TY - JOUR

T1 - The role of the growth hormone (GH) receptor and JAK1 and JAK2 kinases in the activation of Stats 1, 3, and 5 by GH

AU - Smit, L S

AU - Meyer, D J

AU - Billestrup, Nils

AU - Norstedt, G

AU - Schwartz, J

AU - Carter-Su, C

PY - 1996/5

Y1 - 1996/5

N2 - GH has been shown to activate the GH receptor (GHR)-associated tyrosine kinase JAK2 and the Src homology 2 domain-containing transcription factors Stats (signal transducers and activators of transcription) 1, 3, and 5. The present work investigates the role of GHR and JAK2 in the activation of Stats 1, 3, and 5 by GH. The ability of GH to stimulate the tyrosyl phosphorylation of these Stats was assessed in Chinese hamster ovary (CHO) cells expressing truncated and mutated GHR. GH was observed to stimulate tyrosyl phosphorylation of Stats 1, 3, and 5 in CHO cells expressing GHRs that bind JAK2 [GHR1-638 (full-length) and GHR1-454 (lacks approximately half of the cytoplasmic domain)] but not in CHO cells expressing GHR that do not bind JAK2 (GHR1-318 or GHR1-294). GH-dependent tyrosyl phosphorylation of Stat5, but not Stats 1 or 3, was reduced in CHO cells expressing GHR1-454. GH-dependent tyrosyl phosphorylation of Stats 3 and 5 was severely reduced and undetectable for Stat1 in cells expressing GHR1-454 in which tyrosines 333 and 338 (the only tyrosines phosphorylated within 1-454) are mutated to phenylalanine (GHR1-454Y333, 338F). However, GH-dependent phosphorylation of Stats 1, 3, and 5 was observed in cells expressing full-length GHR in which tyrosines 333 and 338 are mutated to phenylalanine (GHR1-638Y333, 338F) GH, whose receptor lacks previously defined Stat1- or Stat3-binding sites, was found in 3T3-F442A fibroblasts and 2fTGH-GHR cells to stimulate tyrosyl phosphorylation of JAK2 to a substantially greater extent than, and JAK1 to a similar extent as, leukemia inhibitory factor (LIF) and/or interferon gamma (IFN gamma), ligands whose receptors contains Stat3- and Stat1-binding sites and activate Stat3 and Stat1, respectively, better than GH. These findings suggest that: 1) JAK2 is required for GH-dependent phosphorylation of Stats 1, 3, and 5; 2) tyrosines 333 and/or 338 are required for maximal tyrosyl phosphorylation of Stats 1, 3, and 5; 3) Stat5 binds to a phosphorylated tyrosine(s) within amino acids 454-638 in addition to tyrosines 333 and/or 338; 4) GH stimulates tyrosyl phosphorylation of JAK1 in addition to JAK2 with JAK2 having a much greater response; 5) some Stat3 and Stat5 (and possibly Stat1) may bind to nonphosphorylated amino acids in GHR or to phosphorylated tyrosines in proteins that bind to GHR (e.g. JAK22) to be maximally activated; and 6) if JAK2, which contains Stat3-binding motifs, does serve as a docking site for some Stat proteins, Stat-JAK2 binding is likely to be more important for GH than LIF or IFN gamma in 3T3-F442A cells since GH induces 15 times more tyrosyl-phosphorylated JAK2 than LIF or IFN gamma.

AB - GH has been shown to activate the GH receptor (GHR)-associated tyrosine kinase JAK2 and the Src homology 2 domain-containing transcription factors Stats (signal transducers and activators of transcription) 1, 3, and 5. The present work investigates the role of GHR and JAK2 in the activation of Stats 1, 3, and 5 by GH. The ability of GH to stimulate the tyrosyl phosphorylation of these Stats was assessed in Chinese hamster ovary (CHO) cells expressing truncated and mutated GHR. GH was observed to stimulate tyrosyl phosphorylation of Stats 1, 3, and 5 in CHO cells expressing GHRs that bind JAK2 [GHR1-638 (full-length) and GHR1-454 (lacks approximately half of the cytoplasmic domain)] but not in CHO cells expressing GHR that do not bind JAK2 (GHR1-318 or GHR1-294). GH-dependent tyrosyl phosphorylation of Stat5, but not Stats 1 or 3, was reduced in CHO cells expressing GHR1-454. GH-dependent tyrosyl phosphorylation of Stats 3 and 5 was severely reduced and undetectable for Stat1 in cells expressing GHR1-454 in which tyrosines 333 and 338 (the only tyrosines phosphorylated within 1-454) are mutated to phenylalanine (GHR1-454Y333, 338F). However, GH-dependent phosphorylation of Stats 1, 3, and 5 was observed in cells expressing full-length GHR in which tyrosines 333 and 338 are mutated to phenylalanine (GHR1-638Y333, 338F) GH, whose receptor lacks previously defined Stat1- or Stat3-binding sites, was found in 3T3-F442A fibroblasts and 2fTGH-GHR cells to stimulate tyrosyl phosphorylation of JAK2 to a substantially greater extent than, and JAK1 to a similar extent as, leukemia inhibitory factor (LIF) and/or interferon gamma (IFN gamma), ligands whose receptors contains Stat3- and Stat1-binding sites and activate Stat3 and Stat1, respectively, better than GH. These findings suggest that: 1) JAK2 is required for GH-dependent phosphorylation of Stats 1, 3, and 5; 2) tyrosines 333 and/or 338 are required for maximal tyrosyl phosphorylation of Stats 1, 3, and 5; 3) Stat5 binds to a phosphorylated tyrosine(s) within amino acids 454-638 in addition to tyrosines 333 and/or 338; 4) GH stimulates tyrosyl phosphorylation of JAK1 in addition to JAK2 with JAK2 having a much greater response; 5) some Stat3 and Stat5 (and possibly Stat1) may bind to nonphosphorylated amino acids in GHR or to phosphorylated tyrosines in proteins that bind to GHR (e.g. JAK22) to be maximally activated; and 6) if JAK2, which contains Stat3-binding motifs, does serve as a docking site for some Stat proteins, Stat-JAK2 binding is likely to be more important for GH than LIF or IFN gamma in 3T3-F442A cells since GH induces 15 times more tyrosyl-phosphorylated JAK2 than LIF or IFN gamma.

KW - Animals

KW - Base Sequence

KW - CHO Cells

KW - Cell Line

KW - Cricetinae

KW - DNA-Binding Proteins

KW - Growth Hormone

KW - Growth Inhibitors

KW - Humans

KW - Interferon-gamma

KW - Interleukin-6

KW - Janus Kinase 1

KW - Janus Kinase 2

KW - Leukemia Inhibitory Factor

KW - Lymphokines

KW - Milk Proteins

KW - Molecular Sequence Data

KW - Phosphotyrosine

KW - Protein-Tyrosine Kinases

KW - Proto-Oncogene Proteins

KW - Receptors, Somatotropin

KW - STAT1 Transcription Factor

KW - STAT3 Transcription Factor

KW - STAT5 Transcription Factor

KW - Signal Transduction

KW - Structure-Activity Relationship

KW - Trans-Activators

U2 - 10.1210/mend.10.5.8732683

DO - 10.1210/mend.10.5.8732683

M3 - Journal article

C2 - 8732683

VL - 10

SP - 519

EP - 533

JO - Molecular Endocrinology

JF - Molecular Endocrinology

SN - 0888-8809

IS - 5

ER -