Photosynthetic organisms have developed sophisticated strategies to fine-tune light energy conversion to meet the metabolic demand, thereby optimizing growth in fluctuating light environments. Although mechanisms such as energy dissipation, photosynthetic control, or the photosystem II (PSII) damage and repair have been widely studied, little is known about the regulation of protein synthesis capacity during light acclimation. By screening a Chlamydomonas reinhardtii insertional mutant library using chlorophyll fluorescence imaging, we isolated a high chlorophyll fluorescence mutant (hf0) defected in a gene encoding a putative plastid targeted DEAD-box RNA helicase called CreRH22. CreRH22 is rapidly induced upon illumination and belongs to the GreenCut, a set of proteins specific to photosynthetic organisms. While photosynthesis is slightly affected in the mutant under low light (LL), exposure to high light (HL) induces a marked decrease in both PSII and PSI, and a strong alteration of the light-induced gene expression pattern. These effects are explained by the inability of hf0 to increase plastid ribosome amounts under HL. We conclude that CreRH22, by promoting ribosomal RNA precursor maturation in a light-dependent manner, enables the assembly of extra ribosomes required to synthesize photosystem subunits at a higher rate, a critical step in the acclimation of algae to HL.

The maize female gametophyte is comprised of four cell types: two synergids, an egg cell, a central cell, and a variable number of antipodal cells. In maize, these cells are produced after three rounds of free-nuclear divisions followed by cellularization, differentiation, and proliferation of the antipodal cells. Cellularization of the eight-nucleate syncytium produces seven cells with two polar nuclei in the central cell. Nuclear localization is tightly controlled in the embryo sac as evidenced by the regular, stereotypical position of the nuclei in all syncytial stages of female gametophyte development. This leads to precise allocation of the nuclei into the cells upon cellularization. Nuclear positioning within the syncytium is highly correlated with their identity after cellularization. Two mutants are described with extra polar nuclei, abnormal antipodal cell morphology, and reduced antipodal cell number, which is correlated with a frequent loss of auxin signaling in the antipodal cell cluster. Mutations in one of these genes, indeterminate gametophyte2 encoding a MICROTUBULE ASSOCIATED PROTEIN65-3 homolog, shows a requirement for MAP65-3 in cellularization of the syncytial embryo sac and that the identity of the nuclei in the syncytial female gametophyte can be changed very late before cellularization.

Photosynthetic algae cope with suboptimal levels of light and CO2. In low CO2 and excess light, the green alga Chlamydomonas reinhardtii activates a CO2 Concentrating Mechanism (CCM) and photoprotection; the latter is mediated by LHCSR1/3 and PSBS. How light and CO2 signals converge to regulate photoprotective responses remains unclear. Here we show that excess light activates expression of photoprotection-and CCM-related genes and that depletion of CO2 drives these responses, even in total darkness. High CO2 levels, derived from respiration or impaired photosynthetic fixation, repress LHCSR3 and CCM genes while stabilizing the LHCSR1 protein. We also show that CIA5, which controls CCM genes, is a major regulator of photoprotection, elevating LHCSR3 and PSBS transcript accumulation while inhibiting LHCSR1 accumulation. Our work emphasizes the importance of CO2 in regulating photoprotection and the CCM, demonstrating that the impact of light on photoprotection is often indirect and reflects intracellular CO2 levels.

In nature, photosynthetic organisms are exposed to different light spectra and intensities depending on the time of day and atmospheric and environmental conditions. When photosynthetic cells absorb excess light, they induce non-photochemical quenching to avoid photo-damage and trigger expression of ‘photoprotective’ genes. In this work, we used the green alga Chlamydomonas reinhardtii to assess the impact of light intensity, light quality, wavelength, photosynthetic electron transport and CO2 on induction of the ‘photoprotective’ genes (LHCSR1, LHCSR3 and PSBS) during dark-to-light transitions. Induction (mRNA accumulation) occurred at very low light intensity, was independently modulated by blue and UV-B radiation through specific photoreceptors, and only LHCSR3 was strongly controlled by CO2 levels through a putative enhancer function of CIA5, a transcription factor that controls genes of the carbon concentrating mechanism. We propose a model that integrates inputs of independent signaling pathways and how they may help the cells anticipate diel conditions and survive in a dynamic light environment.

