In land plants and cyanobacteria, co-translational association of chlorophyll (Chl) to the nascent D1 polypeptide, a reaction center protein of photosystem II (PSII), requires a Chl binding complex consisting of a short-chain dehydrogenase (HIGH CHLOROPHYLL FLUORESCENCE 244 [HCF244]/Uncharacterized Protein 39 [Ycf39]) and ONE-HELIX PROTEINS (OHP1 and OHP2 in chloroplasts) of the light-harvesting antenna complex (LHC) superfamily. Here, we show that an ohp2 mutant of the green alga Chlamydomonas (Chlamydomonas reinhardtii) fails to accumulate core PSII subunits, in particular D1 (encoded by the psbA mRNA). Extragenic suppressors arose at high frequency, suggesting the existence of another route for Chl association to PSII. The ohp2 mutant was complemented by the Arabidopsis (Arabidopsis thaliana) ortholog. In contrast to land plants, where psbA translation is prevented in the absence of OHP2, ribosome profiling experiments showed that the Chlamydomonas mutant translates the psbA transcript over its full length. Pulse labelling suggested that D1 is degraded during or immediately after translation. The translation of other PSII subunits was affected by assembly-controlled translational regulation. Proteomics showed that HCF244, a translation factor which associates with and is stabilized by OHP2 in land plants, still partly accumulates in the Chlamydomonas ohp2 mutant, explaining the persistence of psbA translation. Several Chl biosynthesis enzymes overaccumulate in the mutant membranes. Partial inactivation of a D1-degrading protease restored a low level of PSII activity in an ohp2 background, but not photoautotrophy. Taken together, our data suggest that OHP2 is not required for psbA translation in Chlamydomonas, but is necessary for D1 stabilization.

Excess light causes severe photodamage to photosystem II (PSII) where the primary charge separation for electron transfer takes place. Dissection of mechanisms underlying the PSII maintenance and repair cycle in green algae promotes the usage of genetic engineering and synthetic biology to improve photosynthesis and biomass production. In this study, we systematically analyzed the high light (HL) responsive immunophilin genes in Chlamydomonas (Chlamydomonas reinhardtii) and identified one chloroplast lumen-localized immunophilin, CYN28, as an essential player in HL tolerance. Lack of CYN28 caused HL hypersensitivity, severely reduced accumulation of PSII supercomplexes, and compromised PSII repair in cyn28. The thylakoid FtsH (filamentation temperature-sensitive H) is an essential AAA family metalloprotease involved in the degradation of photodamaged D1 during the PSII repair cycle and was identified as one potential target of CYN28. In the cyn28 mutant, the thylakoid FtsH undergoes inefficient turnover under HL condition. The CYN28-FtsH1/2 interaction relies on the FtsH N-terminal proline residues and is strengthened particularly under HL. Further analyses demonstrated CYN28 displays peptidyl-prolyl isomerase (PPIase) activity, which is necessary for its physiological function. Taken together, we propose that immunophilin CYN28 participates in PSII maintenance and regulates the homeostasis of FtsH under HL stress via its PPIase activity.