Present and future potential role of toxin-producing Synechococcus in the tropical region.

As anthropogenic induced temperature rises and nutrient loadings increase in fresh and brackish environments, the ecological function of the phytoplankton community is expected to favour the picocyanobacteria, of the genus Synechococcus. Synechococcus is already a ubiquitous cyanobacterium found in both freshwater and marine environments, notwithstanding that the toxigenic species still remains unexplored in many freshwaters. Their fast growth rate and their ability to produce toxins make Synechococcus a potential dominant player in harmful algal blooms under climate change scenarios. This study examines the responses of a novel toxin-producing Synechococcus (i.e., one belonging to a freshwater clade; the other belonging to a brackish clade) to environmental changes that reflect climate change effects. We conducted a series of controlled experiments under present and predicted future temperatures, as well as under various N and P nutrients loadings. Our findings highlight how Synechococcus can be altered by the differing reactions to increasing temperature and nutrients, which resulted in considerable variations in cell abundance, growth rate, death rate, cellular stoichiometry and toxin production. Synechococcus had the highest growth observed at 28 °C, and further increases in temperature resulted in a decline for both fresh and brackish waters. Cellular stoichiometry was also altered, where more nitrogen (N) per cell was required, and the plasticity of N:P was more severe for the brackish clade. However, Synechococcus become more toxic under future scenario. Anatoxin-a (ATX) saw the greatest spike when temperature was at 34 °C especially under P-enrichment conditions. In contrast, Cylindrospermopsin (CYN) was promoted at the lowest tested temperature (25 °C) and under N-limitation. Overall, both temperature and external nutrients are the dominant control over Synechococcus toxins production. A model was also created to assess Synechococcus toxicity to zooplankton grazing. Zooplankton grazing was reduced by two folds under nutrient limitation, but temperature accounted for very insignificant change.

KidA, a multi-PAS domain protein, tunes the period of the cyanobacterial circadian oscillator.

The cyanobacterial clock presents a unique opportunity to understand the biochemical basis of circadian rhythms. The core oscillator, composed of the KaiA, KaiB, and KaiC proteins, has been extensively studied, but a complete picture of its connection to the physiology of the cell is lacking. To identify previously unknown components of the clock, we used KaiB locked in its active fold as bait in an immunoprecipitation/mass spectrometry approach. We found that the most abundant interactor, other than KaiC, was a putative diguanylate cyclase protein predicted to contain multiple Per-Arnt-Sim (PAS) domains, which we propose to name KidA. Here we show that KidA directly binds to the fold-switched active form of KaiB through its N-terminal PAS domains. We found that KidA shortens the period of the circadian clock both in vivo and in vitro and alters the ability of the clock to entrain to light-dark cycles. The dose-dependent effect of KidA on the clock period could be quantitatively recapitulated by a mathematical model in which KidA stabilizes the fold-switched form of KaiB, favoring rebinding to KaiC. Put together, our results show that the period and amplitude of the clock can be modulated by regulating the access of KaiB to the fold-switched form.

Acclimation of the photosynthetic apparatus to low light in a thermophilic Synechococcus sp. strain.

Depending upon their growth responses to high and low irradiance, respectively, thermophilic Synechococcus sp. isolates from microbial mats associated with the effluent channels of Mushroom Spring, an alkaline siliceous hot spring in Yellowstone National Park, can be described as either high-light (HL) or low-light (LL) ecotypes. Strains isolated from the bottom of the photic zone grow more rapidly at low irradiance compared to strains isolated from the uppermost layer of the mat, which conversely grow better at high irradiance. The LL-ecotypes develop far-red absorbance and fluorescence emission features after growth in LL. These isolates have a unique gene cluster that encodes a putative cyanobacteriochrome denoted LcyA, a putative sensor histidine kinase; an allophycocyanin (FRL-AP; ApcD4-ApcB3) that absorbs far-red light; and a putative chlorophyll a-binding protein, denoted IsiX, which is homologous to IsiA. The emergence of FRL absorbance in LL-adapted cells of Synechococcus sp. strain A1463 was analyzed in cultures responding to differences in light intensity. The far-red absorbance phenotype arises from expression of a novel antenna complex containing the FRL-AP, ApcD4-ApcB3, which is produced when cells were grown at very low irradiance. Additionally, the two GAF domains of LcyA were shown to bind phycocyanobilin and a [4Fe-4S] cluster, respectively. These ligands potentially enable this photoreceptor to respond to a variety of environmental factors including irradiance, redox potential, and/or oxygen concentration. The products of the gene clusters specific to LL-ecotypes likely facilitate growth in low-light environments through a process called Low-Light Photoacclimation.

