filaments as a consistent feature of the pollen tube. Planta 221:95–104
Lovy-Wheeler A, Cardenas L, Kunkel JG, Hepler PK (2007) Differential organelle movement on
the actin cytoskeleton in lily pollen tubes. Cell Motil Cytoskel 64:217–232
Luu DT, Qin X, Morse D, Cappadocia M (2000) S-RNase uptake by compatible pollen tubes in
gametophytic self-incompatibility. Nature 407:649–651
Malhó R, Trewavas AJ (1996) Localized apical increases of cytosolic free calcium control pollen
tube orientation. Plant Cell 8:1935–1949
Malhó R, Read ND, Pais M, Trewavas AJ (1994) Role of cytosolic calcium in the reorientation of
pollen tube growth. Plant J 5:331–341
Malhó R, Liu Q, Monteiro D, Rato C, Camacho L, Dinis A (2006) Signalling pathways in pollen
germination and tube growth. Protoplasma 228:21–30
Markovic O, Jornvall H (1992) Disulfide bridges in tomato pectinesterase: variations from
pectin esterases of other species; conservation of possible active site segments. Protein Sci
1:1288–1292
Márton ML, Cordts S, Broadhvest J, Dresselhaus T (2005) Micropylar pollen tube guidance by egg
apparatus 1 of maize. Science 307:573–576
Mascarenhas JP (1993) Molecular mechanisms of pollen tube growth and differentiation. Plant
Cell 5:1303–1314
Maurel C, Verdoucq L, Luu DT, V Santoni (2008) Plant aquaporins: membrane channels with
multiple integrated functions. Annu Rev Plant Biol 59:595–624
Mayfield JA, Preuss D (2000) Rapid initiation of Arabidopsis pollination requires the oleosin-
domain protein GRP17. Nature Cell Biol 2:128–130
Mayfield JA, Fiebig A, Johnstone SE, Preuss D (2001) Gene families from the Arabidopsis
thaliana pollen coat proteome. Science 292:2482–2485
Micheli F (2001) Pectin methylesterases: cell wall enzymes with important roles in plant physiol-
ogy. Trends Plant Sci 6:414–419
Micheli F, Sundberg B, Goldberg R, Richard L (2000) Radial distribution pattern of pectin
methylesterases across the cambial region of hybrid aspen at activity and dormancy. Plant
Physiol 124:191–199
Millar AA, Clemens S, Zachgo S, Giblin EM, Taylor DC, Kunst L (1999) CUT1, an Arabidopsis
gene required for cuticular wax biosynthesis and pollen fertility, encodes a very-long-chain
fatty acid condensing enzyme. Plant Cell 11:825–838
Miller DD, Callaham DA, Gross DJ, Hepler PK (1992) Free Ca2+ gradient in growing pollen tubes
of lilium. J Cell Sci 101:7–12
Miller DD, Scordilis SP, Hepler PK (1995) Identification and localization of three classes of
myosins in pollen tubes of Lilium longiflorum and Nicotiana alata. J Cell Sci 108:2549–2563
Miller DD, Lancelle SA, Hepler PK (1996) Actin microfilaments do not form a dense meshwork in
Lilium longiflorum pollen tube tips. Protoplasma 195:123–132
Minc N, Bratman SV, Basu R, Chang F (2009) Establishing new sites of polarization by micro-
tubules. Curr Biol 19:83–94
Molendijk AJ, Ruperti B, Palme K (2004) Small GTPases in vesicle trafficking. Curr Opin Plant
Biol 7:694–700
Mollet JC, Park SY, Nothnagel EA, Lord EM (2000) A lily stylar pectin is necessary for pollen
tube adhesion to an in vitro stylar matrix. Plant Cell 12:1737–1749
Morel J, Fromentin J, Blein JP, Simon-Plas F, Elmayan T (2004) Rac regulation of NtrbohD, the
oxidase responsible for the oxidative burst in elicited tobacco cell. Plant J 37:282–293
Moutinho A, Trewavas AJ, Malhó R (1998) Relocation of a Ca2+-dependent protein kinase activity
during pollen tube reorientation. Plant Cell 10:1499–1510
Muschietti J, Dircks L, Vancanneyt G, McCormick S (1994). LAT52 protein is essential for
tomato pollen development: Pollen expressing antisense LAT52 RNA hydrates and germinates
abnormally and cannot achieve fertilization. Plant J 6:321–338