Photosynthetic organisms frequently experience abiotic stresses that restrict their growth and development. Under such circumstances, most absorbed solar energy cannot be used for CO2 fixation and can cause the photoproduction of reactive oxygen species (ROS) that can damage the photosynthetic reaction centers, photosystems I and II (PSI and PSII), resulting in a decline in primary productivity. This work describes a biological ‘switch’ in the green alga Chlamydomonas reinhardtii that reversibly restricts photosynthetic electron transport (PET) at the cytochrome b6f complex when reductant and ATP generated by PET are in excess of the capacity of carbon metabolism to utilize these products; we specifically show a restriction at this switch when sta6 mutant cells, which cannot synthesize starch, are limited for nitrogen (growth inhibition) and subjected to a dark-to-light transition. This restriction, which may be a form of photosynthetic control, causes diminished electron flow to PSI, which prevents PSI photodamage. When electron flow is blocked the plastid alternative oxidase (PTOX) may also become activated, functioning as an electron valve that dissipates some of the excitation energy absorbed by PSII thereby lessening PSII photoinhibition. Furthermore, illumination of the cells following the dark acclimation gradually diminishes the restriction at cytochrome b6f complex. Elucidating this photoprotective mechanism and its modulating factors may offer new insights into mechanisms associated with photosynthetic control and offer new directions for optimizing photosynthesis.

Photosynthetic eukaryotic organisms contain several chloroplast-associated metabolite transporters that enable energetic/metabolic exchange between the chloroplast and other cellular compartments. In this study, we used the model photosynthetic alga Chlamydomonas reinhardtii to investigate a highly expressed chloroplast triose phosphate transporter. The triose phosphate/phosphate translocator 3 (CreTPT3), located on the Chlamydomonas chloroplast envelope, was found to be highly expressed under both non-stressed/stressed conditions (RNA level) and was characterized for substrate specificity in vitro using a yeast liposome uptake system. The CreTPT3 transporter showed high DHAP and 3-PGA transport activities, but little activity with PEP. Null mutants for CreTPT3, generated by CRISPR-Cas9 editing of the CreTPT3 gene, resulted in a pleiotropic phenotype impacting photosynthetic activity, metabolite pools, carbon partitioning, and storage, the redox status of the chloroplast, and the accumulation of reactive oxygen species. The results presented demonstrate that CreTPT3 is a major conduit on the chloroplast envelope for the intracellular distribution of fixed carbon and reductant generated by photosynthetic electron transport. Its function is critical for optimizing the use of resources supporting cell fitness, especially as light intensities increase, the rate of photosynthetic CO2 fixation is elevated and the chloroplast environment becomes highly reducing.

Photosynthetic organisms frequently experience abiotic stress that restricts their growth and development. Under such circumstances, most absorbed solar energy cannot be used for CO2 fixation and can cause the photoproduction of reactive oxygen species (ROS) that can damage the photosynthetic reaction centers of photosystem I and II (PSI and PSII), resulting in a decline in primary productivity. This work describes a biological 'switch' in the green alga Chlamydomonas reinhardtii that reversibly restricts photosynthetic electron transport (PET) at the cytochrome b6f (Cyt b6f) complex when the capacity for accepting electrons downstream of PSI is severely limited. We specifically show this restriction in STARCHLESS6 (sta6) mutant cells, which cannot synthesize starch when they are limited for nitrogen (growth inhibition) and subjected to a dark-to-light transition. This restriction represents a form of photosynthetic control that causes diminished electron flow to PSI and thereby prevents PSI photodamage but does not appear to rely on a DeltapH. Furthermore, when electron flow is restricted, the plastid alternative oxidase (PTOX) becomes active, functioning as an electron valve that dissipates some excitation energy absorbed by PSII and allows the formation of a proton motive force (PMF) that would drive some ATP production [potentially sustaining PSII repair and non-photochemical quenching (NPQ)]. The restriction at the Cyt b6f complex can be gradually relieved with continued illumination. This study provides insights into how photosynthetic electron transport responds to a marked reduction in availability of downstream electron acceptors and the protective mechanisms involved.

