SOCS/CIS protein inhibition of growth hormone-stimulated STAT5 signaling by multiple mechanisms

J Biol Chem. 1999 Dec 10;274(50):35553-61. doi: 10.1074/jbc.274.50.35553.

Abstract

The inhibition of growth hormone (GH) signaling by five members of the GH-inducible suppressor of cytokine signaling (SOCS/CIS) family was investigated in transfected COS cells. Complete inhibition of GH activation of the signal transducer STAT5b and STAT5b-dependent transcriptional activity was observed upon expression of SOCS-1 or SOCS-3, while partial inhibition (CIS, SOCS-2) or no inhibition (SOCS-6) was seen with other SOCS/CIS family members. SOCS-1, SOCS-2, SOCS-3, and CIS each strongly inhibited the GH receptor (GHR)-dependent tyrosine phosphorylation of JAK2 seen at low levels of transfected JAK2; however, only SOCS-1 strongly inhibited the GHR-independent tyrosine phosphorylation of JAK2 seen at higher JAK2 levels. To probe for interactions with GHR, in vitro binding assays were carried out using glutathione S-transferase-GHR fusion proteins containing variable lengths of GHR's COOH-terminal cytoplasmic domain. CIS and SOCS-2 bound to fusions containing as few as 80 COOH-terminal GHR residues, provided the fusion protein was tyrosine-phosphorylated. By contrast, SOCS-3 binding required tyrosine-phosphorylated GHR membrane-proximal sequences, SOCS-1 binding was tyrosine phosphorylation-independent, and SOCS-6 did not bind the GHR fusion proteins at all. Mutation of GHR's membrane-proximal tyrosine residues 333 and 338 to phenylalanine suppressed the inhibition by SOCS-3, but not by CIS, of GH signaling to STAT5b. SOCS/CIS proteins can thus inhibit GH signaling to STAT5b by three distinct mechanisms, distinguished by their molecular targets within the GHR-JAK2 signaling complex, as exemplified by SOCS-1 (direct JAK2 kinase inhibition), SOCS-3 (inhibition of JAK2 signaling via membrane-proximal GHR tyrosines 333 and 338), and CIS and SOCS-2 (inhibition via membrane-distal tyrosine(s)).

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Animals
  • Carrier Proteins / metabolism*
  • Cell Line
  • Cloning, Molecular
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Growth Hormone / pharmacology*
  • Humans
  • Hypophysectomy
  • Immediate-Early Proteins / metabolism*
  • Intracellular Signaling Peptides and Proteins*
  • Male
  • Milk Proteins*
  • Proteins / metabolism*
  • Rats
  • Rats, Inbred F344
  • Receptors, Somatotropin / drug effects
  • Receptors, Somatotropin / physiology*
  • Recombinant Fusion Proteins / drug effects
  • Recombinant Fusion Proteins / metabolism
  • Repressor Proteins*
  • STAT5 Transcription Factor
  • Signal Transduction / drug effects
  • Signal Transduction / physiology*
  • Suppressor of Cytokine Signaling 1 Protein
  • Suppressor of Cytokine Signaling 3 Protein
  • Suppressor of Cytokine Signaling Proteins
  • Trans-Activators / genetics
  • Trans-Activators / metabolism*
  • Transcription Factors*
  • Transfection
  • src Homology Domains

Substances

  • Carrier Proteins
  • DNA-Binding Proteins
  • Immediate-Early Proteins
  • Intracellular Signaling Peptides and Proteins
  • Milk Proteins
  • Proteins
  • Receptors, Somatotropin
  • Recombinant Fusion Proteins
  • Repressor Proteins
  • SOCS1 protein, human
  • SOCS2 protein, human
  • SOCS3 protein, human
  • STAT5 Transcription Factor
  • STAT5B protein, human
  • Socs1 protein, rat
  • Socs2 protein, rat
  • Socs3 protein, rat
  • Stat5b protein, rat
  • Suppressor of Cytokine Signaling 1 Protein
  • Suppressor of Cytokine Signaling 3 Protein
  • Suppressor of Cytokine Signaling Proteins
  • Trans-Activators
  • Transcription Factors
  • cytokine inducible SH2-containing protein
  • Growth Hormone