SYNTHESIS AND TURNOVER OF PHOTOSYSTEM-II REACTION-CENTER PROTEIN D1 -RIBOSOME PAUSING INCREASES DURING CHLOROPLAST DEVELOPMENT

The chloroplast photosystem II reaction center protein D1 contains fiv e membrane-spanning helices and binds chlorophyll, carotenoid, quinone , iron, and probably manganese. Turnover of pulse-labeled D1 in isolat ed plastids was found to involve cleavage between helix IV and helix V , which releases a 23-kDa N-terminal peptide and two C-terminal peptid es of 10 and 8 kDa. Ribosomes pause at specific sites during translati on of D1, which results in the accumulation of D1 translation intermed iates. Pulse-labeling assays followed by poly some isolation and immun oprecipitation identified paused D1 translation intermediates of 9, 12 .5, 15-18, 20, 21, 24, and 28-32 kDa. Ribosome pausing was not altered when dark grown seedlings were illuminated for up to 1 h, even though this treatment stimulated accumulation of chlorophyll and D1. However , illumination of plants for 16-72 h resulted in increased ribosome pa using and the build-up of D1 translation intermediates. We hypothesize that ribosome pausing during synthesis of D1 im proves the efficiency of chlorophyll binding to D1 nascent chains and enhances accumulation of D1 in mature chloroplasts, which have reduced rates of chlorophyll biosynthesis.