Modulation of photoassimilate export from the chloroplast is essential for controlling the distribution of fixed carbon in the cell and maintaining optimum photosynthetic rates. In this study we identified chloroplast TRIOSE PHOSPHATE/PHOSPHATE TRANSLOCATOR2 (CreTPT2) and CreTPT3 in the green alga Chlamydomonas (Chlamydomonas reinhardtii), which exhibit similar substrate specificities but whose encoding genes are differentially expressed over the diurnal cycle. We focused mostly on CreTPT3 because of its high level of expression and the severe phenotype exhibited by tpt3 relative to tpt2 mutants. Null mutants for CreTPT3 had a pleiotropic phenotype that affected growth, photosynthetic activities, metabolite profiles, carbon partitioning, and organelle-specific accumulation of H2O2. These analyses demonstrated that CreTPT3 is a dominant conduit on the chloroplast envelope for the transport of photoassimilates. In addition, CreTPT3 can serve as a safety valve that moves excess reductant out of the chloroplast and appears to be essential for preventing cells from experiencing oxidative stress and accumulating reactive oxygen species, even under low/moderate light intensities. Finally, our studies indicate subfunctionalization of the CreTPT transporters and suggest that there are differences in managing the export of photoassimilates from the chloroplasts of Chlamydomonas and vascular plants.

Controlling the transition from a multicellular motile state to a sessile biofilm is an important eco-physiological decision for most prokaryotes, including cyanobacteria. Photosynthetic and bio geochemically significant cyanobacterium Synechocystis sp. PCC6803 (Syn6803) uses Type IV pili (TFP) for surface-associated motility and light-directed phototaxis. We report the identification of a novel Chaperone-Usher (CU) system in Syn6803 that regulate secretion of minor pilins as a means of stabilizing TFP morphology. These secreted minor-pilins aid in modifying TFP morphology to suit the adhesion state by forming cell to surface contacts when motility is not required. This morphotype is structurally distinct from TFP assembled during motile phase. We further demonstrate by examining mutants lacking either the CU system or the minor-pilins, which produce aberrant TFP, that are morphologically and functionally distinct from wild-type (WT). Thus, here we report that in Syn6803, CU system work independent of TFP biogenesis machinery unlike reported for other pathogenic bacterial systems and contributes to provide multifunctional plasticity to TFP. cAMP levels play an important role in controlling this switch. This phenotypic plasticity exhibited by the TFP, in response to cAMP levels would allow cells and cellular communities to adapt to rapidly fluctuating environments by dynamically transitioning between motile and sessile states.Significance of this workHow cyanobacterial communities cope with fluctuating or extreme environments is crucial in understanding their role in global carbon and nitrogen cycles. This work addresses the key question: how do cyanobacteria modulate external appendages, called Type IV pili, to effectively switch between motile and sessile biofilm states? We demonstrate that cells transition between forming strong cell-surface interactions indispensable for biofilm formation to forming cell-cell interactions that allow for coordinated movement crucial for social motility by functional/ structural modification of same TFP appendage. The second messenger, cAMP and a Chaperone-Usher secretion are indispensible to achieve these structural modifications of TFP and control the complex phenotypic transition. We have uncovered a strategy that Syn6803 has evolved to deal with molecular decision-making under uncertainty, which we call phenotypic plasticity. Here we demonstrate how a single motility appendage can be structurally modified to attain two antagonistic functions in order to meet the fluctuating environmental demands.

High-throughput RNA sequencing offers unprecedented opportunities to explore the Earth RNA virome. Mining 5,150 diverse metatranscriptomes uncovered >2.5 million RNA viral contigs. Via analysis of the 330k novel RNA-dependent RNA polymerases (RdRP), this expansion corresponds to a five-fold increase of RNA virus diversity. Extended RdRP phylogeny supports monophyly of the five established phyla, reveals two putative new bacteriophage phyla and numerous putative novel classes and orders. The dramatically expanded Lenarviricota phylum, consisting of bacterial and related eukaryotic viruses, now accounts for a third of the RNA virome diversity. Identification of CRISPR spacer matches and bacteriolytic proteins suggests that subsets of picobirnaviruses and partitiviruses, previously associated with eukaryotes, infect prokaryotic hosts. Gene content analysis revealed multiple domains previously not found in RNA viruses and implicated in virus-host interactions. This vast collection of new RNA virus genomes provides insights into RNA virus evolution and should become a major resource for RNA virology.