Nasrallah JB (2000) Cell-cell signaling in the self-incompatibility response. Curr Opin Plant Biol
3:368–373
Nepi M, Pacini E (1993) Pollination, pollen viability and pistil receptivity in Cucurbita pepo. Ann
Bot 72:526–536
Nibau C, Wu HM, Cheung AY (2006) RAC/ROP GTPases: hubs for signal integration and
diversification in plants. Trends Plant Sci 11:309–315
Nishikawa S, Zinkl GM, Swanson RJ, Maruyama D, Preuss D (2005) Callose (b-1,3-glucan) is
essential for Arabidopsis pollen wall patterning, but not tube growth. BMC Plant Biol 5:22–30
Obermeyer G, Weisenseel MH (1991) Calcium channel blocker and calmodulin antagonists affect
the gradient of free calcium ions in lily pollen tubes. Eur J Cell Biol 56:319–327
O’Neill M, Albersheim P, Darvill AG (1990) The pectic polysaccharides of primary cell walls. In:
Dey PM, Harborne JB (eds) Methods in plant biochemistry. Carbohydrates. Academic Press,
London, pp 415–441
Pacini E (1996) Types and meaning of pollen carbohydrate reserves. Sex Plant Reprod 9:362–366
Pacini E (1997) Tapetum character states: analytical keys for tapetum types and activities. Can J
Bot 75:1448–1459
Pagnussat GC, Yu HJ, Sundaresan V (2007) Cell-fate switch of synergid to egg cell in Arabidopsis
eostre mutant embryo sacs arises from misexpression of the BEL1-like homeodomain gene
BLH1. Plant Cell 19:3578–3592
Palanivelu R, Brass L, Edlund AF, Preuss D (2003) Pollen tube growth and guidance is regulated
by POP2, an Arabidopsis gene that controls GABA levels. Cell 114:47–59
Park SK, Twell D (2001) Novel patterns of ectopic cell plate growth and lipid body distribution in
the Arabidopsis gemini pollen1 mutant. Plant Physiol 126:899–909
Park SY, Jauh GY, Mollet JC, Eckard KJ, Nothnagel EA, Walling LL, Lord EM (2000) A lipid
transfer-like protein is necessary for lily pollen tube adhesion to an in vitro stylar matrix. Plant
Cell 12:151–163
Parton RM, Fischer-Parton S, Trewavas AJ, Watahiki MJ (2003) Pollen tubes exhibit
regular periodic membrane trafficking events in the absence of apical extension. J Cell Sci
116:2707–2719
Paxson-Sowders DM, Owen HA, Makaro CA (1997) A comparative ultrastructural analysis of
exine pattern development in wild-type Arabidopsis and a mutant defective in pattern forma-
tion. Protoplasma 198:53–65
Paxon-Sowders DM, Dodrill CH, Owen HA, Makaroff CA (2001) DEX1, a novel plant protein, is
required for exine pattern formation during pollen development in Arabidopsis. Plant Physiol
127:1739–1749
Pelloux J, Rustérucci C, Mellerowicz EJ (2007) New insights into pectin methylesterase structure
and function. Trends Plant Sci 12:267–277
Petroff OA (2002) GABA and glutamate in the human brain. Neuroscientist 8:562–573
Pfeffer S, Aivazian D (2004) Targeting Rab GTPases to distinct membrane compartments. Nature
Rev Mol Cell Biol 5:886–896
Pierson ES, Lichtscheidl IK, Derksen J (1990) Structure and behaviour of organelles in living
pollen tubes of Lilium longiflorum. J Exp Bot 41:1461–1468
Pierson ES, Miller DD, Callaham DA, Shipley AM, Rivers BA, Cresti M, Hepler PK (1994) Pollen
tube growth is coupled to the extracellular calcium ion flux and the intracellular calcium
gradient: effect of BAPTA-type buffers and hypertonic media. Plant Cell 6:1815–1828
Pierson ES, Miller DD, Callaham DA, van Aken J, Hackett G, Hepler PK (1996) Tip-localized
entry fluctuates during pollen tube growth. Dev Biol 174:160–173
Piffanelli P, Ross JHE, Murphy DJ (1997) Intra- and extracellular lipid composition and associated
gene expression patterns during pollen development in Brassica napus. Plant J 11:549–562
Pina C, Pinto F, Feijo JA, Becker JD (2005) Gene family analysis of the Arabidopsis pollen
