GROWTH HORMONE-INDUCED TYROSYL PHOSPHORYLATION AND DEOXYRIBONUCLEIC-ACID BINDING-ACTIVITY OF STAT5A AND STAT5B

Citation
Ls. Smit et al., GROWTH HORMONE-INDUCED TYROSYL PHOSPHORYLATION AND DEOXYRIBONUCLEIC-ACID BINDING-ACTIVITY OF STAT5A AND STAT5B, Endocrinology, 138(8), 1997, pp. 3426-3434
Citations number
43
Language
INGLESE
art.tipo
Article
Categorie Soggetti
Endocrynology & Metabolism
Journal title
ISSN journal
0013-7227
Volume
138
Issue
8
Year of publication
1997
Pages
3426 - 3434
Database
ISI
SICI code
0013-7227(1997)138:8<3426:GHTPAD>2.0.ZU;2-S
Abstract
GH is known to activate JAK2 tyrosine kinase and members of the Stat f amily of transcription factors, including Stats 1, 3, and 5. The recen t observation that at least two Stat5 proteins (Stat5A and Stat5B) exi st in mouse and human, raises the question of whether GH activates bot h Stat5A and Stat5B and, if so, whether the requirements for activatio n are the same. An initial report investigating this issue demonstrate d GH-dependent activation of Stat5A but not Stat5B. In this paper, we demonstrate (in COS cells expressing rat GH receptor (rGHR) and either Stat5A or Stat5B, 3T3-F442A fibroblasts, and CHO cells expressing rGH R) that GH induces tyrosyl phosphorylation of both Stat5A and Stat5B. Similar time courses of phosphorylation were observed for the two prot eins. Interestingly, the pattern of observed bands differs for the two forms of Stat5. Two closely migrating Stat5A bands can be detected in cells treated with?th or without GH. Both of these bands become tyros yl phosphorylated in response to GH. Three species of Stat5B are obser ved in untreated cells. An additional. more slowly migrating Stat5B ba nd, appears upon treatment with GH. The three more slower migrating St at5B bands observed in response to GH contain phosphorylated tyrosyl r esidues. We further demonstrate that GH induces binding of Stat5A and Stat5B, as well as Stat1, to the GAS-like element in the p-casein prom oter. We and others have demonstrated previously that specific regions of GHR are required for GH-dependent activation of what is here ident ified as Stat5B. To gain insight into the mechanism by which GH promot es tyrosyl phosphorylation of Stat5A, GH-dependent tyrosyl phosphoryla tion of Stat5A was examined in CHO cells expressing truncated and muta ted rGWII. The results indicate that Stat5A and Stat5B require the sam e regions of rGHR for maximal activation by GH: the C-terminal half of the cytoplasmic domain tyrosines 333 and/or 338 in the N-terminal hal f of the cytoplasmic domain; and the regions required for JAK2 activat ion. To dissect further the mechanism by which GH activates Stat5A and B, the requirement for JAK2 in GH-dependent Stat5 tyrosyl phosphoryla tion was assessed using JAK2-deficient cells expressing GHR (gamma 2A- GHR) and the wild-type parental cell Line expressing GHR (2C4-GHR). GH -induced tyrosyl phosphorylation of Stat5B in 2C4-GHR cells but not in the JAK2 deficient, gamma 2A-GHR cells, indicating that JAK2 is requi red for GH-dependent tyrosyl phosphorylation of Stat5B. Western blotti ng revealed that Stat5A is not expressed in this cell type. Taken toge ther, these findings suggest that: 1) GH activates both Stat5A and Sta t5B in several cell types; 2) the pattern of bands observed differs fo r Stat5A and Stat5B; 3) GH-dependent tyrosyl phosphorylation of Stat6A requires specific regions of GHR, and these requirements are the same as for Stat5B; and 4) JAK2. kinase is required for GH-dependent tyros yl phosphorylation of Stat5B and,most likely, Stat5A.