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Open Access Publications from the University of California

Scholarly Works (4 results)

  • Thesis
  • Peer Reviewed
UC San Diego Electronic Theses and Dissertations (2006)
Diatoms, a major group of primary producers in the ocean, are estimated to be responsible for over 40% of oceanic carbon fixation. As one of the predominant biosilicifying organisms in the world, diatoms are a model system for studying biological interactions with silicon. This research has focused on characterizing a novel family of transporters, called silicon transporters (SITs), that are specific for the uptake and efflux of silicon in diatoms. SIT sequences were isolated from evolutionarily diverse diatom species. Multi-gene copies were identified in most species, and phylogenetic analysis showed SITs grouped according to species. Structural analysis suggested SITs evolved through an internal gene duplication. Comparative sequence analysis revealed repeats of a conserved sequence motif, GXQ. A model of silicon transport consistent with known aspects of uptake was developed based on this motif. Analysis of SIT protein and mRNA expression, as well as measurements of uptake activity, was done on synchronously growing cultures of the diatom, Thalassiosira pseudonana. Immunoblot analysis using a newly developed SIT-specific antibody showed peaks in SIT protein levels correlated with active periods of silica incorporation. Quantitative PCR showed each T. pseudonana SIT (TpSIT1-3) peaked prior to cell wall synthesis. However, a disconnect between protein and mRNA levels suggested SITs were primarily regulated at the translational or post-translational level. In addition, rates of surge uptake suggested SIT activity was internally controlled by the rate of silica incorporation. Silicon uptake kinetics in diatoms were measured to determine the extent of nonsaturable uptake and the role of SITs. In all diatom species examined, a time-dependent transition from nonsaturable to saturable uptake kinetics was observed. In addition, both forms of uptake were affected by the SIT-specific antibody suggesting SITs were the predominant means of silicon uptake into the cell. Under some conditions, SITs had enormous flexibility in their rate of transport and appeared to act as selectivity gates rather than controlling agents in uptake. A model of diatom silicon uptake, consistent with this and previously published data, was developed based on the interplay between SITs, intracellular soluble silicon pools, and cell wall silica incorporation
    Cover page: Molecular insights into the function and regulation of diatom silicon transporters
    • Thesis
    • Peer Reviewed
    UC Santa Barbara Electronic Theses and Dissertations (2022)

    Diatoms serve as a critical link between the marine carbon (C), nitrogen (N), and silicon (Si) biogeochemical cycles through an obligate requirement for silicon for frustule formation. The siliceous frustules of diatoms serve as ballast, leading to the disproportionate contribution of diatoms to organic carbon export compared to similarly sized non-siliceous cells. Despite accounting for ~30% of marine primary production, diatom growth and productivity are limited in much of the surface ocean by some combination of low concentrations of dissolved iron (dFe), nitrate (NO3-), and silicic acid (Si(OH)4). Furthermore, as obligate phototrophs living in the sunlight region of the ocean, diatom cells can become increasingly light-limited as light quality and quantity decreases over depth. These physiological limitations can lead to shifts in phytoplankton community composition and act as strong controls on phytoplankton physiology, marine biogeochemical cycling, and carbon export.With the growing abundance of marine nucleic acid sequencing data through large collaborative projects such as EXPORTS, BIOS-SCOPE, BioGEOTRACES, Global Ocean Sampling, Tara Oceans, and Bio-GO-SHIP, which all share an interest in moving toward linking cellular metabolism with ocean processes, there is a need to develop and test relationships between molecular markers, phytoplankton physiology, and related biogeochemical stocks and rates. By relating metatranscriptomic patterns to biogeochemical stock and rate measurements during diatom blooms in the California Upwelling Zone we evaluate the ecological relevance of diatom transcriptomic fingerprints of iron, nitrogen, silicon, and light limitations based on those previously characterized in laboratory studies, while also exploring potentially undiscovered Si related genes. The first chapter of this dissertation (Chapter 1) explores the transcriptomic and physiological changes that occur in iron limited diatom assemblages that lead to bulk increases in biogenic silica relative to particulate organic carbon and nitrogen. Under Fe-limitation, diatom dominated communities can double the respective ratios of biogenic silica (bSi) to particulate organic carbon (C) and nitrogen (N), which has implications for carbon export efficiency given the ballasted nature of the silica-based diatom cell wall. Understanding the drivers of this altered cellular stoichiometry will foster a predictive understanding of how diatom carbon export is affected by low Fe. In a simulated upwelling experiment, water from the 10⁰C isotherm was transported from depth, incubated shipboard, and left untreated or amended with dissolved Fe or the Fe-binding siderophore desferrioxamine-B (+DFB) to induce Fe-limitation. The phytoplankton communities within all treatments became dominated by diatoms, which displayed hallmark signatures of Fe-limitation in the +DFB treatment, including elevated particulate Si:C and Si:N ratios. Single-cell, taxon-resolved measurements of elemental content revealed no increase in bSi content during Fe-limitation despite the higher transcript abundance of silicon associated genes. We posit that the observed increase in bSi relative to C and N was primarily due to reductions in C fixation and N assimilation, driven by lower transcript expression of key Fe-dependent genes. Chapter two pairs biogeochemical measurements of Si uptake with metatranscriptomic analysis of communities incubated with and without added dissolved Si to investigate the molecular response of diatom communities to different nutrient statuses. Steep gradients in macronutrients in the surface ocean near Monterey Bay, CA allowed for the sampling of both macronutrient replete phytoplankton communities as well as communities experiencing N-limitation, and N and Si co-limitation. Metatranscriptomic analysis revealed that N-limited communities exhibited dynamic shifts in N and C transcriptional patterns, but despite a 97% reduction in biomass-specific silica production rates, only modest additional changes in transcript abundance occurred with N and Si co-limitation. Transcript abundance of previously characterized Si-metabolism associated genes did not appear to be correlated with changes in Si-limitation. However, several not yet characterized Si-responsive genes were revealed to be potential indicators of kinetic Si-limitation. Transcriptomic data implied that additional Si-limitation within N and Si co-limited samples served a protective role of reducing the degree of N-limitation and the associated heavy physiological toll. The last chapter of this dissertation (Chapter 3) investigates the extent to which ambient light availability at the lower depths of the euphotic zone (EZ) impacted water biogeochemistry and diatom community physiology including bSi production rates and transcript abundance. As light intensity decreased at the base of the EZ (1-5% incident irradiance) all diatom communities reduced bSi-normalized silica production rates. However, there was poor correlation between changes in rates and silicon transporter transcript abundance. Biogeochemical markers of light limitation, such as decreased ratio of chlorophyll a to particulate organic carbon, correlated with reduced transcript abundance of genes related to photosystem II as well as nitrate transport and reduction. In stratified conditions, diatom communities at the base of the euphotic zone exhibited widespread reduction of transcripts for genes related to light harvesting, photosystem II, photosynthetic electron transport, oxidative phosphorylation, and nitrogen assimilation. However, in less stratified water columns diatom communities exhibited minimal changes in transcript abundance and were likely better poised to deal with rapid changes in light availability. The combined results from this dissertation offer a comprehensive understanding of diatom community transcriptomic response to several of the determinant abiotic drivers of biogeochemical cycling within the dynamic California Upwelling Zone. Furthermore, this work validates and supports the use of prior and additional transcriptomic markers of diatom physiology.

