Basal phosphorylation of cyclic AMP-regulated phosphoproteins in intact S49 mouse lymphoma cells.
Protein phosphorylation in intact S49 mouse lymphoma cells was studied by using high-resolution two-dimensional gel electrophoresis of proteins labelled with [35S]methionine or [32P]Pi. In wild-type cells substrates for cyclic AMP-stimulatable phosphorylation exhibited high basal phosphorylation; in mutant cells deficient in activities of either cyclic AMP-dependent protein kinase or adenylate cyclase, basal phosphorylation of most of these substrates was negligible. Analysis of tryptic phosphopeptides from proteins labelled with [32P]Pi in wild-type cells suggested that identical sites were phosphorylated under conditions of both basal and hormonally elevated concentrations of cyclic AMP. These results argue that most basal phosphorylation is a consequence of partial activation of cyclic AMP-dependent protein kinase and that this activation is attributable to basal concentrations of cyclic AMP. For the intermediate filament protein vimentin, basal phosphorylation was largely at a site distinct from that stimulated by increased cyclic AMP, and basal phosphorylation was not markedly different in mutant and wild-type cells. Vimentin phosphorylated at both sites was not observed. Cyclic AMP treatment resulted in enhanced phosphorylation at the cyclic AMP-specific site and decreased phosphorylation at the cyclic AMP-independent site.
Steinberg
R A
RA
Kiss
Z
Z
D000262:Adenylyl Cyclases / Q000172:deficiency; D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D008223:Lymphoma / Q000378:metabolism*; D008715:Methionine / Q000378:metabolism; D051379:Mice; D010446:Peptide Fragments / Q000032:analysis; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000172:deficiency; D014746:Vimentin / Q000378:metabolism
D010446:Peptide Fragments; D010750:Phosphoproteins; D014746:Vimentin; D008715:Methionine; D000242:Cyclic AMP; D011494:Protein Kinases; D000262:Adenylyl Cyclases