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，在病因学上与一种称为成人T细胞白血病（ATL）的侵袭性和通常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基因组的稳健转录需要Tax与细胞转录因子CREB（或ATF/CREB家族的其他成员）合作。 三个21碱基对重复的增强子元件，位于HTLV-1转录控制区，是Tax激活转录的关键。这些元件被称为病毒克雷斯，是Tax/CREB复合物的结合位点。这种含Tax的启动子结合复合物的形成在多功能细胞共激活因子CBP/p300的募集中是至关重要的。CBP和p300是非常大的，高度相关的蛋白质，其抑制乙酰转移酶活性并协调后生动物中高度调节的基因表达。CBP和p300中的几个保守结构域作为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因子相互作用促进共激活因子的基因特异性募集。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， 共激活剂向HTLV-I启动子的募集。这是通过KIX与病毒CRE结合的Tax/CREB复合物的直接结合来实现的。一旦被招募，一些研究已经表明CBP/p300通过共激活因子的乙酰转移酶活性介导Tax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氨基末端的尾巴。相反，我们发现非组蛋白蛋白，包括Tax和CREB，是p300的直接乙酰化靶点。值得注意的是，当两种蛋白质与病毒CRE DNA复合时，通过p300的Tax/CREB乙酰化被显著刺激。在本申请中，我们提出确定与CBP的KIX结构域结合的Tax/CREB/病毒CRE复合物的三维结构。这将与Karolin Luger博士合作进行，他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CBP/p300乙酰化Tax/CREB的生化表征和功能分析上。 复杂.非组蛋白转录因子上赖氨酸残基的乙酰化正迅速成为真核细胞中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.