PHOSPHORYLATION OF ROD OUTER SEGMENT PROTEINS MODULATES PHOSPHATIDYLETHANOLAMINE N-METHYLTRANSFERASE AND PHOSPHOLIPASE A(2) ACTIVITIES IN PHOTORECEPTOR-MEMBRANES

The activities of enzymes involved in lipid metabolism-phospholipase A , (PLA,) and phosphatidylethanolamine N-methyltransferase (PE N-MTase) -were found to be differently affected by pre-incubation of rod outer segments (ROS) under protein phosphorylating or dephosphorylating cond itions. Exposure to cAMP-dependent protein kinase (PKA), under dark or light conditions, produced a significant increase in PE N-MTase activ ity, whereas PLA, activity decreased. Under standard protein kinase C (PKC) phosphorylating conditions in light, PE N-MTase activity was sti mulated and PLA, activity was not affected. When the assays were perfo rmed in the dark, both enzymatic activities were unaffected when compa red to the corresponding controls. Incubation of ROS membranes in ligh t in the presence of PKC activators phorbol 12,13-dibutyrate (PDBu) an d dioctanoylglycerol (DOG) resulted in the same pattern of changes in enzyme activities as described for standard PKC phosphorylating condit ion. Pre-incubation of membranes with the PKC inhibitor H-7 reduced th e stimulation of PDBu on PE N-MTase activity, and had no effect on PLA , activity in ROS membranes incubated with the phorbol ester. Pre-trea tment of isolated ROS with alkaline phosphatase resulted in decreased PE N-MTase activity and produced a significant stimulation of PLA, act ivity under dark as well as under light conditions when compared to th e corresponding controls. These findings suggest that ROS protein phos phorylation and dephosphorylation modulates PE N-MTase and PLA, activi ties in isolated ROS, and that these activities are independently and specifically modulated by particular kinases. Furthermore, dephosphory lation of ROS proteins has the opposite effect to that produced by pro tein phosphorylation on the enzymes studied. (C) 1998 Elsevier Science Inc. All rights reserved.