Growth hormone (GH) induction of tyrosine phosphorylation and activation of mitogen-activated protein kinases in cells transfected with rat GH receptor cDNA.
The mechanism of growth hormone (GH) action was studied in Chinese hamster ovary (CHO) cells transfected with GH receptor cDNA. Cytosolic extracts from GH- or phorbol ester (12-O-tetradecanoyl 4 beta-phorbol 13-acetate)-treated cells, transfected with full-length GH receptor cDNA, had an enhanced ability to phosphorylate myelin basic protein. Myelin basic protein, a substrate for mitogen-activated protein (MAP) kinase, was maximally phosphorylated using extracts from cells treated with 50 nM bovine GH for 10 min. In addition, GH treatment resulted in an increased cell proliferation by 30-60%. GH and 12-O-tetradecanoyl 4 beta-phorbol 13-acetate cause tyrosine phosphorylation of two proteins with M(r) of 40,000 and 42,000 that are also recognized by MAP kinase antibodies. These proteins were identified as MAP kinases by analyzing phosphotyrosine immunoprecipitates on Western blots using MAP kinase antibodies. In addition, GH induces mitogenicity, as well as MAP kinase activation, in CHO cells expressing a receptor in which 184 amino acids had been deleted in the carboxyl-terminal part of the intracellular domain. No GH effects were seen in untransfected cells, in CHO cells expressing a truncated GH receptor containing only 5 of 349 amino acids in the intracellular domain, or in cells expressing the soluble GH-binding protein. In conclusion, our data show that GH treatment of CHO cells, reconstituted with GH receptors, initiates a phosphorylation cascade which includes MAP kinase.
Möller
C
C
Center for Biotechnology, Karolinska Institute, Novum, Huddinge, Sweden.
Hansson
A
A
Enberg
B
B
Lobie
P E
PE
Norstedt
G
G
D000818:Animals; D015153:Blotting, Western; D016466:CHO Cells; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002455:Cell Division / Q000187:drug effects; D006224:Cricetinae; D004247:DNA / Q000235:genetics; D004789:Enzyme Activation; D038362:Glycogen Synthase Kinase 3; D013006:Growth Hormone / Q000378:metabolism / Q000494:pharmacology*; D010766:Phosphorylation; D019000:Phosphotyrosine; D011401:Promoter Regions, Genetic; D011494:Protein Kinases / Q000302:isolation & purification / Q000378:metabolism*; D051381:Rats; D011986:Receptors, Somatotropin / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013937:Thymidine Kinase / Q000235:genetics; D014162:Transfection; D014443:Tyrosine / Q000031:analogs & derivatives / Q000032:analysis
D011986:Receptors, Somatotropin; D019000:Phosphotyrosine; D014443:Tyrosine; D013006:Growth Hormone; D004247:DNA; D011494:Protein Kinases; D013937:Thymidine Kinase; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D038362:Glycogen Synthase Kinase 3; D013755:Tetradecanoylphorbol Acetate