title: Role of tyrosine 441 of interferon-gamma receptor subunit 1 in SOCS-1-mediated attenuation of STAT1 activation.
abstract: Suppressor of cytokine signaling (SOCS)-1, the key negative regulator of interferon (IFN)-gamma-dependent signaling, is induced in response to IFNgamma. SOCS-1 binds to and inhibits the IFNgamma receptor-associated kinase Janus-activated kinase (JAK) 2 and inhibits its function in vitro, but the mechanism by which SOCS-1 inhibits IFNgamma-dependent signaling in vivo is not clear. Upon stimulation, mouse IFNgamma receptor subunit 1 (IFNGR1) is phosphorylated on several cytoplasmic tyrosine residues, and Tyr(419) is required for signal transducer and activator of transcription (STAT) 1 activation in mouse embryo fibroblasts. However, the functions of the other three cytoplasmic tyrosine residues are not known. Here we show that Tyr(441) is required to attenuate STAT1 activation in response to IFNgamma. Several tyrosine to phenylalanine mutants of IFNGR1, expressed at normal levels in stable pools of IFNGR1-null cells, were analyzed for the phosphorylation of STAT1 during a 48-h period, and antiviral activity in response to IFNgamma was also measured. Stronger activation of STAT1 was observed in cells expressing all IFNGR1 variants mutated at Tyr(441), and, consistently, stronger antiviral activity was also observed in these cells. Furthermore, constitutive overexpression of SOCS-1 inhibited IFNgamma-dependent signaling only in cells expressing IFNGR1 variants that included the Tyr(441) mutation. Mutation of Tyr(441) also blocked the ability of SOCS-1 to bind to IFNGR1 and JAK2 in response to IFNgamma and the normal down-regulation of STAT1 activation and antiviral activity. These results, together with data from the literature, suggest a model in which, in response to IFNgamma, phosphorylation of Tyr(441) creates a docking site for SOCS-1, which then binds to JAK2 within the receptor-JAK complex to partially inhibit JAK2 phosphorylation. Furthermore, the virtually complete blockade of STAT1 phosphorylation by overexpressed SOCS-1 in this experiment suggests that the binding of SOCS-1 to Tyr(441) also blocks the access of STAT1 to Tyr(419) and that this effect may be the principal mechanism of inhibition of downstream signaling.
authors.affiliation: Department of Molecular Biology, Lerner Research Institute, Cleveland Clinic Foundation, Cleveland, Ohio 44195, USA.
authors.forename: Yulan
authors.initials: Y
authors.lastname: Qing
authors.forename: Ana P
authors.initials: AP
authors.lastname: Costa-Pereira
authors.forename: Diane
authors.initials: D
authors.lastname: Watling
authors.forename: George R
authors.initials: GR
authors.lastname: Stark
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000502:physiology*; D002460:Cell Line; D004268:DNA-Binding Proteins / Q000378:metabolism*; D047468:Immunoprecipitation; D051379:Mice; D017471:Receptors, Interferon / Q000737:chemistry / Q000502:physiology*; D012097:Repressor Proteins / Q000502:physiology*; D050794:STAT1 Transcription Factor; D000071222:Suppressor of Cytokine Signaling 1 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D015534:Trans-Activators / Q000378:metabolism*; D014443:Tyrosine / Q000502:physiology*; D000096743:Interferon gamma Receptor
chemical_list: D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D017471:Receptors, Interferon; D012097:Repressor Proteins; D050794:STAT1 Transcription Factor; D000071222:Suppressor of Cytokine Signaling 1 Protein; D050826:Suppressor of Cytokine Signaling Proteins; D015534:Trans-Activators; D014443:Tyrosine; D000096743:Interferon gamma Receptor; C491590:Socs1 protein, mouse; C494080:Stat1 protein, mouse