    • 3 supplemental ZIPs
    • 2 supplemental files
    Cover page: Diatom community transcriptomic responses to abiotic stressors
    • Article
    • Peer Reviewed
    UC Merced Previously Published Works (2020)
    The blooming cosmopolitan coccolithophore Emiliania huxleyi and its viruses (EhVs) are a model for density-dependent virulent dynamics. EhVs commonly exhibit rapid viral reproduction and drive host death in high-density laboratory cultures and mesocosms that simulate blooms. Here we show that this system exhibits physiology-dependent temperate dynamics at environmentally relevant E. huxleyi host densities rather than virulent dynamics, with viruses switching from a long-term non-lethal temperate phase in healthy hosts to a lethal lytic stage as host cells become physiologically stressed. Using this system as a model for temperate infection dynamics, we present a template to diagnose temperate infection in other virus-host systems by integrating experimental, theoretical, and environmental approaches. Finding temperate dynamics in such an established virulent host-virus model system indicates that temperateness may be more pervasive than previously considered, and that the role of viruses in bloom formation and decline may be governed by host physiology rather than by host-virus densities.
      Cover page: Temperate infection in a virus–host system previously known for virulent dynamics
      • Article
      • Peer Reviewed
      UC Irvine Previously Published Works (2022)
      High-throughput RNA sequencing offers broad opportunities to explore the Earth RNA virome. Mining 5,150 diverse metatranscriptomes uncovered >2.5 million RNA virus contigs. Analysis of >330,000 RNA-dependent RNA polymerases (RdRPs) shows that this expansion corresponds to a 5-fold increase of the known RNA virus diversity. Gene content analysis revealed multiple protein domains previously not found in RNA viruses and implicated in virus-host interactions. Extended RdRP phylogeny supports the monophyly of the five established phyla and reveals two putative additional bacteriophage phyla and numerous putative additional classes and orders. The dramatically expanded phylum Lenarviricota, consisting of bacterial and related eukaryotic viruses, now accounts for a third of the RNA virome. Identification of CRISPR spacer matches and bacteriolytic proteins suggests that subsets of picobirnaviruses and partitiviruses, previously associated with eukaryotes, infect prokaryotic hosts.
        Cover page: Expansion of the global RNA virome reveals diverse clades of bacteriophagesCreative Commons 'BY-NC-ND' version 4.0 license
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