transcriptome reveals biological implications for cell growth, division control, and gene
expression regulation. Plant Physiol 138:744–756
Pinal CS, Tobin AJ (1998) Uniqueness and redundancy in GABA production. Perspect Dev
Neurobiol 5:109–118
Post-Beittenmiller D (1996) Biochemistry and molecular biology of wax production in plants.
Annu Rev Plant Physiol Plant Mol Biol 47:405–430
Preuss D, Lemieux B, Yen G, Davis RW (1993) A conditional sterile mutation eliminates surface
components from Arabidopsis pollen and disrupts cell signaling during fertilization. Genes
Dev 7:974–985
Preuss ML, Serna J, Falbel TG, Bednarek SY, Nielsen E (2004) The Arabidopsis Rab GTPase
RabA4b localizes to the tips of growing root hair cells. Plant Cell 16:1589–603
Preuss ML, Schmitz AJ, Thole JM, Bonner HK, Otegui MS, Nielsen E (2006) A role for the
RabA4b effector protein PI-4Kbeta1 in polarized expansion of root hair cells in Arabidopsis
thaliana. J Cell Biol 172:991–998
Qualmann B, Kessels MM (2008) Actin nucleation: putting the brakes on Arp2/3. Curr Biol 18:
R420–R423
Raiola A, Camardella L, Giovane A, Mattei B, De Lorenzo G, Cervone F, Bellincampi D (2004)
Two Arabidopsis thaliana genes encode functional pectin methylesterase inhibitors. FEBS Lett
557:199–203
Rathore KS, Cork RJ, Robinson KR (1991) A cytoplasmic gradient of Ca21 is correlated with the
growth of lily pollen tubes. Dev Biol 148:612–619
Rausch T, Greiner S (2004) Plant protein inhibitors of invertases. Biochim Biophys Acta
1696:253–261
Ren C, Kermode AR (2000) An increase in pectin methyl esterase activity accompanies dormancy
breakage and germination of yellow cedar seeds. Plant Physiol 124:231–242
Ren H, Xiang Y (2007) The function of actin-binding proteins in pollen tube growth. Protoplasma
230:171–182
Röckel N, Wolf S, Kost B, Rausch T, Greiner S (2008) Elaborate spatial patterning of cell-wall
PME and PMEI at the pollen tube tip involves PMEI endocytosis, and reflects the distribution
of esterified and de-esterified pectins. Plant J 53:133–143
Romagnoli S, Cai G, Cresti M (2003) In vitro assays demonstrate that pollen tube organelles use
kinesin-related motor proteins to move along microtubules. Plant Cell 15:251–269
Romagnoli S, Cai G, Faleri C, Yokota E, Shimmen T, Cresti M (2007) Microtubule- and actin
filament-dependent motors are distributed on pollen tube mitochondria and contribute differ-
ently to their movement. Plant Cell Physiol 48:345–361
Sarker RH, Elleman CJ, Dickinson HG (1988) Control of pollen hydration in Brassica requires
continued protein-synthesis, and glycosylation is necessary for intraspecific incompatibility.
Proc Natl Acad Sci USA 85:4340–4344
Satin LS, Kinard TA (1998) Neurotransmitters and their receptors in the islets of Langerhans of the
pancreas - What messages do acetylcholine, glutamate, and GABA transmit? Endocrine
8:213–223
Schiøtt M, Romanowsky SM, Baekgaard L, Jakobsen MK, Palmgren MG, Harper JF (2004) A
plant plasma membrane Ca2+ pump is required for normal pollen tube growth and fertilization.
Proc Natl Acad Sci USA 101:9502–9507
Schopfer CR, Nasrallah ME, Nasrallah JB (1999) The male determinant of self-incompatibility in
Brassica. Science 286:1697–1700
Seabra MC, Wasmeier C (2004) Controlling the location and activation of Rab GTPases. Curr
Opin Cell Biol 16:451–457
Shiba H, Takayama S, Iwano M, Shimosato H, Funato M, Nakagawa T, Che FS, Suzuki G,
Watanabe M, Hinata K (2001) A pollen coat protein, SP11SCR, determines the pollen
S-specificity in the self- incompatibility of Brassica species. Plant Physiol 125:2095–2103
Snowman BN, Kovar DR, Shevchenko G, Franklin-Tong VE, Staiger CJ (2002) Signal-mediated
