MOLECULAR ANALYSIS & MODELING OF HIV-1 TRANSMISSION, CONTAINMENT AND ESCAPE

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 这项研究解决了全球健康倡议的重大挑战#6：了解哪些免疫反应提供保护性免疫（针对HIV-1）。全长传播的HIV-1基因组的鉴定可能有助于HIV-1发病机制，杀微生物剂和疫苗研究，使直接分析这些病毒实际上负责生产性临床感染。我们在12例急性感染受试者（9例进化枝B和3例进化枝C）中发现，通过单基因组扩增和血浆病毒体RNA测序，可以推断出传播/创始病毒的完整HIV-1基因组。这使得分子克隆和生物学分析的传播/创始人病毒和全面的全基因组评估的遗传印记留在不断演变的病毒准种的复合宿主选择压力。 传播的病毒编码完整的典型基因（gag-pol-vif-vpr-tat-rev-vpu-env-nef），并在原代人CD 4（+）T淋巴细胞中有效复制，但在单核细胞衍生的巨噬细胞中复制效果较差。传播的病毒是CD 4和CCR 5嗜性的，并表现出包膜桥接片和可变环3的辅助受体结合表面的隐藏。在感染后2个月，3名受试者中的传播/创始病毒几乎完全被在2 - 5个高度选择的基因组位点上不同的病毒所取代;到12-20个月，病毒在17-34个离散位置显示出集中的突变。 这些发现揭示了与粘膜HIV-1传播相关的病毒特性，以及主要但不完全由早期细胞毒性T细胞应答驱动的一组有限的快速进化的适应性突变。 我们最近描述了两种自体单克隆抗体，6.4C和13.6A，从赞比亚受试者产生，每个识别的创始人Env上的V1 V2依赖性目标。使用这些单克隆抗体，我们确定了V1 V2结构域中的两个突变，这两个突变在单抗体水平上参与逃逸。我们已经扩展了这些研究，以测试糖基化的变化是否是Nab抗性所必需的，并进一步确定单克隆抗体的表位。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. This study addresses Grand Challenge #6 of the Global Health Initiative: Learn which immunological responses provide protective immunity (against HIV-1). Identification of full-length transmitted HIV-1 genomes could be instrumental in HIV-1 pathogenesis, microbicide, and vaccine research by enabling direct analysis of those viruses actually responsible for productive clinical infection. We have shown in 12 acutely infected subjects (9 clade B and 3 clade C) that complete HIV-1 genomes of transmitted/founder viruses can be inferred by single genome amplification and sequencing of plasma virion RNA. This allowed for molecular cloning and biological analysis of transmitted/founder viruses and comprehensive genome-wide assessment of the genetic imprint left on the evolving virus quasispecies by a composite of host selection pressures. Transmitted viruses encoded intact canonical genes (gag-pol-vif-vpr-tat-rev-vpu-env-nef) and replicated efficiently in primary human CD4(+) T lymphocytes but much less so in monocyte-derived macrophages. Transmitted viruses were CD4 and CCR5 tropic and demonstrated concealment of coreceptor binding surfaces of the envelope bridging sheet and variable loop 3. At 2 mo after infection, transmitted/founder viruses in three subjects were nearly completely replaced by viruses differing at two to five highly selected genomic loci; by 12-20 mo, viruses exhibited concentrated mutations at 17-34 discrete locations. These findings reveal viral properties associated with mucosal HIV-1 transmission and a limited set of rapidly evolving adaptive mutations driven primarily, but not exclusively, by early cytotoxic T cell responses. We have recently described two autologous monoclonal antibodies, 6.4C and 13.6A, generated from a Zambian subject, that each recognize a V1V2-dependent target on the founder Env. Using these Mabs, we identified two mutations in the V1V2 domain that were involved in escape at the single antibody level. We have extended these studies to test whether a change in glycosylation is necessary for Nab resistance and to further define epitopes of the Mabs.