cAMP-dependent phosphorylation and hexamethylene-bis-acetamide induced dephosphorylation of p19 in murine erythroleukemia cells.
The objective of this study was to investigate cyclic-adenosinemonophosphate (cAMP)-dependent phosphorylation in murine erythroleukemia (MEL) cells and to identify either direct substrates of cAMP-dependent kinase or downstream effectors of cAMP dependent phosphorylation with a potential function in growth and differentiation. MEL-cells rendered deficient in cAMP-dependent protein kinase (A-kinase) activity by stable transfection with DNA encoding for either a mutant regulatory subunit or a specific peptide inhibitor of A-Kinase (PKI) are unable to differentiate normally in response to chemical inducers. We have identified by 2-D Western blotting 2 phosphorylated forms of p19, a highly conserved 18-19 kDa cytosolic protein that is frequently upregulated in transformed cells and undergoes phosphorylation in mammalian cells upon activation of several signal transduction pathways. The phosphorylation of the more acidic phosphorylated form is increased in a cAMP-dependent fashion and impaired in cells deficient in cAMP-dependent kinase (A-kinase). Treatment of MEL-cells with the chemical inducer of differentiation hexamethylene-bisacetamide (HMBA) led to dephosphoryation of this phosphoform. Our data are compatible with previous observations which imply that phosphorylation of Ser 38 in p19 by p34cdc2-kinase leads to a more basic phosphoform and simultaneous phosphorylation by mitogen-activated kinase of Ser 25 in response to protein kinase C and the cAMP-dependent kinase creates the more acidic species.
Scheele
J S
JS
Department of Chemistry and Biochemistry, University of California San Diego, La Jolla 92093-0612, USA.
D000081:Acetamides / Q000494:pharmacology*; D000818:Animals; D000906:Antibodies; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D000242:Cyclic AMP / Q000494:pharmacology*; D017868:Cyclic AMP-Dependent Protein Kinases / Q000172:deficiency; D015180:Electrophoresis, Gel, Two-Dimensional; D004915:Leukemia, Erythroblastic, Acute / Q000201:enzymology / Q000276:immunology / Q000473:pathology; D051379:Mice; D008868:Microtubule Proteins; D010749:Phosphoprotein Phosphatases / Q000187:drug effects; D010750:Phosphoproteins / Q000032:analysis / Q000187:drug effects*; D010766:Phosphorylation / Q000187:drug effects; D011506:Proteins / Q000032:analysis / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D050777:Stathmin; D014407:Tumor Cells, Cultured
D000081:Acetamides; D000906:Antibodies; D000970:Antineoplastic Agents; D008868:Microtubule Proteins; D010750:Phosphoproteins; D011506:Proteins; D050777:Stathmin; C493994:Stmn1 protein, mouse; C061082:Stmn1 protein, rat; D000242:Cyclic AMP; D019908:Proto-Oncogene Proteins c-raf; D017868:Cyclic AMP-Dependent Protein Kinases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D010749:Phosphoprotein Phosphatases; C014026:hexamethylene bisacetamide