REQUIREMENT FOR THE H-PHOSPHOPROTEIN IN PHOTOSYSTEM-II OF CHLAMYDOMONAS-REINHARDTII

To dissect the expression of the psbB gene cluster of the Chlamydomona s reinhardtii chloroplast genome and to assess the role of the photosy stem II H-phosphoprotein (PSII-H) in the biogenesis and/or stabilizati on of PSII, an aadA gene cassette conferring spectinomycin resistance was employed for mutagenesis. Disruption of the gene cluster has no ef fect on the abundance of transcripts of the upstream psbB/T locus. Lik ewise, interruption of psbB/T and psbH with a strong transcriptional t erminator from the rbcL gene does not influence transcript accumulatio n. Thus, psbB/T and psbH may be independently transcribed, and the lat ter gene seems to have its own promoter in C. reinhardtii. In the abse nce of PSII-H, translation and thylakoid insertion of chloroplast PSII core proteins is unaffected, but PSII proteins do not accumulate. Bec ause the deletion mutant also exhibits PSII deficiency when dark-grown , the effect is unrelated to photoinhibition. Turnover of proteins B a nd C of PSII and the polypeptides PSII protein A and PSII protein D is faster than in wild-type cells but is much slower than that observed in other PSII-deficient mutants of C. reinhardtii, suggesting a periph eral location of PSII-H in PSII. The role of PSII-H on PSII assembly w as examined by sucrose gradient fractionation of pulse-labeled thylako ids; the accumulation of high-molecular-weight forms of PSII is severe ly impaired in the psbH deletion mutant. Thus, a primary role of PSII- H may be to facilitate PSII assembly/stability through dimerization. P SII-H phosphorylation, which possibly occurs at two sites, may also be germane to its role in regulating PSII structure, stabilization, or a ctivity.