Reconstitution of an intact clock reveals mechanisms of circadian timekeeping.

Circadian clocks control gene expression to provide an internal representation of local time. We report reconstitution of a complete cyanobacterial circadian clock in vitro, including the central oscillator, signal transduction pathways, downstream transcription factor, and promoter DNA. The entire system oscillates autonomously and remains phase coherent for many days with a fluorescence-based readout that enables real-time observation of each component simultaneously without user intervention. We identified the molecular basis for loss of cycling in an arrhythmic mutant and explored fundamental mechanisms of timekeeping in the cyanobacterial clock. We find that SasA, a circadian sensor histidine kinase associated with clock output, engages directly with KaiB on the KaiC hexamer to regulate period and amplitude of the central oscillator. SasA uses structural mimicry to cooperatively recruit the rare, fold-switched conformation of KaiB to the KaiC hexamer to form the nighttime repressive complex and enhance rhythmicity of the oscillator, particularly under limiting concentrations of KaiB. Thus, the expanded in vitro clock reveals previously unknown mechanisms by which the circadian system of cyanobacteria maintains the pace and rhythmicity under variable protein concentrations.

Marine Synechococcus picocyanobacteria: Light utilization across latitudes.

The most ubiquitous cyanobacteria, Synechococcus, have colonized different marine thermal niches through the evolutionary specialization of lineages adapted to different ranges of temperature seawater. We used the strains of Synechococcus temperature ecotypes to study how light utilization has evolved in the function of temperature. The tropical Synechococcus (clade II) was unable to grow under 16 °C but, at temperatures >25 °C, induced very high growth rates that relied on a strong synthesis of the components of the photosynthetic machinery, leading to a large increase in photosystem cross-section and electron flux. By contrast, the Synechococcus adapted to subpolar habitats (clade I) grew more slowly but was able to cope with temperatures <10 °C. We show that growth at such temperatures was accompanied by a large increase of the photoprotection capacities using the orange carotenoid protein (OCP). Metagenomic analyzes revealed that Synechococcus natural communities show the highest prevalence of the ocp genes in low-temperature niches, whereas most tropical clade II Synechococcus have lost the gene. Moreover, bioinformatic analyzes suggested that the OCP variants of the two cold-adapted Synechococcus clades I and IV have undergone evolutionary convergence through the adaptation of the molecular flexibility. Our study points to an important role of temperature in the evolution of the OCP. We, furthermore, discuss the implications of the different metabolic cost of these physiological strategies on the competitiveness of Synechococcus in a warming ocean. This study can help improve the current hypotheses and models aimed at predicting the changes in ocean carbon fluxes in response to global warming.

Pili allow dominant marine cyanobacteria to avoid sinking and evade predation.

How oligotrophic marine cyanobacteria position themselves in the water column is currently unknown. The current paradigm is that these organisms avoid sinking due to their reduced size and passive drift within currents. Here, we show that one in four picocyanobacteria encode a type IV pilus which allows these organisms to increase drag and remain suspended at optimal positions in the water column, as well as evade predation by grazers. The evolution of this sophisticated floatation mechanism in these purely planktonic streamlined microorganisms has important implications for our current understanding of microbial distribution in the oceans and predator-prey interactions which ultimately will need incorporating into future models of marine carbon flux dynamics.

