Insulin activates the kinase activity of the Raf-1 proto-oncogene by increasing its serine phosphorylation.

Insulin was found to stimulate the serine/threonine kinase activity of the proto-oncogene product Raf-1. This stimulation was observed in HeLa, NIH 3T3, and Chinese hamster ovary cells, all overexpressing the human insulin receptor. In the HeLa cells, 100 pM insulin gave a significant increase in Raf-1 kinase activity, and 100 nM insulin caused a maximal 2-5-fold increase in activity. The increase in activity was detected after 2 min of insulin treatment and peaked after 5 min. In addition to stimulating Raf-1 kinase activity, insulin caused a shift in the electrophoretic mobility of the Raf-1 protein and an increase in the amount of serine phosphorylation of Raf-1. Moreover, a serine/threonine-specific phosphatase, phosphatase 1, but not two tyrosine-specific phosphatases, was found to deactivate the insulin-activated Raf-1 kinase activity. These findings indicate that insulin activates the serine/threonine kinase activity of the Raf-1 proto-oncogene by increasing its content of phosphoserine.