PLATELET-DERIVED GROWTH-FACTOR STIMULATES THE RELEASE OF PROTEIN-KINASE-A FROM THE CELL-MEMBRANE

The mitogenic action of growth factors involves the stimulation of int racellular protein kinases. In this report we have characterized the m ajor protein kinase released from Balb/c 3T3 and normal rat kidney pla sma membranes by the action of platelet-derived growth factor (PDGF). PDGF appears to stimulate the release of approximately 10 proteins, at least one of which is a kinase capable of phosphorylating proteins on Ser or Thr (as determined by the lability of the phosphate to alkali treatment). More than 90% of the Ser/Thr kinase activity was inhibited by PKI5-22, a specific peptide inhibitor of the cAMP-dependent protei n kinase (PKA). We used immunoblotting to confirm that the kinase rele ased in response to PDGF was PRA. cAMP also stimulated the release of PKA, and the set of protein substrates phosphorylated was similar foll owing PDGF or cAMP stimulation. Interestingly, in the presence of a cA MP analogue ((R(p))-cAMPS), cAMP could not induce dissociation of PKA from the membranes, whereas stimulation by PDGF increased the level of PKA activation. Furthermore, unlike Swiss 3T3 cells, neither Balb/c 3 T3 fibroblasts nor normal rat kidney cells accumulate cAMP in response to PDGF, yet the level of PKA in the cytosol of these intact cells in creases in response to PDGF. Thus, it appears as though PDGF activatio n of the membrane-associated form of the PKA holoenzyme occurs by a me chanism independent of an elevation in cAMP levels.