Regulation of the HIV cofactor activity of LEDGF/p75 by interacting proteins

描述(申请人提供)：艾滋病毒感染需要将病毒基因组的DNA拷贝整合到宿主DNA中。因此，防止艾滋病毒DNA整合的治疗策略对于阻止艾滋病毒感染至关重要。细胞因子在这一病毒过程中的作用尚不清楚。晶状体上皮源性生长因子p75(LEDGF/p75)是HIV DNA整合的细胞辅因子。然而，它仍然没有解释为什么慢病毒进化依赖于这种细胞蛋白来有效整合病毒DNA。我的团队和其他人的工作已经确定，LEDGF/p75将HIV整合复合体连接到宿主染色质。然而，这一机制并没有解释染色质结合的HIV前整合复合体为了获得宿主DNA而使用的策略，这是整合到宿主DNA的必要步骤，或者进一步利用细胞DNA修复机制来促进整合后DNA修复。来自我的实验室的新发现为HIV DNA整合过程的这些基本方面提供了潜在的分子解释。我们首次证明了LEDGF/p75中存在几个蛋白质基序，这些蛋白质基序是HIV DNA以染色质拴系不依赖的方式整合所必需的。这些基序包含被磷酸化或SUMO化的残基，或者被预测为蛋白质相互作用模块的一部分。我们推测这些LEDGF/p75基序参与招募与HIV DNA整合过程有关的细胞蛋白。为了进一步支持这一模型，我们的实验室已经确定了新的LEDGF/p75相互作用蛋白，这些蛋白参与染色质重塑、DNA修复和转录延长。综上所述，我们的模型假设慢病毒已经进化到依赖LEDGF/p75的最佳HIV DNA整合，因为这种细胞蛋白：(I)将HIV预整合复合体拴在参与活跃转录的染色质亚区，以及(Ii)通过LEDGF/p75特异的蛋白质-蛋白质相互作用，为病毒提供进入宿主DNA和DNA修复机制的途径，特别是在活跃转录DNA方面。此外，我们推测一些与LEDGF/p75相互作用的蛋白会削弱该蛋白的HIV辅因子活性。在这份提案中，我们将展示这一新模式的基本方面。具体地说，我们将：(I)确定与DNA修复和染色质重塑有关的LEDGF/p75相互作用蛋白在HIV DNA整合中的作用。(2)确定参与翻译后修饰和蛋白质-蛋白质相互作用的LEDGF/p75基序在艾滋病毒DNA整合中的功能影响。(Iii)通过在全基因组范围内确定LEDGF/p75位置与HIV DNA整合位点选择之间的相关性，为我们的模型提供进一步支持。

DESCRIPTION (provided by applicant): HIV infection requires the integration of a DNA copy of the viral genome into the host DNA. Therefore therapeutic strategies to prevent HIV DNA integration are essential to block HIV infection. The role of cellular factors in this viral process is poorly defined. The Lens Epithelium-Derived Growth Factor p75 (LEDGF/p75) is a well characterized cellular cofactor of HIV DNA integration. However, it remains unexplained why Lentiviruses have evolved dependency of this cellular protein for efficient viral DNA integration. Work from my group and others have determined that LEDGF/p75 tethers the HIV integration complex to the host chromatin. However, this mechanism does not explain the strategies used by the chromatin-bound HIV pre-integration complex to gain access to the host DNA, a necessary step for integration to the host DNA, or to exploit further the cellular DNA repair machinery to promote post-integration DNA repair. New findings from my laboratory provide potential molecular explanations for these essential aspects of the HIV DNA integration process. We have demonstrated for the first time the existence of several protein motifs in LEDGF/p75 necessary for the HIV DNA integration in a chromatin tethering-independent manner. These motifs contain residues that are phosphorylated or SUMOylated, or that are predicted to be part of protein-protein interaction modules. We have postulated that these LEDGF/p75 motifs are involved in recruiting cellular proteins implicated in the HIV DNA integration process. In further support of this model, our laboratory has identified novel LEDGF/p75- interacting proteins known to be involved in chromatin remodeling, DNA repair, and transcriptional elongation. In summary, our model postulate that Lentiviruses have evolved LEDGF/p75-dependency for optimal HIV DNA integration because this cellular protein: (i) tethers the HIV pre-integration complex to subregions of the chromatin engaged in active transcription, and (ii) provides to the virus, through LEDGF/p75-specific protein- protein interactions, access to the host DNA and to DNA repair mechanisms particularly active in actively transcribe DNA. In addition, we postulate that some LEDGF/p75-interacting proteins will impair the HIV cofactor activity of this protein. In this proposal we will demonstrate essential aspects of this novel model. Specifically we will: (i) Define the role in HIV DNA integration of LEDGF/p75-interacting proteins implicated in DNA repair and chromatin remodeling. (ii) Determine the functional implications in HIV DNA integration of LEDGF/p75 motifs involved in post-translational modifications and protein-protein interactions. (iii) Provide further support to our model by determining genome-wide the correlation between LEDGF/p75 location and HIV DNA integration site selection.