A Chlamydomonas reinhardtii mutant lacking CGL71, a thylakoid membrane protein previously shown to be involved in photosystem I (PSI) accumulation, exhibited photosensitivity and highly reduced abundance of PSI under photoheterotrophic conditions. Remarkably, the PSI content of this mutant declined to nearly undetectable levels under dark, oxic conditions, demonstrating that reduced PSI accumulation in the mutant is not strictly the result of photodamage. Furthermore, PSI returns to nearly wild-type levels when the O-2 concentration in the medium is lowered. Overall, our results suggest that the accumulation of PSI in the mutant correlates with the redox state of the stroma rather than photodamage and that CGL71 functions under atmospheric O-2 conditions to allow stable assembly of PSI. These findings may reflect the history of the Earth's atmosphere as it transitioned from anoxic to highly oxic (1-2 billion years ago), a change that required organisms to evolve mechanisms to assist in the assembly and stability of proteins or complexes with O-2-sensitive cofactors.

The Plant Cell Atlas (PCA) community hosted a virtual symposium on December 9 and 10, 2021 on single cell and spatial omics technologies. The conference gathered almost 500 academic, industry, and government leaders to identify the needs and directions of the PCA community and to explore how establishing a data synthesis center would address these needs and accelerate progress. This report details the presentations and discussions focused on the possibility of a data synthesis center for a PCA and the expected impacts of such a center on advancing science and technology globally. Community discussions focused on topics such as data analysis tools and annotation standards; computational expertise and cyber-infrastructure; modes of community organization and engagement; methods for ensuring a broad reach in the PCA community; recruitment, training, and nurturing of new talent; and the overall impact of the PCA initiative. These targeted discussions facilitated dialogue among the participants to gauge whether PCA might be a vehicle for formulating a data synthesis center. The conversations also explored how online tools can be lever-aged to help broaden the reach of the PCA (i.e., online contests, virtual networking, and social media stakeholder engagement) and decrease costs of conducting research (e.g., virtual REU opportunities). Major recommendations for the future of the PCA included establishing standards, creating dashboards for easy and intuitive access to data, and engaging with a broad community of stakeholders. The discussions also identified the following as being essential to the PCA's success: identifying homologous cell-type markers and their biocuration, publishing datasets and computational pipelines, utilizing online tools for communication (such as Slack), and user-friendly data visualization and data sharing. In conclusion, the development of a data synthesis center will help the PCA community achieve these goals by providing a centralized repository for existing and new data, a platform for sharing tools, and new analytical approaches through collaborative, multidisciplinary efforts. A data synthesis center will help the PCA reach milestones, such as community-supported data evaluation metrics, accelerating plant research necessary for human and environmental health.

Porphyra is a macrophytic red alga of the Bangiales that is important ecologically and economically. We describe the genomes of three bacteria in the phylum Planctomycetes (designated P1, P2 and P3) that were isolated from blades of Porphyra umbilicalis (P. um. 1). These three Operational Taxonomic Units (OTUs) belong to distinct genera; P2 belongs to the genus Rhodopirellula, while P1 and P3 represent undescribed genera within the Planctomycetes. Comparative analyses of the P1, P2 and P3 genomes show large expansions of distinct gene families, which can be widespread throughout the Planctomycetes (e.g., protein kinases, sensors/response regulators) and may relate to specific habitat (e.g., sulfatase gene expansions in marine Planctomycetes) or phylogenetic position. Notably, there are major differences among the Planctomycetes in the numbers and sub-functional diversity of enzymes (e.g., sulfatases, glycoside hydrolases, polysaccharide lyases) that allow these bacteria to access a range of sulfated polysaccharides in macroalgal cell walls. These differences suggest that the microbes have varied capacities for feeding on fixed carbon in the cell walls of P. um. 1 and other macrophytic algae, although the activities among the various bacteria might be functionally complementary in situ. Additionally, phylogenetic analyses indicate augmentation of gene functions through expansions arising from gene duplications and horizontal gene transfers; examples include genes involved in cell wall degradation (e.g., kappa-carrageenase, alginate lyase, fucosidase) and stress responses (e.g., efflux pump, amino acid transporter). Finally P1 and P2 contain various genes encoding selenoproteins, many of which are enzymes that ameliorate the impact of environmental stresses that occur in the intertidal habitat.

