Pattern of Protein Phosphorylation in Rat Hepatocytes Stimulated by Glucagen or by the Ca2+-Linked Hormones

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Abstract
The high-resolution electrophoretic technique of O''Farrell was adapted to analyze phosphorylated proteins from rat hepatocytes. Total proteins were extracted from rat hepatocytes incubated in the presence of [32P]phosphate and with 2 types of stimuli: glucagon on the one hand and the Ca2+-linked hormones on the other. About 200 phosphorylated polypeptides were separated. Glucagon modifies the incorporation of [32P]phosphate in at least 17 polypeptides and dibutyryl AMP mimics this hormonal effect, implying a common mechanism of action. Phenylephrine (in the presence of the .beta.-antagonist propranolol), vasopressin and angiotensin all modify the incorporation of [32P]phosphate in about 13 polypeptides; since the Ca2+ ionophore A23187 reproduces the effect of these agents it may be concluded that Ca2+ mediates their effects. Not all of the substrates affected by the 2 types of hormones are identical. Both types of stimuli increase the phosphorylation of a same set of 7 proteins and decrease the phosphorylation of a same set of 3 proteins but 7 proteins have their phosphorylation uniquely enhanced by glucagon whereas 3 other specific proteins get more phosphorylated by the Ca2+-linked hormones. The clear differences between the patterns of protein phosphorylation observed in the presence of glucagon and dibutyryl AMP on the one hand and by the Ca2+-linked hormones on the other strongly suggest different mechanisms of action for these 2 types of stimuli.