Relationships between tropism and infectivity in HIV

HIV的趋向性和传染性之间的关系

• 批准号: 6545137
• 负责人: KEITH PEDEN
• 金额: --
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6545137
• 关键词: cytokine receptors; evolution; gene expression; gene mutation; genetic strain; host organism interaction; human immunodeficiency virus; human immunodeficiency virus 2; immunologic assay /test; intermolecular interaction; molecular cloning; natural selections; protein isoforms; protein structure function; simian immunodeficiency virus; tissue /cell culture; virus genetics; virus infection mechanism; virus protein; virus receptors

Summary: The identification of chemokine receptors as coreceptors for HIV entry, not only has contributed to the understanding on viral tropism but has provided an additional target for therapeutic intervention for HIV disease. Several chemokine receptors have been shown to function as coreceptors for HIV-1 entry. The main ones are CXCR4 (for T-cell line tropic viruses) and CCR5 (for macrophage-tropic viruses). Because of the capacity of HIV to adapt when selective pressures are imposed, it is likely that any drug designed to block the interaction of HIV with one coreceptor will force the virus to use additional coreceptors. Thus, the determination of the complete coreceptor repertoire is necessary. Because CXCR6 (STRL33) is expressed in all lymphoid tissues, in collaboration with Dr J. Farber, NIAID, we tested it for coreceptor activity with HIV. CXCR6 expression in Jurkat cells conferred increased permissivity to infection by the ELI1 isolate of HIV-1. Thus, CXCR6 can act as an HIV-1 coreceptor in vitro. As well as testing the coreceptor activity of CXCR6 with a number of HIV-1 strains of different phenotypes, we have begun studies with HIV-2 and SIV. We have shown, in an infectivity assay, that the MAL strain of HIV-1 and the mac239 isolate of SIV use CXCR6 but not as well as they use CCR5. The appearance of virus only after about 30 days in culture was indicative of adaptation. To confirm this, virus emerging after about 35 days was used to infect fresh Jurkat-CXCR6 cells as well as the parent Jurkat cells. In this second passage, virus production was seen after about 12 days, thus demonstrating that both SIVmac239 and HIV-1 MAL had adapted to use CXCR6 more efficiently. Importantly, these passaged viruses were still unable to infect Jurkat cells. That the passaged virus had adapted to use CXCR6 was demonstrated by the fact that an antibody raised to CXCR6 inhibited virus infection. We have cloned 12 envelope genes from the adapted MAL virus and 6 env genes from the adapted SIVmac239. We have inserted 8 of the MAL envs into the MAL infectious clone; 7 were infectious. Four of the 6 envs from CXCR6-adapted SIVmac239 were infectious in the SIVmac239 clone. All of the adapted clones had enhanced capacity to use CXCR6 over CCR5 in both a single-cycle assay and a productive infection assay. Sequncing of the MAL envs demonstrated that, while changes were found in several regions of gp120 and gp41, changes in the V3 region of gp120 were sufficient to confer the adapted phenotype. Based on the cell tropism of a number of viruses, both SIV and HIV-1, we predicted the existence of another coreceptor on SUP-T1 cells. We have cloned this coreceptor and identified it as CCR9, a chemokine receptor for TECK (CCL25). CCR9 exists in two forms, A and B, that differ by 12 amino acids at the N terminus. CCR9B has activity with several strains of SIV, including SIVagmSAB and SIVagmTAN, but as yet no HIV-1 has been found that uses this molecule as an entry cofactor. Additional HIV-1 isolates including primary isolates are being screened. Since CCR9A/B is expressed in thymocytes and in the lymphocyte subset that trafficks to the gut, and because of the obvious importance of these tissues to HIV disease and AIDS pathogenesis, we are expanding our studies to include the coreceptors expressed in these cells as well as to the infection of these cells and of dendritic cells, which secrete CCL25 and thus may recruit T cells. We have demonstrated that infection of monocyte-derived macrophages (MDM) and monocyte-derived dendritic cells (MDDC) by R5 strains of HIV-1 results in the induction of IP-10 and I-TAC, chemokines for CXCR3, but not of CCR5, CXCR4, or CCR5 ligands. This is of interest for HIV disease, since all CCR5-expressing memory CD4 cells also express CXCR3 and thus infected tissue macrophages or DC would secrete IP-10 and I-TAC, which would then recruit memory T cells, which could be infected and thus disseminate infection.

