Abstract
Accumulation of monomer and dimer photosystem (PS) II reaction center core complexes has been analyzed by two-dimensional Blue-native/SDS-PAGE in Synechocystis PCC 6803 wild type and in mutant strains lacking genes psbA, psbB, psbC, psbDIC/DII, or the psbEFLJ operon. In vivo pulse-chase radiolabeling experiments revealed that mutant cells assembled PSII precomplexes only. In DeltapsbC and DeltapsbB, assembly of reaction center cores lacking CP43 and reaction center complexes was detected, respectively. In DeltapsbA, protein subunits CP43, CP47, D2, and cytochrome b559 were synthesized, but proteins did not assemble. Similarly, in DeltapsbD/C lacking D2, and CP43, the de novo synthesized proteins D1, CP47, and cytochrome b559 did not form any mutual complexes, indicating that assembly of the reaction center complex is a prerequisite for assembly with core subunits CP47 and CP43. Finally, although CP43 and CP47 accumulated in DeltapsbEFLJ, D2 was neither expressed nor accumulated. We, furthermore, show that the amount of D2 is high in the strain lacking D1, whereas the amount of D1 is low in the strain lacking D2. We conclude that expression of the psbEFLJ operon is a prerequisite for D2 accumulation that is the key regulatory step for D1 accumulation and consecutive assembly of the PSII reaction center complex.
Highlights
Accumulation of monomer and dimer photosystem (PS) II reaction center core complexes has been analyzed by two-dimensional Blue-native/SDS-PAGE in Synechocystis PCC 6803 wild type and in mutant strains lacking genes psbA, psbB, psbC, psbDIC/DII, or the psbEFLJ operon
Synthesis of photosystem II (PSII) subunits D2, D1, CP47, and CP43 and their assembly was monitored by radiolabel accumulation in protein complexes, whereas the steady state level of protein subunits in complexes was monitored by Coomassie staining or by Western blotting after two-dimensional native/SDS-PAGE
Protein subunits of PSII, CP47, CP43, D2, and D1 accumulated in two complexes at about 300 and 600 kDa in wild type (WT) cells, corresponding to monomeric and dimeric reaction center core complexes RCC1 and RCC2, respectively
Summary
Accumulation of monomer and dimer photosystem (PS) II reaction center core complexes has been analyzed by two-dimensional Blue-native/SDS-PAGE in Synechocystis PCC 6803 wild type and in mutant strains lacking genes psbA, psbB, psbC, psbDIC/DII, or the psbEFLJ operon. In the green alga Chlamydomonas reinhardtii, de Vitry et al [22] described the importance of certain PSII subunits for the accumulation of PSII Using several mutants they concluded that the D2 protein is necessary for synthesis of other large subunits like CP47 and D1, whereas D1 synthesis was not required for synthesis of D2 and CP47. Recent characterization of psbJ and psbL deletion mutants in plants confirmed that these proteins do not significantly affect accumulation of PSII RCC monomers but may be involved in proper assembly of the oxygen evolving apparatus in cyanobacteria and higher plant chloroplast (34 –38)
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