Hydrogen photoproduction by eukaryotic microalgae results from a connection between the photosynthetic electron transport chain and a plastidial hydrogenase. Algal H₂ production is a transitory phenomenon under most natural conditions, often viewed as a safety valve protecting the photosynthetic electron transport chain from overreduction. From the colony screening of an insertion mutant library of the unicellular green alga Chlamydomonas reinhardtii based on the analysis of dark-light chlorophyll fluorescence transients, we isolated a mutant impaired in cyclic electron flow around photosystem I (CEF) due to a defect in the Proton Gradient Regulation Like1 (PGRL1) protein. Under aerobiosis, nonphotochemical quenching of fluorescence (NPQ) is strongly decreased in pgrl1. Under anaerobiosis, H₂ photoproduction is strongly enhanced in the pgrl1 mutant, both during short-term and long-term measurements (in conditions of sulfur deprivation). Based on the light dependence of NPQ and hydrogen production, as well as on the enhanced hydrogen production observed in the wild-type strain in the presence of the uncoupling agent carbonyl cyanide p-trifluoromethoxyphenylhydrazone, we conclude that the proton gradient generated by CEF provokes a strong inhibition of electron supply to the hydrogenase in the wild-type strain, which is released in the pgrl1 mutant. Regulation of the trans-thylakoidal proton gradient by monitoring pgrl1 expression opens new perspectives toward reprogramming the cellular metabolism of microalgae for enhanced H₂ production.

Cytosolic/nuclear molecular chaperones of the heat shock protein families HSP90 and HSC70 are conserved and essential proteins in eukaryotes. These proteins have essentially been implicated in the innate immunity and abiotic stress tolerance in higher plants. Here, we demonstrate that both chaperones are recruited in Arabidopsis (Arabidopsis thaliana) for stomatal closure induced by several environmental signals. Plants overexpressing HSC70-1 or with reduced HSP90.2 activity are compromised in the dark-, CO2-, flagellin 22 peptide-, and abscisic acid (ABA)-induced stomatal closure. HSC70-1 and HSP90 proteins are needed to establish basal expression levels of several ABA-responsive genes, suggesting that these chaperones might also be involved in ABA signaling events. Plants overexpressing HSC70-1 or with reduced HSP90.2 activity are hypersensitive to ABA in seed germination assays, suggesting that several chaperone complexes with distinct substrates might tune tissue-specific responses to ABA and the other biotic and abiotic stimuli studied. This study demonstrates that the HSC70/HSP90 machinery is important for stomatal closure and serves essential functions in plants to integrate signals from their biotic and abiotic environments.