DEVELOPMENTALLY-REGULATED IN-VITRO PHOSPHORYLATION OF A 85 KDA TRITON-INSOLUBLE PROTEIN OF THE CEREBRAL-CORTEX OF RATS

We studied the ontogeny of concentration and in vitro phosphorylation of an 85 kDa Triton-insoluble protein from cerebral cortex of 7, 15, 2 1 and 90 day old rats. The Triton-insoluble cytoskeletal fraction cont ains an 85 kDa basic phosphoprotein different ii-sm synapsin 1, as det ermined by nonequilibrium pH gradient electrophoresis and phosphopepti de mapping with VS protease. The concentration of the 85 kDa cytoskele tal associated phosphoprotein was analyzed during development. Results indicated that the concentration of this protein oscillated during su ckling, presenting a maximal value at day 15 and decreasing again to s tabilize at values near those of 7 day old rats, remaining constant in 21 and 90 day old animals. However, in vitro P-32 incorporation, expr essed as cpm/mu g, presented a developmentally regulated pattern, with maximal values In young rats, declining with age to negligible values in 90 day old animals. The endogenous phosphorylating system responsi ble for in vitro P-32 incorporation into the 85 kDa protein was determ ined by the addition of specific activators of second-messenger protei n kinases (cAMP, Ca2+/calmodulin and Ca2+/phosphatidylserine/phorbol e ster) and a protein phosphatase inhibitor (okadaic acid to the incubat ion system. Results suggested that the in vitro phosphorylation system is composed of protein kinase A, Ca2+/calmodulin dependent protein ki nase and protein phosphatase 1.