Light dependence and kinetics of reversible phosphorylation of the D1 reaction center protein of Photosystem II was studied in pumpkin leaves. At growth light, maximal phosphorylation of D1 was observed after illumination of 1 h, with higher phosphorylation rates at stronger irradiances. 70-85% of D1 became phosphorylated, corresponding to the proportion of the protein in appressed thylakoid membranes. Comparison of the kinetics of D1 phosphorylation and photoinactivation of Photosystem II revealed that D1 phosphorylation became saturated before any significant photoinhibition of Photosystem II could be detected. Dephosphorylation of D1 in both dim light and darkness was determined in leaves preilluminated with high light for various periods. Similar rates of D1 dephosphorylation were observed after short preillumination conditions that induce no significant loss of functional Photosystem II centers. In contrast, photodamage to Photosystem II centers significantly decreased the dephosphorylation rate of D1 in darkness, and no dephosphorylation occurred in leaves containing mainly damaged Photosystem II centers. Darkness also blocked the degradation of damaged D1 after photoinhibitory preillumination. Degradation of damaged D1 could be prevented even in dim light by sodium fluoride, an inhibitor of protein phosphatases, indicating that dephosphorylation is a prerequisite for D1 proteolysis. We conclude that in higher plants (i) high light induced photodamage to Photosystem II occurs in the centers containing phosphorylated D1. (ii) Dephosphorylation of phosphorylated and photodamaged D1 is associated with the repair cycle of inactivated Photosystem II and is a light-dependent reaction in vivo. (iii) Dephosphorylation of D1 in functional Photosystem II centers, however, occurs rapidly and independent of light. We suggest that two reversible phosphorylation cycles with spatially segregated protein phosphatases are involved in dephosphorylation of functional and damaged phosphorylated D1, respectively.
Carbon Dioxide Chlorophyll