HIV/SIV viral reservoirs in lymphocytes and macrophages

拟议的研究涉及HIV-1在接受高效抗逆转录病毒治疗（HAART）的患者中持续存在的机制。在依从的患者中，HAART可以将血浆中的游离病毒水平降低至检测限以下。然而，在最初资助期间完成的研究表明，HIV-1在携带HIV-1基因组完整拷贝的一小部分潜伏感染的静息记忆CD 4 + T细胞中终生存在。由于潜伏感染的细胞是HIV-1根除的主要障碍，因此了解维持潜伏的分子机制非常重要。基于体外研究，已经提出了几种潜在的分子机制，但仍不清楚这些机制中的任何一种是否准确地解释了HIV-1在体内静息CD 4 + T细胞中的持久性。在拟议的研究中，我们将使用遗传学方法来研究体内潜伏期的机制，重点是 宿主基因组中整合位点的影响。前两个特定目的涉及克隆和表征HIV-1在体内静息CD 4 + T细胞中整合的位点。一种新的方法将被用来确定的小子集的集成前病毒的功能显着。这些整合位点的分析应该提供一个潜在的前病毒所在的染色体环境的一般图片。在第三个具体目标中，我们将使用整合位点信息来研究宿主染色体上作为HIV-1整合位点的区域的转录活性和染色质结构。在最后的目标，我们将测试一个新的假设，HIV-1潜伏期的机制，这是与大多数已发表的工作是一致的，并代表了几个建议的机制的合成。总之，这些研究应该有助于阐明HIV-1潜伏期如何在体内运作。

The proposed studies deal with the mechanism by which HIV-1 persists in patients on highly active antiretroviral therapy (HAART). In compliant patients, HAART can decrease levels of free virus in the plasma to below the limit of detection. However, studies completed during the initial funding period demonstrated that HIV-1 persists for life in a small pool of latently infected resting memory CD4 + T cells that carry an integrated copy of the HIV-1 genome. Because latently infected cells represent a major barrier to HIV-1 eradication, it is important to understand the molecular mechanisms that maintain latency. Several potential molecular mechanisms have been proposed based on in vitro studies, but it remains unclear whether any of these accurately account for the persistence of HIV-1 in resting CD4 + T cells in vivo. In the proposed studies, we will use genetic approaches to examine the mechanism of latency in vivo, focusing on the influence of the integration site in the host genome. The first two Specific Aims involve cloning and characterizing sites of HIV-1 integration in resting CD4 + T cells in vivo. A novel method will be used to identify the small subset of integrated proviruses that are functionally significant. Analysis of these integration sites should provide a general picture the chromosomal environments where latent proviruses reside. In the third Specific Aim, we will use integration site information to study the transcriptional activity and chromatin structure of regions of the host chromosome that are sites for HIV-1 integration. In the final Aim, we will test a novel hypothesis concerning the mechanism of HIV-1 latency that is consistent with most published work and that represents a synthesis of several proposed mechanisms. Together, these studies should help to elucidate how HIV-1 latency operates in vivo.