The cytokine Interferon (IFN) (/( mediates its antiviral, antiproliferative, and immunomodulatory effects through the engagement of the Jak/Stat signaling pathway. Ligation of the IFN(/( receptor results in activation of Jak tyrosine kinases and subsequent tyrosine phosphorylation of Stat1. Phosphorylated Stat1 either forms homodimers or heterodimerizes with Stat2 via reciprocal SH2 domain interactions. Dimerized Stats then translocate to the nucleus where they bind to regulatory elements in IFN response genes. In addition to tyrosine kinases, protein arginine methyltransferases (PRMT) have been implicated in IFN(/( signaling pathways through their physical association with members of the Jak/Stat pathway and through genetic studies. The Hepatitis C Virus interferes with IFN signaling by targeting arginine methylation of Stat1. We have now found that cells deficient in the Carm1 arginine methyltransferase exhibited elevated responses to IFN(-driven Stat1 activation, including a deficiency in Stat1 dephosphorylation and. Furthermore, Carm1 can methylate Stat1 in in vitro assays. Our HYPOTHESIS is that Carm1 negatively regulates Stat1 driven transcription by promoting the dephosphorylation of Stat1 by TCPTP.
Specific Aims : 1) Define the intersection of Carm1 within IFN1/2 signaling pathways. We will examine Stat1 DNA binding, tetramerization, and association with target genes. We will also characterize the activity of Jak1, Tyk2, Stat1, Stat2, Stat3, and Stat5. 2) Determine the mechanism by which Carm1 regulates Stat1 dephosphorylation, including examining the importance of Carm1 catalytic activity and the effects on Carm1 on Stat1 interaction with Pias1 and TCPTP. The Stat1 methylation site will be determined. 3) Investigate the regulation of Carm1 by the IFN(/( signaling pathway, including expression, enzymatic activity, subcellular localization, and posttranslational modifications. Significance. These studies will provide insight into the mechanisms by which PRMTs regulate Type I IFN signaling, which may be altered during viral infection.
The interferon family of cytokines control pathogen infections through regulation of cellular proliferation and antiviral defenses, as well as through direct modulation of the immune response. For these reasons, the interferons are used clinically to treat viral and malignant diseases. Our project will address the molecular mechanisms of by which interferon signals are attenuated.
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