Regulation of PD-1 Gene Expression

程序性死亡-1（PD-1）在免疫激活后短暂出现在T细胞表面， 在长期暴露于抗原的T细胞中高度表达。虽然PD-1可能在 控制对自身抗原的反应，有令人信服的证据表明PD-1介导的抑制信号也 在慢性病毒感染期间T细胞的功能衰竭中发挥核心作用。这是 在包括HIV在内的几种动物和人类病毒感染中得到证实。最值得注意的是， PD-1与其配体之间的相互作用导致“耗尽的”抗原特异性T细胞的再活化。这种效果 已经在许多持续性病毒感染中得到证实，包括人类和 淋巴细胞性脉络膜脑膜炎病毒（LCMV）感染小鼠。这些数据强烈表明PD-1比 而不仅仅是T细胞耗竭的标志，而是这种表型的关键介质。这些数据还表明， 操纵PD-1表达、信号传导和/或功能可能导致新的基于免疫的疗法， 治疗慢性感染。令人惊讶的是，事实上对PD-1基因表达的调控一无所知。 在分子水平上，PD-1生物学和免疫学的这一基本原则将在本提案中阐明。 我们的初步数据提供了证据表明PD-1在多个水平上受到调节。运用比较 基因组学，DNase 1超敏试验，基因报告试验，和亚硫酸氢盐序列，我们已经确定 小鼠和人类基因组中可能参与PD-1调控的区域， 将其中两个区域与转录活性相关联。我们还发现证据表明PD-1 基因在体内病毒特异性CDS T细胞中差异甲基化，表明表观遗传控制的作用 的基因。本申请中的实验将鉴定顺式作用元件，Aim 1;确定顺式作用元件的分子量; 表观遗传机制的广度和动态，目的2;并确定转录因子， 与PD-1基因调控相关，目的3。在所有的目标中，我们将开始与模型细胞系，确认 并在原代小鼠和人类CDS T细胞中验证该信息，测试这些发现的有效性。 LCMV感染后抗原特异性CDS T细胞;并检查HIV特异性CDS T细胞以确定 PD-1调节如何影响HIV疾病进展。这些方法将提供分子靶点 对于可能最终用于治疗HIV疾病、慢性感染、自身免疫和帮助 移植成功。

Programmed death-1 (PD-1) appears transiently on the surface of T cells following immune activation and is highly expressed in T cells chronically exposed to antigen. While PD-1 likely plays an important role in governing responses to self antigen, there is compelling evidence that PD-1-mediated inhibitory signals also play a central role in the functional exhaustion of T cells during chronic viral infection. This has been demonstrated in several animal and human viral infections including HIV. Most notably, antibody blockade between PD-1 and its ligands results in reactivation of the "exhausted" antigen-specific T cells. This effect has been demonstrated in a number of persistent viral infections, including those from HIV in humans and in the mouse with lymphocytic choriomenigitis virus (LCMV). These data strongly suggest that PD-1 is more than just a marker of T cell exhaustion but rather a critical mediator of this phenotype. These data also imply that manipulating PD-1 expression, signaling, and/or function could lead to novel immune based therapies to treat chronic infection. Surprisingly, virtually nothing is known about the regulation of PD-1 gene expression at the molecular level ¿ this basic tenet of PD-1 biology and immunology will be elucidated in this proposal. Our preliminary data provide evidence that PD-1 Is regulated at multiple levels. Using comparative genomics, DNasel hypersensitivity assays, gene reporter assays, and bisulfite sequence, we have identified regions in both mouse and human genomes that are likely to be involved in the regulation of PD-1 and have correlated two of these regions with transcriptional activity. We have also found evidence that the PD-1 gene is differentially methylated in viral-specific CDS T cells in vivo, suggesting a role for epigenetic control of the gene. Experiments in this application will identify the cis-acting elements, Aim 1; determine the breadth and dynamics of epigenetic mechanisms, Aim 2; and identify transcription factors that are associated with PD-1 gene regulation, Aim 3. In all of the aims we will begin with model cell lines, confirm and verify that information in primary mouse and human CDS T cells, test the validity .of these findings in antigen-specific CDS T cell following LCMV infection; and examine HIV-specific CDS T cells to determine how PD-1 regulation factors into HIV disease progression. These approaches will provide molecular targets for pathways that may be ultimately used for treating HIV disease, chronic infection, autoimmunity and aiding to transplantation success.