Many microalgae acquire vitamin B-12 from marine prokaryotes. A new study demonstrates that vitamin B-12 is synthesized by planktonic cyanobacteria as pseudocobalamin, a form not bioactive in microalgae. However, some microalgae can remodel pseudocobalamin to the active cobalamin form, adding complexity to our assessment of active vitamin B-12 in the environment.

When photosynthetic organisms are deprived of nitrogen (N), the capacity to grow and assimilate carbon becomes limited, causing a decrease in the productive use of absorbed light energy and likely a rise in the cellular reduction state. Although there is a scarcity of N in many terrestrial and aquatic environments, a mechanistic understanding of how photosynthesis adjusts to low-N conditions and the enzymes/activities integral to these adjustments have not been described. In this work, we use biochemical and biophysical analyses of photoautotrophically grown wild-type and mutant strains of Chlamydomonas reinhardtii to determine the integration of electron transport pathways critical for maintaining active photosynthetic complexes even after exposure of cells to N deprivation for 3 d. Key to acclimation is the type II NADPH dehydrogenase, NDA2, which drives cyclic electron flow (CEF), chlororespiration, and the generation of an H+ gradient across the thylakoid membranes. N deprivation elicited a doubling of the rate of NDA2-dependent CEF, with little contribution from PGR5/PGRL1-dependent CEF. The H+ gradient generated by CEF is essential to sustain nonphotochemical quenching, while an increase in the level of reduced plastoquinone would promote a state transition; both are necessary to down-regulate photosystem II activity. Moreover, stimulation of NDA2-dependent chlororespiration affords additional relief from the elevated reduction state associated with N deprivation through plastid terminal oxidase-dependent water synthesis. Overall, rerouting electrons through the NDA2 catalytic hub in response to photoautotrophic N deprivation sustains cell viability while promoting the dissipation of excess excitation energy through quenching and chlororespiratory processes.

Paulinella chromatophora is a cercozoan amoeba that contains "chromatophores," which are photosynthetic inclusions of cyanobacterial origin. The recent discovery that chromatophores evolved independently of plastids, underwent major genome reduction, and transferred at least two genes to the host nucleus has highlighted P. chromatophora as a model to infer early steps in the evolution of photosynthetic organelles. However, owing to the paucity of nuclear genome sequence data, the extent of endosymbiotic gene transfer (EGT) and host symbiont regulation are currently unknown. A combination of 454 and Illumina next generation sequencing enabled us to generate a comprehensive reference transcriptome data set for P. chromatophora on which we mapped short Illumina cDNA reads generated from cultures from the dark and light phases of a diel cycle. Combined with extensive phylogenetic analyses of the deduced protein sequences, these data revealed that 1) about 0.3-0.8% of the nuclear genes were obtained by EGT compared with 11-14% in the Plantae, 2) transferred genes show a distinct bias in that many encode small proteins involved in photosynthesis and photoacclimation, 3) host cells established control over expression of transferred genes, and 4) not only EGT, but to a minor extent also horizontal gene transfer from organisms that presumably served as food sources, helped to shape the nuclear genome of P. chromatophora. The identification of a significant number of transferred genes involved in photosynthesis and photoacclimation of thylakoid membranes as well as the observed transcriptional regulation of these genes strongly implies import of the encoded gene products into chromatophores, a feature previously thought to be restricted to canonical organelles. Thus, a possible mechanism by which P. chromatophora exerts control over the performance of its newly acquired photosynthetic organelle may involve controlling the expression of nuclear-encoded chromatophore-targeted regulatory components of the thylakoid membranes.

A sensor histidine kinase of Synechococcus sp. strain PCC7942, designated nblS, was previously identified and shown to be critical for the acclimation of cells to high-light and nutrient limitation conditions and to influence the expression of a number of light-responsive genes. The nblS orthologue in Synechocystis sp. strain PCC6803 is designated dspA (also called hik33). We have generated a dspA null mutant and analyzed global gene expression in both the mutant and wild-type strains under high- and low-light conditions. The mutant is aberrant for the expression of many genes encoding proteins critical for photosynthesis, phosphate and carbon acquisition, and the amelioration of stress conditions. Furthermore, transcripts from a number of genes normally detected only during exposure of wild-type cells to high-light conditions become partially constitutive in the low-light-grown dspA mutant. Other genes for which transcripts decline upon exposure of wild-type cells to high light are already lower in the mutant during growth in low light. These results suggest that DspA may influence gene expression in both a positive and a negative manner and that the dspA mutant behaves as if it were experiencing stress conditions (e.g., high-light exposure) even when maintained at near-optimal growth conditions for wild-type cells. This is discussed with respect to the importance of DspA for regulating the responses of the cell to environmental cues.