总结： 趋化因子受体作为HIV进入的辅助受体的鉴定，不仅有助于理解病毒的嗜性，而且为HIV疾病的治疗干预提供了另一个靶点。 几种趋化因子受体已被证明作为HIV-1进入的辅助受体。 主要的是CXCR 4（针对T细胞系嗜性病毒）和CCR 5（针对巨噬细胞嗜性病毒）。 由于艾滋病毒在选择性压力下的适应能力，任何旨在阻断艾滋病毒与一种辅助受体相互作用的药物都可能迫使病毒使用其他辅助受体。 因此，确定完整的辅助受体库是必要的。 由于CXCR 6（STRL 33）在所有淋巴组织中表达，因此我们与NIAID的J. Farber博士合作，测试了它与HIV的共受体活性。 Jurkat细胞中CXCR 6的表达增加了HIV-1的ELI 1分离株的感染概率。 因此，CXCR 6在体外可作为HIV-1辅助受体。 除了用不同表型的HIV-1毒株测试CXCR 6的辅助受体活性外，我们还开始了HIV-2和SIV的研究。 我们已经在感染性试验中表明，HIV-1的MAL株和SIV的mac 239分离株使用CXCR 6，但不如它们使用CCR 5。 仅在培养约30天后出现病毒，表明适应。 为了证实这一点，使用约35天后出现的病毒感染新鲜Jurkat-CXCR 6细胞以及亲本Jurkat细胞。 在第二次传代中，在约12天后观察到病毒产生，从而证明SIVmac 239和HIV-1 MAL均已适应更有效地使用CXCR 6。 重要的是，这些传代病毒仍然不能感染Jurkat细胞。 传代的病毒已经适应使用CXCR 6，这一点通过针对CXCR 6产生的抗体抑制病毒感染的事实得到证明。 我们已经从适应的MAL病毒中克隆了12个包膜基因，从适应的SIVmac 239中克隆了6个env基因。 我们已经将8个MAL env插入MAL感染性克隆中; 7个具有感染性。 来自CXCR 6适应的SIVmac 239的6个env中的4个在SIVmac 239克隆中具有感染性。 所有适应的克隆在单循环测定和生产性感染测定中使用CXCR 6的能力都比CCR 5增强。 MAL env的测序表明，虽然在gp 120和gp 41的几个区域中发现了变化，但gp 120的V3区域中的变化足以赋予适应的表型。 基于SIV和HIV-1等多种病毒的细胞嗜性，我们预测在SUP-T1细胞上存在另一种辅助受体。 我们已经克隆了这种辅助受体，并将其鉴定为CCR 9，一种TECK（CCL 25）的趋化因子受体。 CCR 9以两种形式存在，A和B，在N末端相差12个氨基酸。 CCR 9 B对包括SIVagmSAB和SIVagmTAN在内的几种SIV毒株具有活性，但迄今为止还没有发现使用该分子作为进入辅因子的HIV-1。 正在筛选其他HIV-1分离株，包括主要分离株。 由于CCR 9 A/B在胸腺细胞和运输到肠道的淋巴细胞亚群中表达，并且由于这些组织对HIV疾病和AIDS发病机制的明显重要性，我们正在扩大我们的研究，包括在这些细胞中表达的辅助受体以及这些细胞和树突状细胞的感染，树突状细胞分泌CCL 25，因此可以募集T细胞。 我们已经证明，HIV-1的R5株感染单核细胞衍生的巨噬细胞（MDM）和单核细胞衍生的树突状细胞（MDDC）导致诱导IP-10和I-TAC，CXCR 3的趋化因子，但不诱导CCR 5、CXCR 4或CCR 5配体。 这对于HIV疾病是有意义的，因为所有表达CCR 5的记忆性CD 4细胞也表达CXCR 3，因此感染的组织巨噬细胞或DC将分泌IP-10和I-TAC，其然后将募集记忆性T细胞，其可以被感染并因此传播感染。