INHIBITION OF CHLOROPLAST PROTEIN-PHOSPHORYLATION BY CAMP IN LEMNA-PAUCICOSTATA-6746

Phosphorylation was carried out in chloroplasts, purified on sucrose d ensity gradient, using [gamma-P-32]ATP. Among several phosphopolypepti des detected, polypeptides in the M(r) range of 24-22 k, which are kno wn to constitute the light-harvesting chlorophyll a/b-binding protein complex (LHCP), and 16-18 k were highly phosphorylated. Cyclic AMP (10 0 mu M) decreased the overall phosphorylation of chloroplast polypepti des and its effect was more striking on LHCP. Other nucleotides such a s cGMP, 2',3'-cAMP, 2',3'-cGMP and 5'-AMP, at equimolar level, did not show any significant effect on phosphorylation of chloroplast polypep tides. The effect of cAMP was also studied on light-dependent in vitro phosphorylation of chloroplast polypeptides by incubating a crude pre paration of intact chloroplasts with [(32)Pi]. In dark control, two po lypeptides of M(r) 66 and 64 k were distinctly phosphorylated. On illu mination, several other polypeptides in the range of 97-12 k were also phosphorylated, with the 26-24 k LHCP fraction being the major phosph opolypeptides. Cyclic AMP (at 100 mu M and 1 mM) specifically caused a decrease in phosphorylation of the 26-24, 21-20 and 12 k polypeptides . The phosphorylation status of the 66-64 k doublet, as attained in da rk, remained unaffected in the presence of light and/or cAMP. Sodium f luoride, a protein phosphatase inhibitor, enhanced the phosphorylation of 26-24, 21-20 and 12 k polypeptides but did not affect the inhibito ry action of cAMP. It is thus likely that cAMP down-regulates protein kinase(s) responsible for the phosphorylation of these polypeptides. T he present study also provides evidence that this protein kinase is se nsitive to staurosporine, a protein kinase inhibitor.