Tax regulation of HTLV-I Gene Expression through CBP

人类T细胞白血病病毒1型(HTLV-1)是一种复杂的逆转录病毒，其病因与一种称为成人T细胞白血病(ATL)的侵袭性且通常是致命的恶性肿瘤有关。HTLV-I相关肿瘤发生的分子基础与病毒编码的Tax蛋白的表达有关。HTLV-I基因组的强健转录需要与细胞转录因子CREB(或ATF/CREB家族的其他成员)合作的TAX。 位于HTLV-I转录控制区的三个21碱基对重复序列增强子元件对Tax激活的转录起关键作用。这些元件被称为病毒Cres，是Tax/CREB复合体的结合位点。这种含税启动子结合复合体的形成对多功能细胞共激活因子CBP/p300的招募至关重要。CBP和p300是非常大的、高度相关的蛋白质，具有乙酰转移酶活性，并协调后生动物中高度调控的基因表达。CBP和p300中的几个保守结构域是多种细胞和病毒转录因子的结合位点。由这些结构域介导的转录因子共激活子相互作用促进了辅激活子的基因特异性招募。最近几个实验室的研究表明，CBP/p300的KIX结构域被利用，也许是排他性的 HTLV-I启动子共激活子的招募。这是通过将KIX直接结合到病毒Cre结合的Tax/CREB复合体来实现的。一旦招募，几项研究表明，CBP/p300通过辅活化子的乙酰转移酶活性介导税务反式激活。我们实验室最近的实验表明，乙酰转移酶的活性不是针对组蛋白的氨基末端。相反，我们发现包括Tax和CREB在内的非组蛋白是p300的直接乙酰化靶点。值得注意的是，当两种蛋白都与病毒Cre DNA形成复合体时，p300对Tax/CREB的乙酰化作用被显著地刺激。在本应用中，我们建议确定与CBP的KIX结构域结合的Tax/CREB/病毒Cre复合体的三维结构。这项工作将与卡罗琳·卢格博士合作进行，后者参与了核小体核心颗粒的结构测定。其他目标将主要集中在TAX/CREB的CBP/p300乙酰化的生化特征和功能分析上 很复杂。在真核细胞中，非组蛋白转录因子上赖氨酸残基的乙酰化正迅速成为一种重要的调控机制。

The human T-cell leukemia virus type-1 (HTLV-1) is acomplex retrovirus etiologically linked to an aggressive and generally fatal malignancy called adult Tcell leukemia (ATL). The molecular basis of HTLV-I associated oncogenesis is linked to the expression of the virally-encoded Tax protein. Robust transcription of the HTLV-I genome requires Tax in cooperation with the cellular transcription factor CREB (or other members of the ATF/CREB family). Three 21 base pair repeat enhancer elements, located in the HTLV-I transcriptional control region, are critical to Tax-activated transcription. These elements, which are referred to as viral CREs, are binding sites for the Tax/CREB complex. The formation of this Tax-containing promoter-bound complex is critical in the recruitment of the multifunctional cellular coactivators CBP/p300. CBP and p300 are very large, highly related proteins that posses acetyltransferase activity and coordinate highly regulated gene expression in metazoans. Several conserved domains in CBP and p300 serve as binding sites for a wide variety of cellular and viral transcription factors. The transcription factor coactivator interactions mediated by these domains facilitate gene-specific recruitment of the coactivators. Recent studies in several laboratories have demonstrated the KIX domain of CBP/p300 is utilized, perhaps exclusive, in the recruitment of the coactivators to the HTLV-I promoter. This is accomplished via direct binding of KIX to the viral CRE-bound Tax/CREB complex. Once recruited, several studies have shown that CBP/p300 mediate Tax-transactivation via the acetyltransferase activity of the coactivators. Recent experiments in our laboratory demonstrate that the acetyltransferase activity is not directed at the histone amino terminal tails. Instead, we find that non-histone proteins, including Tax and CREB, are the direct acetylation targets of p300. Significantly, Tax/CREB acetylation by p300 is dramatically stimulated when both proteins are in complex with the viral CRE DNA. In this application, we propose to determine the three dimensional structure of the Tax/CREB/viral CRE complex bound to the KIX domain of CBP. This will be carried out in collaboration with Dr. Karolin Luger, who participated in the structure determination of the nucleosome core particle. The additional aims will focus primarily on the biochemical characterization and functional analysis of CBP/p300 acetylation of the Tax/CREB complex. Acetylation of lysine residues on nonhistone transcription factors is rapidly emerging as a prominent regulatory mechanism in eukaryotic cells.