A Cyanobacterial Component Required for Pilus Biogenesis Affects the Exoproteome.

Protein secretion as well as the assembly of bacterial motility appendages are central processes that substantially contribute to fitness and survival. This study highlights distinctive features of the mechanism that serves these functions in cyanobacteria, which are globally prevalent photosynthetic prokaryotes that significantly contribute to primary production. Our studies of biofilm development in the cyanobacterium Synechococcus elongatus uncovered a novel component required for the biofilm self-suppression mechanism that operates in this organism. This protein, which is annotated as "hypothetical," is denoted EbsA (essential for biofilm self-suppression A) here. EbsA homologs are highly conserved and widespread in diverse cyanobacteria but are not found outside this clade. We revealed a tripartite complex of EbsA, Hfq, and the ATPase homolog PilB (formerly called T2SE) and demonstrated that each of these components is required for the assembly of the hairlike type IV pili (T4P) appendages, for DNA competence, and affects the exoproteome in addition to its role in biofilm self-suppression. These data are consistent with bioinformatics analyses that reveal only a single set of genes in S. elongatus to serve pilus assembly or protein secretion; we suggest that a single complex is involved in both processes. A phenotype resulting from the impairment of the EbsA homolog in the cyanobacterium Synechocystis sp. strain PCC 6803 implies that this feature is a general cyanobacterial trait. Moreover, comparative exoproteome analyses of wild-type and mutant strains of S. elongatus suggest that EbsA and Hfq affect the exoproteome via a process that is independent of PilB, in addition to their involvement in a T4P/secretion machinery.IMPORTANCE Cyanobacteria, environmentally prevalent photosynthetic prokaryotes, contribute ∼25% of global primary production. Cyanobacterial biofilms elicit biofouling, thus leading to substantial economic losses; however, these microbial assemblages can also be beneficial, e.g., in wastewater purification processes and for biofuel production. Mechanistic aspects of cyanobacterial biofilm development were long overlooked, and genetic and molecular information emerged only in recent years. The importance of this study is 2-fold. First, it identifies novel components of cyanobacterial biofilm regulation, thus contributing to the knowledge of these processes and paving the way for inhibiting detrimental biofilms or promoting beneficial ones. Second, the data suggest that cyanobacteria may employ the same complex for the assembly of the motility appendages, type 4 pili, and protein secretion. A shared pathway was previously shown in only a few cases of heterotrophic bacteria, whereas numerous studies demonstrated distinct systems for these functions. Thus, our study broadens the understanding of pilus assembly/secretion in diverse bacteria and furthers the aim of controlling the formation of cyanobacterial biofilms.

The structural basis of far-red light absorbance by allophycocyanins.

Phycobilisomes (PBS), the major light-harvesting antenna in cyanobacteria, are supramolecular complexes of colorless linkers and heterodimeric, pigment-binding phycobiliproteins. Phycocyanin and phycoerythrin commonly comprise peripheral rods, and a multi-cylindrical core is principally assembled from allophycocyanin (AP). Each AP subunit binds one phycocyanobilin (PCB) chromophore, a linear tetrapyrrole that predominantly absorbs in the orange-red region of the visible spectrum (600-700 nm). AP facilitates excitation energy transfer from PBS peripheral rods or from directly absorbed red light to accessory chlorophylls in the photosystems. Paralogous forms of AP that bind PCB and are capable of absorbing far-red light (FRL; 700-800 nm) have recently been identified in organisms performing two types of photoacclimation: FRL photoacclimation (FaRLiP) and low-light photoacclimation (LoLiP). The FRL-absorbing AP (FRL-AP) from the thermophilic LoLiP strain Synechococcus sp. A1463 was chosen as a platform for site-specific mutagenesis to probe the structural differences between APs that absorb in the visible region and FRL-APs and to identify residues essential for the FRL absorbance phenotype. Conversely, red light-absorbing allophycocyanin-B (AP-B; ~ 670 nm) from the same organism was used as a platform for creating a FRL-AP. We demonstrate that the protein environment immediately surrounding pyrrole ring A of PCB on the alpha subunit is mostly responsible for the FRL absorbance of FRL-APs. We also show that interactions between PCBs bound to alpha and beta subunits of adjacent protomers in trimeric AP complexes are responsible for a large bathochromic shift of about ~ 20 nm and notable sharpening of the long-wavelength absorbance band.

