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The amount of messenger RNA (mRNA) for polypeptides of the chlorophyll a/bprotein complex of thylakoid membranes in etiolated and greening cells of Chlamydomonas reinhardtii y-1 was examined by immunoprecipitation and electrophoresis of products of in vitro translation to determine at which stage production of these polypeptides is regulated. Cells grown 4 d in the dark at 25°C contained small amounts of translatable mRNA for the major membrane polypeptides. Exposure of these etiolated cells to light, under conditions in which the membrane polypeptides accumulated, resulted in a significant increase in the quantity of the mRNA. In contrast, when etiolated cells were incubated for 1-2 h in the dark at 38°C, translation assays indicated that mRNA for the membrane polypeptides became abundant. Moreover, the quantity of the mRNA did not increase when these cells subsequently were exposed to light. Therefore, at 38°C the cellular level of the polypeptides is not regulated by synthesis of mRNA. The in vitro synthesized polypeptides, which were precipitated with antibodies prepared against the purified thylakoid polypeptides, had apparent molecular weights of 31,500 and 30,000. The corresponding immunoprecipitated polypeptides made in vivo had apparent molecular weights of 29,500 and 26,000. Thus, the membrane polypeptides are made as precursors.No net accumulation of the polypeptides occurred in cells in the dark at 38°C, but immunoreactive polypeptides the size of the mature membrane components were labeled during incubation of cells with [14C]acetate in the dark. These results indicated that the mRNA was translated in the dark, but since the polypeptides did not accumulate, the products of translation were probably degraded. We conclude from our experiments that at 25°C production of the polypeptides is regulated by the level of translatable mRNA in the cells. At 38°C, however, the accumulation of the polypeptides is controlled by posttranslational processes.

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The amount of chloroplast ribosomal RNAs of Chlamydomonas reinhardtii which sediment at 15,000 g is increased when cells are treated with chloramphenicol . Preparations of chloroplast membranes from chloramphenicol-treated cells contain more chloroplast ribosomal RNAs than preparations from untreated cells . The membranes from treated cells also contain more ribosome-like particles, some of which appear in polysome-like arrangements . About 50% of chloroplast ribosomes are released from membranes in vitro as subunits by 1 mM puromycin in 500 mM KC1 . A portion of chloroplast ribosomal subunits is released by 500 mM KCl alone, a portion by 1 mM puromycin alone, and a portion by 1 mM puromycin in 500 mM KCl . Ribosomes are not released from isolated membranes by treatment with ribonuclease . Membranes in chloroplasts of chloramphenicol-treated cells show many ribosomes associated with membranes, some of which are present in polysome-like arrangements . This type of organization is less frequent in chloroplasts of untreated cells . Streptogramin, an inhibitor of initiation, prevents chloramphenicol from acting to permit isolation of membrane-bound ribosomes .