Date of Award

2020

Degree Type

Dissertation

Degree Name

Doctor of Philosophy in Biological and Environmental Sciences

Specialization

Cell & Molecular Biology

Department

Cell & Molecular Biology

First Advisor

Bethany D. Jenkins

Abstract

Within the marine ecosystem, phytoplankton such as diatoms experience regions of nutrient limitation including macronutrients such as nitrogen, silica, and phosphate. In regions with high concentration of these nutrients but low chlorophyll a concentrations (HNLCs), phytoplankton are limited for micronutrients such as iron (Fe) an element that serves as a co-factor in proteins involved in cellular redox reactions. Of particular interest for diatoms is the Fe requirement of cellular processes localized to the chloroplast. These Fe requirements are in multiple metabolic pathways such as light reactions of photosynthesis that generate the necessary energy and reducing power necessary for anabolic processes including carbon fixation and assimilation of reduced nitrogen into amino acids.

To better understand the impacts of Fe-limitation on the diatom chloroplast, Chapter 1 focuses on the generation of a sub-cellular chloroplast proteome from a model diatom Thalassiosira pseudonana and its regulation under both Fe-limited and Fe-replete conditions. This serves a two-fold purpose, first the identification of the full complement of proteins synthesized within and localized to the diatom chloroplast, and second to provide insight to changes in chloroplast metabolic regulation in response to changes in Fe-status. Following the analysis of the diatom chloroplast proteome, this dataset was used in conjunction with incubation experiments to assess chloroplast-associated gene regulation in total diatom communities using metatranscriptome data within two distinct HNLC regions, the Southern Ocean (Chapter 2) and Subarctic Pacific (Chapter 3), and to aid in the interpretation of shifts in diatom community composition assessed using 18s rDNA community analyses.

To accomplish this, a Meta-plastid approach was taken to assess transcripts derived from both nuclear and chloroplast genomes, under ambient low-Fe and Feamended conditions. Metatranscriptome library construction was based on rRNA pulldown, allowing for the sequencing of both organellar and nuclear derived transcripts. Metatranscriptomes generated from these incubations were then queried using the T. pseudonana chloroplast proteome through a sequence similarity approach, resulting in the generation of the diatom community Meta-plastid.

Available for download on Sunday, May 08, 2022

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