Divergent functions of two clades of flavodoxin in diatoms mitigate oxidative stress and iron limitation.

Phytoplankton rely on diverse mechanisms to adapt to the decreased iron bioavailability and oxidative stress-inducing conditions of today's oxygenated oceans, including replacement of the iron-requiring ferredoxin electron shuttle protein with a less-efficient iron-free flavodoxin under iron-limiting conditions. Yet, diatoms transcribe flavodoxins in high-iron regions in contrast to other phytoplankton. Here, we show that the two clades of flavodoxins present within diatoms exhibit a functional divergence, with only clade II flavodoxins displaying the canonical role in acclimation to iron limitation. We created CRISPR/Cas9 knock-outs of the clade I flavodoxin from the model diatom Thalassiosira pseudonana and found that these cell lines are hypersensitive to oxidative stress, while maintaining a wild-type response to iron limitation. Within natural diatom communities, clade I flavodoxin transcript abundance is regulated over the diel cycle rather than in response to iron availability, whereas clade II transcript abundances increase either in iron-limiting regions or under artificially induced iron limitation. The observed functional specialization of two flavodoxin variants within diatoms reiterates two major stressors associated with contemporary oceans and illustrates diatom strategies to flourish in diverse aquatic ecosystems.

The dynamic trophic architecture of open-ocean protist communities revealed through machine-guided metatranscriptomics.

Intricate networks of single-celled eukaryotes (protists) dominate carbon flow in the ocean. Their growth, demise, and interactions with other microorganisms drive the fluxes of biogeochemical elements through marine ecosystems. Mixotrophic protists are capable of both photosynthesis and ingestion of prey and are dominant components of open-ocean planktonic communities. Yet the role of mixotrophs in elemental cycling is obscured by their capacity to act as primary producers or heterotrophic consumers depending on factors that remain largely uncharacterized. Here, we develop and apply a machine learning model that predicts the in situ trophic mode of aquatic protists based on their patterns of gene expression. This approach leverages a public collection of protist transcriptomes as a training set to identify a subset of gene families whose transcriptional profiles predict trophic mode. We applied our model to nearly 100 metatranscriptomes obtained during two oceanographic cruises in the North Pacific and found community-level and population-specific evidence that abundant open-ocean mixotrophic populations shift their predominant mode of nutrient and carbon acquisition in response to natural gradients in nutrient supply and sea surface temperature. Metatranscriptomic data from ship-board incubation experiments revealed that abundant mixotrophic prymnesiophytes from the oligotrophic North Pacific subtropical gyre rapidly remodeled their transcriptome to enhance photosynthesis when supplied with limiting nutrients. Coupling this approach with experiments designed to reveal the mechanisms driving mixotroph physiology provides an avenue toward understanding the ecology of mixotrophy in the natural environment.

Fe limitation decreases transcriptional regulation over the diel cycle in the model diatom Thalassiosira pseudonana.

Iron (Fe) is an important growth factor for diatoms and its availability is further restricted by changes in the carbonate chemistry of seawater. We investigated the physiological attributes and transcriptional profiles of the diatom Thalassiosira pseudonana grown on a day: night cycle under different CO2/pH and iron concentrations, that in combination generated available iron (Fe') concentrations of 1160, 233, 58 and 12 pM. We found the light-dark conditions to be the main driver of transcriptional patterns, followed by Fe' concentration and CO2 availability, respectively. At the highest Fe' (1160 pM), 55% of the transcribed genes were differentially expressed between day and night, whereas at the lowest Fe' (12 pM), only 28% of the transcribed genes displayed comparable patterns. While Fe limitation disrupts the diel expression patterns for genes in most central metabolism pathways, the diel expression of light- signaling molecules and glycolytic genes was relatively robust in response to reduced Fe'. Moreover, we identified a non-canonical splicing of transcripts encoding triose-phosphate isomerase, a key-enzyme of glycolysis, generating transcript isoforms that would encode proteins with and without an active site. Transcripts that encoded an active enzyme maintained a diel expression at low Fe', while transcripts that encoded the non-active enzyme lost the diel expression. This work illustrates the interplay between nutrient limitation and transcriptional regulation over the diel cycle. Considering that future ocean conditions will reduce the availability of Fe in many parts of the oceans, our work identifies some of the regulatory mechanisms that may shape future ecological communities.