The Chlamydomonas reinhardtii transcription factor PSR1 is required for the control of activities involved in scavenging phosphate from the environment during periods of phosphorus limitation. Increased scavenging activity reflects the development of high-affinity phosphate transport and the expression of extracellular phosphatases that can cleave phosphate from organic compounds in the environment. A comparison of gene expression patterns using microarray analyses and quantitative PCRs with wild-type and psr1 mutant cells deprived of phosphorus has revealed that PSR1 also controls genes encoding proteins with potential "electron valve" functions-these proteins can serve as alternative electron acceptors that help prevent photodamage caused by overexcitation of the photosynthetic electron transport system. In accordance with this finding, phosphorus-starved psr1 mutants die when subjected to elevated light intensities; at these intensities, the wild-type cells still exhibit rapid growth. Acclimation to phosphorus deprivation also involves a reduction in the levels of transcripts encoding proteins involved in photosynthesis and both cytoplasmic and chloroplast translation as well as an increase in the levels of transcripts encoding stress-associated chaperones and proteases. Surprisingly, phosphorus-deficient psr1 cells (but not wild-type cells) also display expression patterns associated with specific responses to sulfur deprivation, suggesting a hitherto unsuspected link between the signal transduction pathways involved in controlling phosphorus and sulfur starvation responses. Together, these results demonstrate that PSR1 is critical for the survival of cells under conditions of suboptimal phosphorus availability and that it plays a key role in controlling both scavenging responses and the ability of the cell to manage excess absorbed excitation energy.

Global photosynthesis consumes ten times more CO2 than net anthropogenic emissions, and microalgae account for nearly half of this consumption(1). The high efficiency of algal photosynthesis relies on a mechanism concentrating CO2 (CCM) at the catalytic site of the carboxylating enzyme RuBisCO, which enhances CO2 fixation(2). Although many cellular components involved in the transport and sequestration of inorganic carbon have been identified(3,4), how microalgae supply energy to concentrate CO2 against a thermodynamic gradient remains unknown(4-6). Here we show that in the green alga Chlamydomonas reinhardtii, the combined action of cyclic electron flow and O-2 photoreduction-which depend on PGRL1 and flavodiiron proteins, respectively-generate a low luminal pH that is essential for CCM function. We suggest that luminal protons are used downstream of thylakoid bestrophin-like transporters, probably for the conversion of bicarbonate to CO2. We further establish that an electron flow from chloroplast to mitochondria contributes to energizing non-thylakoid inorganic carbon transporters, probably by supplying ATP. We propose an integrated view of the network supplying energy to the CCM, and describe how algal cells distribute energy from photosynthesis to power different CCM processes. These results suggest a route for the transfer of a functional algal CCM to plants to improve crop productivity.

Responses of photosynthetic organisms to sulfur starvation include (i) increasing the capacity of the cell for transporting and/or assimilating exogenous sulfate, (ii) restructuring cellular features to conserve sulfur resources, and (iii) modulating metabolic processes and rates of cell growth and division. We used microarray analyses to obtain a genome-level view of changes in mRNA abundances in the green alga Chlamydomonas reinhardtii during sulfur starvation. The work confirms and extends upon previous findings showing that sulfur deprivation elicits changes in levels of transcripts for proteins that help scavenge sulfate and economize on the use of sulfur resources. Changes in levels of transcripts encoding members of the light-harvesting polypeptide family, such as LhcSR2, suggest restructuring of the photosynthetic apparatus during sulfur deprivation. There are also significant changes in levels of transcripts encoding enzymes involved in metabolic processes (e.g., carbon metabolism), intracellular proteolysis, and the amelioration of oxidative damage; a marked and sustained increase in mRNAs for a putative vanadium chloroperoxidase and a peroxiredoxin may help prolong survival of C. reinhardtii during sulfur deprivation. Furthermore, many of the sulfur stress-regulated transcripts (encoding polypeptides associated with sulfate uptake and assimilation, oxidative stress, and photosynthetic function) are not properly regulated in the sac1 mutant of C. reinhardtii, a strain that dies much more rapidly than parental cells during sulfur deprivation. Interestingly, sulfur stress elicits dramatic changes in levels of transcripts encoding putative chloroplast-localized chaperones in the sac1 mutant but not in the parental strain. These results suggest various strategies used by photosynthetic organisms during acclimation to nutrient-limited growth.