Resolving the molecular ecology of marine microbial eukaryotes with metatranscriptomes

Abstract

The surface waters of the North Pacific ocean host diverse populations of microbial eukaryotes, who collectively mediate the flux of energy and matter in their environment with a significant impact on global biogeochemical cycles. The evolutionary history of these organisms spans billions of years and includes multiple endosymbiotic events that radiated into successful lineages with complex, chimeric genomes. The trophic mode of marine microbial eukaryotes ranges from purely photo-autotrophic to heterotrophic, including a number of flexible mixotrophic strategies. The metabolism, behavior, and physiology of a majority of microbial eukaryote species are not fully understood. The emergence of environmental genomics and transcriptomics has allowed researchers to elucidate the functional capacity and activity of microorganisms in situ, including perspectives on species and entire lineages that evade isolation and culture. The influx of massive amounts of high-throughput sequence data necessitates new approaches to digesting and extracting information from these valuable data. In this dissertation I leverage poly-A+ selected, deeply sequenced metatranscriptomes to resolve the molecular ecology of in situ marine microbial eukaryote communities. In Chapter 1, I studied a diel-resolved metatranscriptome time series in the North Pacific Subtropical Gyre. I developed a method for assembly, annotation, and quantification of these metatranscriptomes to reveal oscillating patterns of gene transcription over diel cycles. I identified differences in the magnitude of diel transcript regulation across different environmental genera and examined the enrichment of diel-regulated genes in key metabolic pathways to show orchestrated metabolic re-arrangement over day and night cycles. In Chapter 2, I constructed an updated and reproducible protein reference library for marine microbial eukaryotes (MarFERReT), with the aim of improving the taxonomic annotations of environmental metatranscriptomes. I collected reference sequence material from a variety of sources, including recently available sequence data from novel and uncultured taxa, and ingested sequences through a standardized and open-source pipeline. I identify sets of core transcribed genes from these reference species that I use to estimate the coverage of environmental taxa bins and show how the incorporation of new sequence material improves the specificity of annotations. I combine the methodological approaches of Chapter 1 with the enhanced annotation capacities generated from Chapter 2 to consolidate and standardize metatranscriptome data from four different cruises into a North Pacific Eukaryotic Gene Catalog (Appendix 1). This catalog contains clustered protein sequences from all metatranscriptome assemblies, together with taxonomic annotations from the MarFERReT reference library (Chapter 2), functional annotations, and transcript abundances. Chapter 3 explores the picoeukaryotes of the dynamic North Pacific Transition Zone, using data from 3 cruises included in the North Pacific Eukaryotic Gene Catalog (Appendix 1). I focus on the small size fraction samples from latitudinal transections of the transition zone, where I observe shifts in the transcript inventory of species across biogeochemical and physical gradients. Perturbing the resource ratio of these communities in on-deck incubations revealed a subset of fast-responding species responsive to different nitrogen and iron ratios, and their differentially transcribed functions indicate a wide assortment of adaptive metabolic strategies. This dissertation has elucidated numerous strategies that marine microbial eukaryotes use to sustain, survive and thrive in their environments, ranging from the fine-tuned diel transcription of metabolic machinery in the North Pacific Subtropical Gyre, to the rapid response of species with mixed trophic modes under changing nutrient conditions in the North Pacific Transition Zone. In the process, I have developed public resources for enhanced taxonomic annotation and improved reproducibility and accessibility of these valuable metatranscriptome data sets, so they can continue to provide insight and discovery of microbial eukaryotes in the oceans.