Phosphorylation of P20 is associated with the actions of insulin in rat skeletal and smooth muscle

Although a large number of protein kinases/phosphatases involved in insulin 's actions have been characterized recently, relatively few of the downstre am phosphoproteins have been identified. We have employed two-dimensional g el electrophoresis-based proteome analysis to investigate the insulin-evoke d phosphorylation cascade in rat soleus muscle. Insulin reproducibly increa sed phosphorylation of a 20-kDa protein with a pi value of 6.0, which was i dentified subsequently as a phospho-isoform of P20, a small heat-shock-rela ted protein. The adenylyl cyclase activator, forskolin, decreased phosphory lation of this P20 isoform and increased phosphorylation of another two P20 isoforms, with pi values of 5.9 and 5.6. Two-dimensional peptide mapping r evealed that the phospho-peptides of these three P20 isoforms are different . In contrast to its action in soleus muscle, insulin decreased phosphoryla tion of the P20 isoform with pi 6.0 and increased phosphorylation of the tw o isoforms with pi values of 5.9 and 5.6 in vascular smooth muscle. This ef fect is similar to that induced by vasodilatory stimuli, suggesting that in sulin could exert its vasodilatory action by affecting phosphorylation of P 20. In summary, these results demonstrate that insulin differently modulate s phosphorylation of P20 in skeletal and smooth muscle, and suggest that P2 0 could be a potential modulator of insulin's functions in these tissues.