TYK2 AND IFNAR1 IN INTERFERON ALPHA SIGNALING

JAK/STAT信号通路在细胞表面的配体结合和细胞核内的基因转录之间提供了一种相对简单和直接的联系。这些途径介导了对多种多肽配体的反应，这些多肽配体调节细胞的生长、分化和效应器功能。最具代表性的JAK/STAT信号级联通路之一是干扰素-α(IFNpha)刺激的通路。许多实验室对这一信号转导级联的广泛研究提供了对调控IFNpha诱导基因转录的初始分子事件的基本了解，并为研究其他JAK-STAT通路建立了一个范例。在这笔赠款的前一期中，我们探索了这一级联中不同组成部分之间的分子相互作用：TYK2酪氨酸激酶、IFNAR1受体亚单位以及STAT1和STAT2转录因子。特别是，我们已经：i)鉴定和鉴定了TYK2激酶-IFNAR1受体复合体；ii)鉴定和鉴定了IFNaR1上的配体诱导的磷酸酪氨酸对接位点，它招募了Stat2的SH2结构域；iii)利用嵌合受体部分重组了IFNpha信号通路。然而，控制这一途径的分子相互作用的许多细节仍然未知，包括TYK2激酶的哪些部分是与IFNaR1相互作用所必需的，Stat2-STAT1异二聚体是如何从受体中释放出来的，IFNaR1亚单位远端部分的作用以及哪些特定的分子参与了这一途径的负调控。为了研究其中的一些细节，我们提出了四个特定的目标：1.确定TYK2中与IFNAR1受体亚单位相互作用所需的残基。2.明确了STAT1-STAT2异二聚体的形成机制。3.确定更多调控IFNpha信号的IFNAR1相互作用蛋白。4.研究CIS蛋白在TYK2蛋白和/或IFNaR1亚基调控中的作用。

JAK/STAT signaling cascades provide a relative simple and direct connection between ligand binding on the cell surface and gene transcription in the nucleus. These pathways mediate the response to a diverse group of polypeptide ligands which regulate cell growth, differentiation and effector function. One of the best characterized JAK/STAT signaling cascades is the pathway stimulated by interferon- alpha (IFNalpha). Extensive studies of this signal transduction cascade by a number of laboratories has provided a basic understanding of the initial molecular events regulating IFNalpha-induced gene transcription and has established a paradigm for studying other JAK-STAT pathways, as well. During the previous term of this grant, we have explored the molecular interactions between the various components in this cascade: the Tyk2 tyrosine kinase, the IFNaR1 receptor subunit, and the Stat1 and Stat2 transcription factors. In particular, we have: i) identified and characterized the Tyk2 kinase-IFNaR1 receptor complex; ii) identified and characterized the ligand-inducible Phosphotyrosine docking site on IFNaR1 which recruits the SH2 domain of Stat2; and iii) partially reconstituted the IFNalpha signaling pathway using chimeric receptors. Many details of the molecular interactions controlling this pathway, however, remain unknown, including which parts of the Tyk2 kinase are required for the interaction with IFNaR1, how the Stat2-Stat1 heterodimer is released from the receptor, the role of the distal portion of the IFNaR1 subunit and which specific molecules are involved in the negative regulation of this pathway. Four specific aims are proposed to investigate some of the these details: 1. Identify the residues in Tyk2 required for interaction with the IFNaR1 receptor subunit. 2. Define the mechanism of Stat1-Stat2 heterodimer formation. 3. Identify additional IFNaR1-interacting proteins which regulate IFNalpha signaling. 4. Investigate the role of CIS proteins in the regulation of the Tyk2 kinase and/or the IFNaR1 subunit.