The effect of growth state on the activity of the protein kinase isoenzymes. An increase in type II cyclic AMP-dependent protein kinase is correlated with growth arrest in G1 phase.
Native polyacrylamide gels have been used to resolve protein kinase isoenzymes from cultured cells and the protein kinases have been identified by carrying out phosphorylation reactions in the gel. Following electrophoresis the gels were incubated with histone and [gamma-32P]ATP. The gels were then thoroughly washed and dried down, and the protein kinases were located by autoradiography. Protein kinase activity as measured in the gel system was a linear function of cytosol protein concentration up to about 100 microgram per channel and incorporation of 32P into histone was time dependent. Three bands of protein kinase activity were resolved in cytosol samples from baby hamster kidney (BHK) fibroblasts. The band with the lowest relative mobility utilized histone IIA or casein equally well as substrate protein whereas bands 2 and 3 demonstrated a clear preference for histone. Bands 2 and 3 displayed a relative mobility in electrophoresis that was identical to that observed for cyclic AMP-dependent protein kinases I and II from rat liver. Treatment of cytosol samples with cyclic AMP prior to electrophoresis resulted in the disappearance of cyclic AMP-dependent protein kinases from the gel profile. This method was employed to identify bands 2 and 3 as cyclic AMP-dependent protein kinases. The protein kinases in growth-arrested cells were compared with proliferating cells. We have observed a 3.5-fold increase in the activity of Type II protein kinase as the cells arrest growth in G1 phase of the cell cycle. This increase in Type II is correlated with the increase in cells blocked in G1 and a decrease in Type II activity appears to be an early event in permitting cells to leave G1 and resume growth.
D000818:Animals; D002453:Cell Cycle; D002460:Cell Line; D006224:Cricetinae; D000242:Cyclic AMP / Q000494:pharmacology; D007527:Isoenzymes / Q000032:analysis / Q000378:metabolism*; D007668:Kidney / Q000166:cytology / Q000201:enzymology; D008099:Liver / Q000201:enzymology*; D011494:Protein Kinases / Q000378:metabolism*; D051381:Rats; D011919:Rats, Inbred Strains
D007527:Isoenzymes; D000242:Cyclic AMP; D011494:Protein Kinases