Mechanisms of HTLV-I Tax-mediated NF-kB activation

描述（由申请人提供）：人T细胞白血病病毒I型（HTLV-I）与成人T细胞白血病（ATL）的发生相关，ATL是一种侵袭性CD 4 + T细胞恶性肿瘤。Tax是由HTLV-I基因组编码的癌蛋白，其调节病毒和细胞基因表达。Tax通过激活NF-κ B/Rel转录因子家族促进细胞转化。Tax需要NF-kB用于原代T细胞的永生化，并且HTLV-I转化的细胞依赖于NF-kB用于它们的存活，从而为研究Tax用于激活NF-kB的机制提供了强有力的理论基础。Tax通过促进持续性IkB激酶（IKK）激活来激活NF-kB，尽管其机制尚不清楚。我们的支持性研究表明，多聚泛素化是NF-κ B Tax激活的关键翻译后调节因子。NF-κ B的税务激活与IKK在高尔基体的泛素依赖性重定位相关。Tax通过Lys 63连接的泛素链被多聚泛素化，并且需要泛素缀合酶Ubc 13用于Tax泛素化和NF-κ B活化。Tax的泛素化不促进其降解，而是调节其与IKK调节亚基IKKg的相互作用。此外，我们已经确定，Tax相互作用细胞蛋白TAX 1BP 1是一个关键的负调节剂，通过作为泛素编辑酶A20的一个必需亚基发挥作用，从而介导的NF-κ B信号转导。我们假设Tax靶向TAX 1BP 1作为促进持续NF-kB激活机制的一部分。在这个提议中，我们将进一步研究泛素化如何调节Tax和IKK进入高尔基体的靶向。我们还将进一步阐明调控Tax泛素化的宿主因素的要求和识别。最后，我们将确定Tax用于抑制TAX 1BP 1和A20泛素编辑复合物的机制。我们的目标是通过完成以下具体目标来实现：（1）确定Tax介导的IKK再定位机制;（2）研究Tax泛素化和NF-κ B激活的分子决定因素;（3）确定Tax与TAX 1BP 1结合在NF-κ B激活中的作用。这些研究将阐明逆转录病毒癌蛋白如何篡夺宿主泛素化/去泛素化机制，持续激活促进细胞转化的转录因子。公共卫生相关性：HTLV-I Tax癌蛋白是NF-κ B/Rel转录因子家族的有效激活剂。Tax需要NF-kB用于T细胞的永生化，并且NF-kB也是HTLV-1转化细胞系的存活所需的。该提案的重点是确定Tax激活NF-kB的分子机制，具体来说，我们将进一步确定Tax泛素化的决定因素，并研究Tax与细胞蛋白TAX 1BP 1相互作用的作用。

DESCRIPTION (provided by applicant): Human T cell leukemia virus type I (HTLV-I) is linked to the genesis of adult T cell leukemia (ATL), an aggressive CD4+ T cell malignancy. Tax is an oncoprotein encoded by the HTLV-I genome that regulates viral and cellular gene expression. Tax promotes cell transformation, in part, by activating the NF-kB/Rel transcription factor family. Tax requires NF-kB for the immortalization of primary T cells and HTLV-I transformed cells are dependent on NF-kB for their survival, thereby providing a strong rationale for studying the mechanisms used by Tax to activate NF-kB. Tax activates NF-kB by promoting persistent IkB kinase (IKK) activation although the mechanisms are poorly understood. Our supporting studies indicate that polyubiquitination is a key post- translational regulator of Tax activation of NF-kB. Tax activation of NF-kB correlates with ubiquitin-dependent relocalization of IKK to the Golgi apparatus. Tax is polyubiquitinated via Lys 63-linked ubiquitin chains and requires the ubiquitin conjugating enzyme Ubc13 for Tax ubiquitination and NF-kB activation. Ubiquitination of Tax does not promote its degradation, but rather regulates its interaction with the IKK regulatory subunit IKKg. In addition, we have determined that the Tax interacting cellular protein TAX1BP1 is a key negative regulator of cytokine-mediated NF-kB signaling by functioning as an essential subunit for the ubiquitin-editing enzyme A20. We hypothesize that Tax targets TAX1BP1 as part of a mechanism to promote persistent NF-kB activation. In this proposal, we will further investigate how ubiquitination regulates the targeting of Tax and IKK into the Golgi. We will also further delineate the requirements and the identification of host factors regulating Tax ubiquitination. Finally, we will determine the mechanisms used by Tax to inactivate TAX1BP1 and the A20 ubiquitin-editing complex. Our objectives for this proposal will be satisfied by completion of the following specific aims: (1) Determine the mechanisms of Tax-mediated relocalization of IKK; (2) investigate the molecular determinants of Tax ubiquitination and NF-kB activation and (3) determine the role of Tax binding to TAX1BP1 in NF-kB activation. These studies will shed light on how a retroviral oncoprotein usurps host ubiquitination/deubiquitination machinery to persistently activate transcription factors that promote cell transformation. PUBLIC HEALTH RELEVANCE: The HTLV-I Tax oncoprotein is a potent activator of the NF-kB/Rel transcription factor family. Tax requires NF-kB for the immortalization of T cells, and NF-kB is also required for the survival of HTLV-1 transformed cell lines. The focus of the proposal is to determine the molecular mechanisms used by Tax to activate NF-kB, specifically we will further establish the determinants of Tax ubiquitination and examine the role of Tax interaction with the cellular protein TAX1BP1.