Proximity-based proteomics reveals the thylakoid lumen proteome in the cyanobacterium Synechococcus sp. PCC 7002.

Cyanobacteria possess unique intracellular organization. Many proteomic studies have examined different features of cyanobacteria to learn about the intracellular structures and their respective functions. While these studies have made great progress in understanding cyanobacterial physiology, the conventional fractionation methods used to purify cellular structures have limitations; specifically, certain regions of cells cannot be purified with existing fractionation methods. Proximity-based proteomics techniques were developed to overcome the limitations of biochemical fractionation for proteomics. Proximity-based proteomics relies on spatiotemporal protein labeling followed by mass spectrometry of the labeled proteins to determine the proteome of the region of interest. We performed proximity-based proteomics in the cyanobacterium Synechococcus sp. PCC 7002 with the APEX2 enzyme, an engineered ascorbate peroxidase. We determined the proteome of the thylakoid lumen, a region of the cell that has remained challenging to study with existing methods, using a translational fusion between APEX2 and PsbU, a lumenal subunit of photosystem II. Our results demonstrate the power of APEX2 as a tool to study the cell biology of intracellular features and processes, including photosystem II assembly in cyanobacteria, with enhanced spatiotemporal resolution.

Spatial and temporal variations in Synechococcus microdiversity in the Southern California coastal ecosystem.

The Synechococcus cyanobacterial population at the Scripps Institution of Oceanography pier in La Jolla, CA, shows large increases in abundance, typically in the spring and summer followed, by rapid declines within weeks. Here we used amplicon sequencing of the ribosomal RNA internal transcribed spacer region to examine the microdiversity within this cyanobacterial genus during these blooms as well as further offshore in the Southern California coastal ecosystem (CCE). These analyses revealed numerous Synechococcus amplicon sequence variants (ASVs) and that clade and ASV composition can change over the course of blooms. We also found that a large bloom in August 2016 was highly anomalous both in its overall Synechococcus abundance and in terms of the presence of normally oligotrophic Synechococcus clade II. The dominant ASVs at the pier were found further offshore and in the California Current, but we did observe more oligotrophic ASVs and clades along with depth variation in Synechococcus diversity. We also observed that the dominant sequence variant switched during the peak of multiple Synechococcus blooms, with this switch occurring in multiple clades, but we present initial evidence that this apparent ASV switch is a physiological response rather than a change in the dominant population.

Involvement of glycogen metabolism in circadian control of UV resistance in cyanobacteria.

Most organisms harbor circadian clocks as endogenous timing systems in order to adapt to daily environmental changes, such as exposure to ultraviolet (UV) light. It has been hypothesized that the circadian clock evolved to prevent UV-sensitive activities, such as DNA replication and cell division, during the daytime. Indeed, circadian control of UV resistance has been reported in several eukaryotic organisms, from algae to higher organisms, although the underlying mechanisms remain unknown. Here, we demonstrate that the unicellular cyanobacterium Synechococcus elongatus PCC 7942 exhibits a circadian rhythm in resistance to UV-C and UV-B light, which is higher during subjective dawn and lower during subjective dusk. Nullification of the clock gene cluster kaiABC or the DNA-photolyase phr abolished rhythmicity with constitutively lower resistance to UV-C light, and amino acid substitutions of KaiC altered the period lengths of the UV-C resistance rhythm. In order to elucidate the molecular mechanism underlying the circadian regulation of UV-C resistance, transposon insertion mutants that alter UV-C resistance were isolated. Mutations to the master circadian output mediator genes sasA and rpaA and the glycogen degradation enzyme gene glgP abolished circadian rhythms of UV-C resistance with constitutively high UV-C resistance. Combining these results with further experiments using ATP synthesis inhibitor and strains with modified metabolic pathways, we showed that UV-C resistance is weakened by directing more metabolic flux from the glycogen degradation to catabolic pathway such as oxidative pentose phosphate pathway and glycolysis. We suggest glycogen-related metabolism in the dark affects circadian control in UV sensitivity, while the light masks this effect through the photolyase function.