depolymerization of actin in pollen during the self-incompatibility response. Plant Cell
14:2613–2626
Speranza A, Calzoni GL, Pacini E (1997) Occurrence of mono- or disaccharides and polysaccha-
ride reserves in mature pollen grains. Sex Plant Reprod 10:110–115
Staiger CJ, Hussey PJ (2004) Actin and actin-modulating proteins. In: Hussey PJ (ed) The plant
cytoskeleton in cell differentiation and development. Blackwell, Oxford, pp 32–80
Steer MW, Steer JM (1989) Pollen tube tip growth. New Phytol 111:323–358
Stenmark H, Olkkonen VM (2001) The Rab GTPase family. Genome Biol 2:3007.1–3007.7
Sterling JD, Quigley HF, Orellana A, Mohnen D (2001) The catalytic site of the pectin biosyn-
thetic enzyme a-1,4-galacturonosyltransferase is located in the lumen of the Golgi. Plant
Physiol 127:360–371
Surpin M, Raikhel N (2004) Traffic jams affect plant development and signal transduction. Nature
Rev Mol Cell Biol 5:100–109
Swanson R, Edlund AF, Preuss D (2004) Species specificity in pollen-pistil interactions. Annu Rev
Genet 38:793–818
Tamm LK, Crane J, Kiessling V (2003) Membrane fusion: a structural perspective on the interplay
of lipids and proteins. Curr Opin Struct Biol 13:453–466
Tang W, Ezcurra I, Muschietti J, McCormick S (2002) A cysteine-rich extracellular protein,
LAT52, interacts with the extracellular domain of the pollen receptor kinase LePRK2. Plant
Cell 14:2277–2287
Tang W, Kelley D, Ezcurra I, Cotter R, McCormick S (2004) LeSTIG1, an extracellular binding
partner for the pollen receptor kinases LePRK1 and LePRK2, promotes pollen tube growth
in vitro. Plant J 39:343–353
Tao WJ, Liang SP, Lu YT (2004) The roles of calmodulin polar distribution during pollen
hydration and germination. Can J Bot 82:774–780
Tian GW, Chen MH, Zaltsman A, Citovsky V (2006) Pollen-specific pectin methylesterase
involved in pollen tube growth. Dev Biol 294:83–91
Tieman DM, Handa AK (1994) Reduction in pectin methylesterase activity modifies tissue
integrity and action levels in ripening tomato (Lycopersicon esculentum Mill.) fruits. Plant
Physiol 106:429–436
Tirlapur UK, Scali M, Moscatelli A, Casino CD, Cai G, Tiezzi A, Gresti M (1994) Confocal image
analysis of spatial variations in immunocytochemically identified calmodulin during pollen
hydration, germination and pollen tube tip growth in Nicotiana tabacum L. Zygote 2:63–68
Tiwari SC, Polito VS (1990) An analysis of the role of actin during pollen activation leading to
germination in pear (Pyrus communis L.): treatment with cytochalasin D. Sex Plant Reprod
3:121–129
Tung CW, Dwyer KG, Nasrallah ME, Nasrallah JB (2005) Genome-wide identification of genes
expressed in Arabidopsis pistils specifically along the path of pollen tube growth. Plant Physiol
138:977–989
Ueda T, Uemura T, Sato MH, Nakano A (2004) Functional differentiation of endosomes in
Arabidopsis cells. Plant J 40:783–789
Valster AH, Hepler PK, Chernoff J (2000) Plant GTPases: the Rhos in bloom. Trends Cell Biol
10:141–146
Vernoud V, Horton AC, Yang Z, Nielsen E (2003) Analysis of the small GTPase gene superfamily
of Arabidopsis. Plant Physiol 131:1191–1208
Vidali L, McKenna ST, Hepler PK (2001) Actin polymerization is necessary for pollen tube
growth. Mol Biol Cell 12:2534–2545
Wakeley PR, Rogers HJ, Rozycka M, Greenland AJ, Hussey PJ (1998) A maize pectin methylesterase-
like gene, ZmC5, specifically expressed in pollen. Plant Mol Biol 37:187–192
Wang H, Wu HM, Cheung AY (1993) Development and pollination regulated accumulation and
glycosylation of a stylar transmitting tissue-specific proline-rich protein. Plant Cell 5:1639–1650
Wang Y, Fan L, Zhang W, Zhang W, Wu W (2004) Ca2+-Permeable channels in the plasma
