Human Retrovirus Effects on Blood Cells

该项目的目标是更好地了解宿主细胞和其他辅助因子对人类逆转录病毒复制和发病机制的影响。高效联合抗病毒药物治疗（HAART）有望将生产性HIV感染转化为慢性、非病毒血症、非进展性疾病。然而，在一些患者中，已经观察到有限但连续的病毒产生，并且在其他患者中，已经在HAART治疗的HIV感染个体中鉴定出潜伏感染的长寿命外周T细胞，其可以在正常免疫应答期间重新激活。在开发用于HIV感染个体的长期疗法中，将需要通过辅助疗法如免疫疗法来控制来自这些潜伏储库的病毒产生和/或根除这些潜伏储库。我们以前已经确定了两种激活潜伏HIV前病毒表达的机制：免疫激活，这需要潜伏感染的细胞和未感染的活化CD 4+细胞之间的接触，和低甲基化。我们最近证明，急性HIV感染增加了细胞甲基化基因的能力。HIV感染细胞中DNA甲基转移酶（DNMT）活性的增加可通过HIV LTR的甲基化和随后的病毒基因表达沉默导致潜伏库的增加。此外，增加的DNMT活性可导致细胞基因的异常甲基化，包括干扰素-γ（IFN-γ）。IFN-γ的这种高甲基化导致IFN-γ的产生减少，1型免疫应答降低和无细胞病毒体的扩散增加。在试图了解甲基化在病毒和基因调控中的作用时，我们在几个领域取得了进展。首先，HIV前病毒整合对于HIV感染增加细胞甲基化基因的能力不是必需的。第二，在HIV感染淋巴细胞期间，DNMT-1而不是DNMT 3a和DNMT 3b增加。第三，在一些（49/96）接受长期HAART的患者中，CD 4 + T细胞产生IFN-γ的能力大大降低，这可以通过IFN-γ启动子的低甲基化来恢复， 使用低甲基化药物。该启动子的亚硫酸氢盐基因组测序表明，IFN-γ启动子在HAART治疗期间可以保持高甲基化。第四，使用新开发的基于阵列的测定（差异甲基化杂交），我们发现来自HIV感染个体的CD 4 + T淋巴细胞具有比正常对照更广泛甲基化的几个CpG岛。我们已经确定了这些基因之一为p16（Ink 4a）基因 表明CpG岛通常是低甲基化的，在HIV感染后可以是高甲基化的。由于DNMT 1是在HIV感染后主要上调的DNMT，并且最近的证据表明，除了其酶促功能之外，它还可以与其他免疫调节因子形成复合物。 包括转录因子如达特或Vpr的蛋白质可以以新的方式拴系到含有DNMT-1的复合物上，使转录控制失调。虽然在很大程度上是未知的，由HTLV-1感染引发的淋巴增殖过程中的细胞事件是非常重要的白血病的建立。我们最近的特点是HTLV-1的感染性克隆以及易感表达细胞系，以研究这种感染性克隆的分子相互作用。被膜的HTLV-1表面单元已经融合到IG分子上，并被用于研究HIV-1受体的调节。 该项目以前为Z 01 CM 09251-12 LLB。 艾滋病标题：艾滋病毒复制的负调节-致病作用

The goal of this project is to better understand the effects of host cells and other co-factors on human retroviral replication and pathogenesis. Highly active combination antiviral drug therapy (HAART) holds promise to convert productive HIV infections into a chronic, non-viremic, non-progressive condition. However, in some patients, limited but continuous viral production has been observed and, in others, latently infected long-lived peripheral T cells which can be reactivated during a normal immune response have been identified in HAART-treated HIV infected individuals. In developing long term therapy for HIV infected individuals it will be necessary to control viral production from and/or eradicate these latent reservoirs through adjuvant therapies like immunotherapy. We have previously identified two mechanisms which activate expression of latent HIV provirus: immune activation, which requires contact between latently infected cells and uninfected activated CD4+ cells, and hypomethylation. We have recently demonstrated that acute HIV infection increases the cellular capacity to methylate genes. Increased DNA methyltransferase (DNMT) activity in HIV infected cells can result in an increase in the latent reservoir through methylation of the HIV LTR and the subsequent silencing of viral gene expression. In addition, increased DNMT activity can result in aberrant methylation of cellular genes including, interferon-gamma (IFN-gamma). This hypermethylation of IFN-gamma results in decreased production of IFN-gamma, decreased type 1 immune response and increased spread of cell free virions. In attempting to understand the role of methylation in the regulation of viral and gene regulation, we have made progress in several areas. First, HIV proviral integration is not necessary for HIV infection to increase the cellular capacity to methylate genes. Second, DNMT-1 but not DNMT3a and DNMT3b are increased during HIV infection of lymphoid cells. Third, in some (49/96) patients on long-term HAART, CD4+ T cells have greatly diminished ability to produce IFN-gamma which can be restored through hypomethylation of IFN-gamma promoter, using hypomethylating drugs. Bisulfite genomic sequencing of this promoter has shown that the IFN-gamma promoter can remain hypermethylated during HAART therapy. Fourth, using a newly developed array based assay (differential methylation hybridization), we have found that CD4+ T lymphocytes from HIV infected individuals have several CpG islands which are more extensively methylated than normal controls. We have identified one of these genes as the p16(Ink4a) gene indicating that CpG islands, normally hypomethylated, can be hypermethlated after HIV infection. Since DNMT1 is the DNMT that is predominantly upregulated following HIV infection and recent evidence suggests that, in addition to its enzymatic function, it can form complexes with other proteins including transcriptional factors such as Tat or Vpr can tether to complexes containing DNMT-1 dysregulate transcription control in a novel manner. Although largely unknown, the cellular events triggered by HTLV-1 infection in the lymphoproliferative process are of major importance to the establishment of leukemia. We have recently characterized an infectious clone of HTLV-1 as well as a susceptible expressing cell line to study the molecular interactions of this infectious clone. The HTLV-1 surface unit of the envelope has been fused to the Ig molecule and is being used to study the regulation of the HIV-1 receptor. This project was formerly Z01 CM 09251-12 LLB. AIDS Title: Negative Regulation of HIV replication - Pathogenic Effects