Co-culture with Synechococcus facilitates growth of Prochlorococcus under ocean acidification conditions.

Anthropogenic CO2 emissions are projected to lower the pH of the ocean 0.3 units by 2100. Previous studies suggested that Prochlorococcus and Synechococcus, the numerically dominant phytoplankton in the oceans, have different responses to elevated CO2 that may result in a dramatic shift in their relative abundances in future oceans. Here we showed that the exponential growth rates of these two genera respond to future CO2 conditions in a manner similar to other cyanobacteria, but Prochlorococcus strains had significantly lower realized growth rates under elevated CO2 regimes due to poor survival after exposure to fresh culture media. Despite this, a Synechococcus strain was unable to outcompete a Prochlorococcus strain in co-culture at elevated CO2 . Under these conditions, Prochlorococcus' poor response to elevated CO2 disappeared, and Prochlorococcus' relative fitness showed negative frequency dependence, with both competitors having significant fitness advantages when initially rare. These experiments suggested that the two strains should be able to coexist indefinitely in co-culture despite sharing nearly identical nutritional requirements. We speculate that negative frequency dependence exists due to reductive Black Queen evolution that has resulted in a passively mutualistic relationship analogous to that connecting Prochlorococcus with the 'helper' heterotrophic microbes in its environment.

Solution NMR structure of Se0862, a highly conserved cyanobacterial protein involved in biofilm formation.

Biofilms are accumulations of microorganisms embedded in extracellular matrices that protect against external factors and stressful environments. Cyanobacterial biofilms are ubiquitous and have potential for treatment of wastewater and sustainable production of biofuels. But the underlying mechanisms regulating cyanobacterial biofilm formation are unclear. Here, we report the solution NMR structure of a protein, Se0862, conserved across diverse cyanobacterial species and involved in regulation of biofilm formation in the cyanobacterium Synechococcus elongatus PCC 7942. Se0862 is a class α+ß protein with ααßßßßαα topology and roll architecture, consisting of a four-stranded ß-sheet that is flanked by four α-helices on one side. Conserved surface residues constitute a hydrophobic pocket and charged regions that are likely also present in Se0862 orthologs.

ß-Ν-Methylamino-L-alanine interferes with nitrogen assimilation in the cyanobacterium, non-BMAA producer, Synechococcus sp. TAU-MAC 0499.

The production of ß-Ν-methylamino-L-alanine (BMAA) in cyanobacteria is triggered by nitrogen-starvation conditions and its biological role, albeit unknown, is associated with nitrogen assimilation. In the present study, the effect of BMAA (773 µg L-1) on nitrogen metabolism and physiology of the non-diazotrophic cyanobacterium and non-BMAA producer, Synechococcus sp. TAU-MAC 0499, was investigated. In order to study the combined effect of nitrogen availability and BMAA, nitrogen-starvation conditions were induced by transferring cells in nitrogen-free medium and subsequently exposing the cultures to BMAA. After short-term treatment (180 min) and in the presence of nitrogen, BMAA inhibited glutamine synthetase, which resulted in low concentration of glutamine. In the absence of nitrogen, although there was no effect on glutamine synthetase, a possible perturbation in nitrogen assimilation is reflected on the significant decrease in glutamate levels. During the long-term exposure (24-96 h), growth, photosynthetic pigments and total protein were not affected by BMAA exposure, except for an increase in protein and phycocyanin levels at 48 h in nitrogen replete conditions. Results suggest that BMAA interferes with nitrogen assimilation, in a different way, depending on the presence or absence of combined nitrogen, providing novel data on the potential biological role of BMAA.

