Biogenesis of chloroplast membranes IX. Development of photophosphorylation and proton pump activities in greening Chlamydomonas reinhardi y-1 as measured with an open-cell preparation

Chlamydomonas reinhardi y-1 cells are rendered permeable to substrates and cofactors for photosynthetic electron transfer and photophosphorylation by applying low shearing forces in a hypertonic viscous medium. The chloroplast integrity is well preserved in such open-cell preparations which exhibit high and stable photophosphorylation rates for both photosystems and proton pump activity. These activities were measured during greening of dark-grown y-1 cells using open-cell preparations. It was found that photophosphorylation and light-dependent proton uptake are absent in preparations obtained from cells grown in the dark for 5-6 generations which still contained 1-2 μg chlorophyll per 10⁷ cells. The activities became measurable following exposure of the cells to the light and increased rapidly reaching a maximal rate per chlorophyll unit 3-4 times higher than that exhibited by light-grown cells after 2-3 h of illumination when the chlorophyll content of the cells has increased only slightly. As chlorophyll continues to accumulate the activity per chlorophyll unit declined and became equal to that usually found in light grown cells. Photosynthetic ATP formation and pH rise do not develop in cells greening in the presence of chloramphenicol which specifically inhibits the synthesis of chloroplast made proteins. Chloroplast membranes formed in the presence of chloramphenicol which are enriched in proteins of cytoplasmic origin resume the light-dependent proton pump activity and photophosphorylation by both photosystems if the cells are further incubated in absence of chloramphenicol. The repair of the inactive membranes requires synthesis of proteins within the chloroplast and does not require light and additional synthesis of chlorophyll or proteins of cytoplasmic origin.