membrane of Arabidopsis pollen are regulated by actin microfilaments. Plant Physiol
132:3892–3904
Wang Y, Zhang WZ, Song LF, Zou JJ, Su Z, Wu WH (2008) Transcriptome analyses show
changes in gene expression to accompany pollen germination and tube growth in Arabidopsis.
Plant Physiol 148:1201–1211
Wen FS, Zhu YM, Haves MC (1999) Effect of pectin methylesterase gene expression on pea root
development. Plant Cell 11:1129–1140
Wetzel CLR, Jensen WA (1992) Studies of pollen maturation in cotton: the storage reserve
accumulation phase. Sex Plant Reprod 5:117–127
Wolf S, Grsic-Rausch S, Rausch T, Greiner S (2003) Identification of pollen-expressed pectin
methylesterase inhibitors in Arabidopsis. FEBS Lett 555:551–555
Wolters-Arts M, Lush WM, Mariani C (1998) Lipids are required for directional pollen-tube
growth. Nature 392:818–821
Wolters-Arts M, Van Der Weerd L, Van Aelst AC, Van As H, Mariani C (2002) Water-conducting
properties of lipids during pollen hydration. Plant Cell Environ 25:513–519
Wu HM, Wang H, Cheung AY (1995) A pollen tube growth stimulatory glycoprotein is deglyco-
sylated by pollen tubes and displays a glycosylation gradient in the flower. Cell 82:395–403
Wu HM, Wong E, Ogdahl J, Cheung AY (2000) A pollen tube growth-promoting arabinogalactan
protein from Nicotiana alata is similar to the tobacco TTS protein. Plant J 22:165–176
Xiang Y, Huang X, Wang T, Zhang Y, Liu Q, Hussey PJ, Ren H (2007) ACTIN BINDING
PROTEIN29 from Lilium pollen plays an important role in dynamic actin remodeling. Plant
Cell 19:1930–1946
Yalovsky S, Bloch D, Sorek N, Kost B (2008) Regulation of membrane trafficking, cytoskeleton
dynamics, and cell polarity by ROP/RAC GTPases. Plant Physiol 147:1527–1543
Yang Z (2002) Small GTPases: versatile signaling switches in plants. Plant Cell 14 Suppl:S375–
S388
Yang Z, Fu Y (2007) ROP/RAC GTPase signaling. Curr Opin Plant Biol 10:490–494
Yang WC, Sundaresan V (2000) Genetics of gametophyte biogenesis in Arabidopsis. Curr Opin
Plant Biol 3:53–57
Yeoh S, Pope B, Mannherz HG, Weeds A (2002) Determining the differences in actin binding by
human ADF and cofilin. J Mol Biol 315:911–925
Yokota E, Shimmen T (2000) Characterization of native actin-binding proteins from pollen.
Myosin and the actin-binding proteins, 135-ABP and 115-ABP. In: Staiger CJ, Balŭska F,
Volkmann D, Barlow P (eds) Actin: a dynamic framework for multiple plant cell functions.
Kluwer, Dordrecht, pp 103–118
Yokota E, McDonald AR, Liu B, Shimmen T, Palevitz BA (1995) Localization of a 170 kDa
myosin heavy chain in plant cells. Protoplasma 185:178–187
Yoon GM, Dowd PE, Gilroy S, McCubbin AG (2006) Calcium-dependent protein kinase isoforms
in petunia have distinct functions in pollen tube growth, including regulating polarity. Plant
Cell 18:867–878
Zerial M, McBride H (2001) Rab proteins as membrane organizers. Nature Rev Mol Cell Biol
2:107–117
Zhang Y, McCormick S (2007) A distinct mechanism regulating a pollen-specific guanine
nucleotide exchange factor for the small GTPase Rop in Arabidopsis thaliana. Proc Natl
Acad Sci USA 104:18830–18835
Zhang D, Wengier D, Shuai B, Gui CP, Muschietti J, McCormick S, Tang WH (2008) The pollen
receptor kinase LePRK2 mediates growth-promoting signals and positively regulates pollen
germination and tube growth. Plant Physiol 148:1368–1379
Zheng ZL, Yang Z (2000) The Rop GTPase: an emerging signaling switch in plants. Plant Mol
Biol 44:1–9
Zinkl GM, Zwiebel BI, Grier DG, Preuss D (1999) Pollen-stigma adhesion in Arabidopsis: a
species-specific interaction mediated by lipophilic molecules in the pollen exine. Development
126:5431–5440
View publication stats