The response of Synechococcus sp. PCC 7002 to micro-/nano polyethylene particles - Investigation of a key anthropogenic stressor.

Microplastics or plastic particles less than 5 mm in size are a ubiquitous and damaging pollutant in the marine environment. However, the interactions between these plastic particles and marine microorganisms are just starting to be understood. The objective of this study was to measure the responses of a characteristic marine organism (Synechococcus sp. PCC 7002) to an anthropogenic stressor (polyethelene nanoparticles and microparticles) using molecular techniques. This investigation showed that polyethylene microparticles and nanoparticles have genetic, enzymatic and morphological effects on Synechococcus sp. PCC 7002. An RT-PCR analysis showed increases in the expression of esterase and hydrolase genes at 5 days of exposure to polyethylene nanoparticles and at 10 days of exposure to polyethylene microparticles. A qualitative enzymatic assay also showed esterase activity in nanoparticle exposed samples. Cryo-scanning electron microscopy was used to assess morphological changes in exopolymer formation resulting from exposure to polyethylene microparticles and nanoparticles. The data from this paper suggests that microplastic and nanoplastics could be key microbial stressors and should be investigated in further detail.

Structural variability, coordination and adaptation of a native photosynthetic machinery.

Cyanobacterial thylakoid membranes represent the active sites for both photosynthetic and respiratory electron transport. We used high-resolution atomic force microscopy to visualize the native organization and interactions of photosynthetic complexes within the thylakoid membranes from the model cyanobacterium Synechococcus elongatus PCC 7942. The thylakoid membranes are heterogeneous and assemble photosynthetic complexes into functional domains to enhance their coordination and regulation. Under high light, the chlorophyll-binding proteins IsiA are strongly expressed and associate with Photosystem I (PSI), forming highly variable IsiA-PSI supercomplexes to increase the absorption cross-section of PSI. There are also tight interactions of PSI with Photosystem II (PSII), cytochrome b6f, ATP synthase and NAD(P)H dehydrogenase complexes. The organizational variability of these photosynthetic supercomplexes permits efficient linear and cyclic electron transport as well as bioenergetic regulation. Understanding the organizational landscape and environmental adaptation of cyanobacterial thylakoid membranes may help inform strategies for engineering efficient photosynthetic systems and photo-biofactories.

Methylation deficiency disrupts biological rhythms from bacteria to humans.

The methyl cycle is a universal metabolic pathway providing methyl groups for the methylation of nuclei acids and proteins, regulating all aspects of cellular physiology. We have previously shown that methyl cycle inhibition in mammals strongly affects circadian rhythms. Since the methyl cycle and circadian clocks have evolved early during evolution and operate in organisms across the tree of life, we sought to determine whether the link between the two is also conserved. Here, we show that methyl cycle inhibition affects biological rhythms in species ranging from unicellular algae to humans, separated by more than 1 billion years of evolution. In contrast, the cyanobacterial clock is resistant to methyl cycle inhibition, although we demonstrate that methylations themselves regulate circadian rhythms in this organism. Mammalian cells with a rewired bacteria-like methyl cycle are protected, like cyanobacteria, from methyl cycle inhibition, providing interesting new possibilities for the treatment of methylation deficiencies.

Damped circadian oscillation in the absence of KaiA in Synechococcus.

Proteins KaiA, KaiB and KaiC constitute a biochemical circadian oscillator in the cyanobacterium Synechococcus elongatus. It has been reported kaiA inactivation completely abolishes circadian oscillations. However, we show here that kaiBC promoter activity exhibits a damped, low-amplitude oscillation with a period of approximately 24 h in kaiA-inactivated strains. The damped rhythm resonates with external cycles with a period of 24-26 h, indicating that its natural frequency is similar to that of the circadian clock. Double-mutation experiments reveal that kaiC, kaiB, and sasA (encoding a KaiC-binding histidine kinase) are all required for the damped oscillation. Further analysis suggests that the kaiA-less damped transcriptional rhythm requires KaiB-KaiC complex formation and the transcription-translation feedback loop, but not the KaiC phosphorylation cycle. Our results provide insights into mechanisms that could potentially underlie the diurnal/circadian behaviors observed in other bacterial species that possess kaiB and kaiC homologues but lack a kaiA homologue.

