Loss-of-function mutations in ORGANELLE RNA RECOGNITION MOTIF PROTEIN6 (ORRM6) result in the near absence of RNA editing of psbF-C77 and the reduction in accD-C794 editing in Arabidopsis (Arabidopsis thaliana). The orrm6 mutants have decreased levels of photosystem II (PSII) proteins, especially PsbF, lower PSII activity, pale green pigmentation, smaller leaf and plant sizes, and retarded growth. Stable expression of ORRM6 rescues the orrm6 editing defects and mutant phenotype. Unlike ORRM1, the other known ORRM plastid editing factor, ORRM6, does not contain RNA editing interacting protein/multiple organellar RNA editing factor (RIP/MORF) boxes, which are required for ORRM1 to interact with site-specific pentatricopeptide repeat protein editing factors. ORRM6 interacts with RIP1/MORF8, RIP2/MORF2, and RIP9/MORF9, known components of RNA editosomes. While some plastid RRM proteins are involved in other forms of RNA processing and translation, the primary function of ORRM6 is evidently to mediate psbF-C77 editing, like the essential site-specific pentatricopeptide repeat protein LOW PSII ACCUMULATION66. Stable expression in the orrm6 mutants of a nucleus-encoded, plastid-targeted PsbF protein from a psbF gene carrying a T at nucleotide 77 significantly increases leaf and plant sizes, chlorophyll content, and PSII activity. These transformants demonstrate that plastid RNA editing can be bypassed through the expression of nucleus-encoded, edited forms of plastid genes.

Nitrogen-fixation-subunit-U (NFU)-type proteins have been shown to be involved in the biogenesis of iron-sulfur clusters. We investigated the molecular function of a chloroplastic NFU-type iron-sulfur scaffold protein, NFU3, in Arabidopsis (Arabidopsis thaliana) using genetics approaches. Loss-of-function mutations in the NFU3 gene caused yellow pigmentation in leaves, reductions in plant size, leaf size, and growth rate, delay in flowering and seeding, and decreases in seed production. Biochemical and physiological analyses indicated that these defects are due to the substantial reductions in the abundances of 4Fe-4S-containing photosystem I (PSI) core subunits PsaA (where Psa stands for PSI), PsaB, and PsaC and a nearly complete loss of PSI activity. In addition to the substantial decreases in the amounts of PSI core proteins, the content of 3Fe-4S-containing ferredoxin-dependent glutamine oxoglutarate aminotransferases declined significantly in the nfu3 mutants. Furthermore, the absorption spectrum of the recombinant NFU3 protein showed features characteristic of 4Fe-4S and 3Fe-4S clusters, and the in vitro reconstitution experiment indicated an iron-sulfur scaffold function of NFU3. These data demonstrate that NFU3 is involved in the assembly and transfer of 4Fe-4S and 3Fe-4S clusters and that NFU3 is required for the accumulation of 4Fe-4S-and 3Fe-4S-containing proteins, especially 4Fe-4S-containing PSI core subunits, in the Arabidopsis chloroplast.

The chloroplasts and mitochondria of photosynthetic eukaryotes contain proteins that are closely related to cytosolic Guided Entry of Tail-anchored proteins3 (Get3). Get3 is a targeting factor that efficiently escorts tail-anchored proteins to the ER. Because other components of the cytosolic targeting pathway appear to be absent in organelles, previous investigators have asserted that organellar Get3 homologs are unlikely to act as targeting factors. However, we show here both that the Arabidopsis thaliana chloroplast homolog designated as GET3B is structurally similar to cytosolic Get3 proteins and that it selectively binds a thylakoid-localized tail-anchored protein. Based on genetic interactions between a get3b mutation and mutations affecting the chloroplast signal recognition particle targeting pathway, as well as changes in the abundance of photosynthesis-related proteins in mutant plants, we propose that GET3B acts primarily to direct proteins to the thylakoids. Furthermore, through molecular complementation experiments, we show that function of GET3B depends on its ability to hydrolyze ATP, and this is consistent with action as a targeting factor. We propose that GET3B and related organellar Get3 homologs play a role that is analogous to that of cytosolic Get3 proteins, and that GET3B acts as a targeting factor in the chloroplast stroma to deliver tail-anchored proteins in a membrane-specific manner.

Photosynthetic species are subjected to a variety of environmental stresses, including suboptimal irradiance. In oxygenic photosynthetic organisms, a major effect of high light exposure is damage to the Photosystem II (PSII) reaction-center protein D1. This process even happens under low or moderate light. To cope with photodamage to D1, photosynthetic organisms evolved an intricate PSII repair and reassembly cycle, which requires the participation of different auxiliary proteins, including thiol/disulfide-modulating proteins. Most of these auxiliary proteins exist ubiquitously in oxygenic photosynthetic organisms. Due to differences in mobility and environmental conditions, land plants are subject to more extensive high light stress than algae and cyanobacteria. Therefore, land plants evolved additional thiol/disulfide-modulating proteins, such as Low Quantum Yield of PSII 1 (LQY1), to aid in the repair and reassembly cycle of PSII. In this study, we introduced an Arabidopsis thaliana homolog of LQY1 (AtLQY1) into the cyanobacterium Synechocystis sp. PCC6803 and performed a series of biochemical and physiological assays on AtLQY1-expressing Synechocystis. At a moderate growth light intensity (50 µmol photons m-2 s-1), AtLQY1-expressing Synechocystis was found to have significantly higher F v /F m, and lower nonphotochemical quenching and reactive oxygen species levels than the empty-vector control, which is opposite from the loss-of-function Atlqy1 mutant phenotype. Light response curve analysis of PSII operating efficiency and electron transport rate showed that AtLQY1-expressing Synechocystis also outperform the empty-vector control under higher light intensities. The increases in F v /F m, PSII operating efficiency, and PSII electron transport rate in AtLQY1-expressing Synechocystis under such growth conditions most likely come from an increased amount of PSII, because the level of D1 protein was found to be higher in AtLQY1-expressing Synechocystis. These results suggest that introducing AtLQY1 is beneficial to Synechocystis.

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