Human T-cell lymphotropic virus type 1 (HTLV-1) Tax oncoprotein is required for viral gene expression. Tax transactivates the viral promoter by recruiting specific transcription factors but also by interfering with general transcription factors involved in the preinitiation step, such as TFIIA and TFIID. However, data are lacking regarding Tax interplay with TFIIH, which intervenes during the last step of preinitiation. We previously reported that XPB, the TFIIH subunit responsible for promoter opening and promoter escape, is required for Tat-induced human-immunodeficiency virus promoter transactivation. Here, we investigated whether XPB may also play a role in HTLV-1 transcription. We report that Tax and XPB directly interact in vitro and that endogenous XPB produced by HTLV-1-infected T cells binds to Tax and is recruited on proviral LTRs. In contrast, XPB recruitment at the LTR is not detected in Tax-negative HTLV-1-infected T cells and is strongly reduced when Tax-induced HTLV-1 LTR transactivation is blocked. XPB overexpression does not affect basal HTLV-1 promoter activation but enhances Tax-mediated transactivation in T cells. Conversely, downregulating XPB strongly reduces Tax-mediated transactivation. Importantly, spironolactone (SP)-mediated inhibition of LTR activation can be rescued by overexpressing XPB but not XPD, another TFIIH subunit. Furthermore, an XPB mutant defective for the ATPase activity responsible for promoter opening does not show rescue of the effect of SP. Finally, XPB downregulation reduces viability of Tax-positive but not Tax-negative HTLV-1-transformed T cell lines. These findings reveal that XPB is a novel cellular cofactor hijacked by Tax to facilitate HTLV-1 transcription.IMPORTANCE HTLV-1 is considered the most potent human oncovirus and is also responsible for severe inflammatory disorders. HTLV-1 transcription is undertaken by RNA polymerase II and is controlled by the viral oncoprotein Tax. Tax transactivates the viral promoter first via the recruitment of CREB and its cofactors to the long terminal repeat (LTR). However, how Tax controls subsequent steps of the transcription process remains unclear. In this study, we explore the link between Tax and the XPB subunit of TFIIH that governs, via its ATPase activity, the promoter-opening step of transcription. We demonstrate that XPB is a novel physical and functional partner of Tax, recruited on HTLV-1 LTR, and required for viral transcription. These findings extend the mechanism of Tax transactivation to the recruitment of TFIIH and reinforce the link between XPB and transactivator-induced viral transcription.

中文翻译：

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人类T细胞淋巴病毒1型反式激活因子税在病毒转录过程中利用了TFIIH的XPB亚基。

病毒基因表达需要1型人类T细胞淋巴病毒（HTLV-1）税收癌蛋白。Tax通过募集特定的转录因子，也通过干扰预启动步骤中涉及的一般转录因子（例如TFIIA和TFIID）来使病毒启动子反式激活。但是，缺少有关税收与TFIIH相互作用的数据，TFIIH在预初始化的最后一步进行干预。我们先前曾报道XPA，即负责启动子打开和启动子逃逸的TFIIH亚基，是Tat诱导的人类免疫缺陷病毒启动子反式激活所必需的。在这里，我们调查XPB是否也可能在HTLV-1转录中起作用。我们报告说，税收和XPB直接在体外相互作用，并且由HTLV-1感染的T细胞产生的内源性XPB与税收结合，并在前病毒LTRs上募集。相反，在Tax阴性的HTLV-1感染的T细胞中未检测到LTR处的XPB募集，并且在阻止Tax诱导的HTLV-1 LTR反式激活时，XPB的募集会大大减少。XPB的过表达不会影响基础HTLV-1启动子的激活，但会增强T细胞中Tax介导的反式激活。相反，下调XPB会大大减少Tax介导的反式激活。重要的是，螺内酯（SP）介导的LTR激活抑制可通过过量表达XPB而非XPD（另一种TFIIH亚基）来挽救。此外，负责启动子打开的ATPase活性有缺陷的XPB突变体未显示出SP作用的挽救。最后，XPB的下调降低了Tax-阳性但未阴性的HTLV-1转化T细胞系的活力。这些发现表明XPB是被Tax劫持以促进HTLV-1转录的新型细胞辅因子。重要提示HTLV-1被认为是最有效的人类肿瘤病毒，也可导致严重的炎症性疾病。HTLV-1转录由RNA聚合酶II进行，并受病毒癌蛋白税的控制。税收首先通过募集CREB及其辅因子至长末端重复序列（LTR）使病毒启动子反式激活。但是，税收如何控制转录过程的后续步骤仍不清楚。在这项研究中，我们探索了Tax和TFIIH的XPB亚基之间的联系，后者通过其ATPase活性控制转录的启动子开放步骤。我们证明XPB是Tax的新型物理和功能伙伴，在HTLV-1 LTR上募集，并且是病毒转录所必需的。