Pollen Germination and Tube Growth

Uploaded by

Copyright:

Available Formats

Pollen Germination and Tube Growth

Uploaded by

Document Information

Original Description:

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Pollen Germination and Tube Growth

Uploaded by

Copyright:

Available Formats

See discussions, stats, and author profiles for this publication at: https://www.researchgate.

Pollen Germination and Tube Growth

Chapter · October 2010

The user has requested enhancement of the downloaded file.

Pollen Germination and Tube Growth

D.-Q. Shi and W.-C. Yang

Plant Developmental Biology -

D.-Q. Shi and W.-C. Yang

D.-Q. Shi and W.-C. Yang

E.C. Pua and M.R. Davey (eds.), 245

13.2 Mature Pollen Grains

13.2.1 Pollen Wall

13.2.2 Pollen Maturation

Before anther dehiscence, pollen undergoes a maturation process, which prepares

The maturation processes include pollen dehydration and accumulation of storage

13.2.2.1 Pollen Dehydration

13.2.2.2 Reserves in Mature Pollen Grains

Magnolia grandiflora, Passiflora caerulea and Pinus halepensis, pollen grains

13.2.2.3 Transcriptome and Proteome of Pollen Grains

13.3 Pollen-Stigma Interaction

The pistil is a female sexual organ, an elongated structure located in a central

13.3.1 The Stigma

13.3.2 Pollen Recognition

epidermal cells, to initiate an SI reaction (Schopfer et al. 1999; Nasrallah 2000). In

13.3.3 Pollen Adherence and Hydration

13.4 Pollen Germination and Tube Growth

13.4.1 Calcium Signalling in Pollen Germination

13.4.2 The Cytoskeleton

Cytoskeleton dynamics regulate many important cellular processes, such as organ-

13.4.2.1 Actin Cytoskeleton

13.4.2.2 Microtubule Cytoskeleton

13.4.3 Crosstalk Between Calcium Signalling and Cytoskeleton

a number of reversible phenotypes, including elimination of the acidic domain at

13.4.4 Small GTPases and Pollen Tube Growth

13.4.4.1 Rab GTPases

a member of the Rab11 subfamily of tobacco, is localized predominantly to the

13.4.4.2 Rho-Like GTPases

13.4.5 Pectin Methyltransferase and Pectin Modification

AtPMEI2 may be constantly and selectively internalized in the region 5–15 mm

13.4.6 Pollen Tube Guidance

Another possible attracting signal is the maize EGG APPARATUS 1 (ZmEA1)

View publication stats

You might also like