Synechococcus elongatus PCC7942: a cyanobacterium cell factory for producing useful chemicals and fuels under abiotic stress conditions.

Sucrose, a compatible osmolyte in cyanobacteria, functions both as an energy reserve and as osmoprotectant. Sugars are the most common substrates used by microorganisms to produce hydrogen (H2) by means of anaerobic dark fermentation. Cells of the unicellular, non-nitrogen fixing, freshwater cyanobacterium Synechococcus elongatus PCC7942 accumulate sucrose under salt stress. In the present work, we used this cyanobacterium and a genetically engineered strain of it (known as PAMCOD) to investigate the optimal conditions for (a) photosynthetic activity, (b) cell proliferation and (c) sucrose accumulation, which are necessary for H2 production via anaerobic dark fermentation of the accumulated sucrose. PAMCOD (Deshnium et al. in Plant Mol Biol 29:897-902, 1995) contains the gene codA that codes for choline oxidase, the enzyme which converts choline to the zwitterion glycine betaine. Glycine betaine is a compatible osmolyte which increases the salt tolerance of Synechococcus elongatus PCC7942. Furthermore, glycine betaine maintains cell proliferation under salt stress and results in increased sucrose accumulation. In the present study, we examine the environmental factors, such as the NaCl concentration, the culture medium pH, and the carbon dioxide content of the air bubbled through it. At optimal conditions, sucrose accumulated in the cyanobacteria cells up to 13.5 mol per mole Chl a. Overall, genetically engineered Synechococcus elongatus PCC7942 produces sucrose in sufficient quantities such that it may be a viable alternative (a) to sucrose synthesis, and (b) to H2 formation via anaerobic dark fermentation.

Phytoplankton transcriptomic and physiological responses to fixed nitrogen in the California current system.

Marine phytoplankton are responsible for approximately half of photosynthesis on Earth. However, their ability to drive ocean productivity depends on critical nutrients, especially bioavailable nitrogen (N) which is scarce over vast areas of the ocean. Phytoplankton differ in their preferences for N substrates as well as uptake efficiencies and minimal N requirements relative to other critical nutrients, including iron (Fe) and phosphorus. In this study, we used the MicroTOOLs high-resolution environmental microarray to examine transcriptomic responses of phytoplankton communities in the California Current System (CCS) transition zone to added urea, ammonium, nitrate, and also Fe in the late summer when N depletion is common. Transcript level changes of photosynthetic, carbon fixation, and nutrient stress genes indicated relief of N limitation in many strains of Prochlorococcus, Synechococcus, and eukaryotic phytoplankton. The transcriptomic responses helped explain shifts in physiological and growth responses observed later. All three phytoplankton groups had increased transcript levels of photosynthesis and/or carbon fixation genes in response to all N substrates. However, only Prochlorococcus had decreased transcript levels of N stress genes and grew substantially, specifically after urea and ammonium additions, suggesting that Prochlorococcus outcompeted other community members in these treatments. Diatom transcript levels of carbon fixation genes increased in response to Fe but not to Fe with N which might have favored phytoplankton that were co-limited by N and Fe. Moreover, transcription patterns of closely related strains indicated variability in N utilization, including nitrate utilization by some high-light adapted Prochlorococcus. Finally, up-regulation of urea transporter genes by both Prochlorococcus and Synechococcus in response to filtered deep water suggested a regulatory mechanism other than classic control via the global N regulator NtcA. This study indicated that co-existing phytoplankton strains experience distinct nutrient stresses in the transition zone of the CCS, an understudied region where oligotrophic and coastal communities naturally mix.