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Nuclear transit of the intracellular domain of the interferon receptor subunit IFNaR2 requires Stat2 and Irf9.

ABSTRACT: Regulated intramembrane proteolysis (RIP) is the primary signaling mechanism for some receptors, such as Notch and the amyloid precursor protein. In addition, some receptor type tyrosine kinases, such as HER4, are able to signal via both kinase activation and regulated receptor proteolysis. Previously, we showed that the IFNaR2 subunit of the type I interferon receptor can be cleaved in a two step process that resembles RIP and that the IFNaR2 intracellular domain (IFNaR2-ICD) can mediate gene transcription in a Stat2 dependent manner. Here, we demonstrate that IFNaR2-ICD, Stat2 and Irf9 form a ternary complex. Furthermore, Stat2 and Irf9 are required for the nuclear transit of a GFP-linked IFNaR2-ICD construct (GFP-ICD). Additional experiments monitoring the nuclear localization of GFP-ICD demonstrate that Stat2 serves an adaptor role, mediating the interaction between the IFNaR2-ICD and Irf9, while the bipartite nuclear localization signal within Irf9 is the primary determinant driving nuclear transit of the ICD containing complex. Overall, the data suggest that liberation of the IFNaR2-ICD by regulated proteolysis could trigger a novel mechanism for moving the transcription factor Stat2 to the nucleus.

SUBMITTER: El Fiky A

PROVIDER: S-EPMC2494602 | biostudies-literature | 2008 Jul

REPOSITORIES: biostudies-literature

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Nuclear transit of the intracellular domain of the interferon receptor subunit IFNaR2 requires Stat2 and Irf9.

El Fiky Ashraf A Pioli Pete P Azam Arif A Yoo Kiwon K Nastiuk Kent L KL Krolewski John J JJ

Cellular signalling 20080321 7

Regulated intramembrane proteolysis (RIP) is the primary signaling mechanism for some receptors, such as Notch and the amyloid precursor protein. In addition, some receptor type tyrosine kinases, such as HER4, are able to signal via both kinase activation and regulated receptor proteolysis. Previously, we showed that the IFNaR2 subunit of the type I interferon receptor can be cleaved in a two step process that resembles RIP and that the IFNaR2 intracellular domain (IFNaR2-ICD) can mediate gene t ...[more]

PMID: 18456457

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Project description:Measles virus (MeV) is a highly immunotropic and contagious pathogen that can even diminish preexisting antibodies and remains a major cause of childhood morbidity and mortality worldwide despite the availability of effective vaccines. MeV is one of the most extensively studied viruses with respect to the mechanisms of JAK-STAT antagonism. Of the three proteins translated from the MeV P gene, P and V are essential for inactivation of this pathway. However, the lack of data from direct analyses of the underlying interactions means that the detailed molecular mechanism of antagonism remains unresolved. Here, we prepared recombinant MeV V protein, which is responsible for human JAK-STAT antagonism, and a panel of variants, enabling the biophysical characterization of V protein, including direct V/STAT1 and V/STAT2 interaction assays. Unambiguous direct interactions between the host and viral factors, in the absence of other factors such as Jak1 or Tyk2, were observed, and the dissociation constants were quantified for the first time. Our data indicate that interactions between the C-terminal region of V and STAT2 is 1 order of magnitude stronger than that of the N-terminal region of V and STAT1. We also clarified that these interactions are completely independent of each other. Moreover, results of size exclusion chromatography demonstrated that addition of MeV-V displaces STAT2-core, a rigid region of STAT2 lacking the N- and C-terminal domains, from preformed complexes of STAT2-core/IRF-associated domain (IRF9). These results provide a novel model whereby MeV-V can not only inhibit the STAT2/IRF9 interaction but also disrupt preassembled interferon-stimulated gene factor 3.IMPORTANCE To evade host immunity, many pathogenic viruses inactivate host Janus kinase signal transducer and activator of transcription (STAT) signaling pathways using diverse strategies. Measles virus utilizes P and V proteins to counteract this signaling pathway. Data derived largely from cell-based assays have indicated several amino acid residues of P and V proteins as important. However, biophysical properties of V protein or its direct interaction with STAT molecules using purified proteins have not been studied. We have developed novel molecular tools enabling us to identify a novel molecular mechanism for immune evasion whereby V protein disrupts critical immune complexes, providing a clear strategy by which measles virus can suppress interferon-mediated antiviral gene expression.

Project description:Foot-and-mouth disease virus (FMDV) causes a highly contagious, debilitating disease in cloven-hoofed animals with devastating economic consequences. To survive in the host, FMDV has evolved to antagonize the host type I interferon (IFN) response. Previous studies have reported that the leader proteinase (L(pro)) and 3C(pro) of FMDV are involved in the inhibition of type I IFN production. However, whether the proteins of FMDV can inhibit type I IFN signaling is less well understood. In this study, we first found that 3C(pro) of FMDV functioned to interfere with the JAK-STAT signaling pathway. Expression of 3C(pro) significantly reduced the transcript levels of IFN-stimulated genes (ISGs) and IFN-stimulated response element (ISRE) promoter activity. The protein level, tyrosine phosphorylation of STAT1 and STAT2, and their heterodimerization were not affected. However, the nuclear translocation of STAT1/STAT2 was blocked by the 3C(pro) protein. Further mechanistic studies demonstrated that 3C(pro) induced proteasome- and caspase-independent protein degradation of karyopherin ?1 (KPNA1), the nuclear localization signal receptor for tyrosine-phosphorylated STAT1, but not karyopherin ?2, ?3, or ?4. Finally, we showed that the protease activity of 3C(pro) contributed to the degradation of KPNA1 and thus blocked STAT1/STAT2 nuclear translocation. Taken together, results of our experiments describe for the first time a novel mechanism by which FMDV evolves to inhibit IFN signaling and counteract host innate antiviral responses.We show that 3C(pro) of FMDV antagonizes the JAK-STAT signaling pathway by blocking STAT1/STAT2 nuclear translocation. Furthermore, 3C(pro) induces KPNA1 degradation, which is independent of proteasome and caspase pathways. The protease activity of 3C(pro) contributes to the degradation of KPNA1 and governs the ability of 3C(pro) to inhibit the JAK-STAT signaling pathway. This study uncovers a novel mechanism evolved by FMDV to antagonize host innate immune responses.

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Nuclear transit of the intracellular domain of the interferon receptor subunit IFNaR2 requires Stat2 and Irf9.

Publications

Nuclear transit of the intracellular domain of the interferon receptor subunit IFNaR2 requires Stat2 and Irf9.

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