Functional and biochemical relationships between tropism, infectivity, and neut

趋化因子受体作为HIV进入的辅助受体的鉴定，不仅有助于对病毒趋向性的理解，而且为HIV疾病的治疗干预提供了额外的靶点。一些趋化因子受体已被证明是HIV-1进入的辅助受体。主要有CXCR4（用于t细胞系病毒）和CCR5（用于巨噬细胞病毒）。由于艾滋病毒在施加选择性压力时具有适应能力，因此很可能任何旨在阻止艾滋病毒与一种辅助受体相互作用的药物都会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STRL33在所有淋巴组织中表达，我们与NIAID的J. Farber博士合作，对其与HIV的共受体活性进行了测试。STRL33在Jurkat细胞中的表达增加了对HIV-1的ELI1分离物感染的容忍度。因此，STRL33在体外可以作为HIV-1的辅助受体。除了测试STRL33与许多不同表型的HIV-1毒株的共受体活性外，我们还开始了对HIV-2和SIV的研究。我们已经在感染性实验中证明，HIV-1的MAL株和SIV的mac239分离株使用STRL33，但不像它们使用CCR5那样好。只有在培养30天后才出现病毒，这表明了适应。为了证实这一点，用35天后出现的病毒感染新鲜Jurkat- STRL33细胞和亲本Jurkat细胞。在第二代中，大约12天后观察到病毒产生，从而表明SIVmac239和HIV-1 MAL都适应了更有效地使用STRL33。重要的是，这些传代的病毒仍然无法感染Jurkat细胞。一种针对STRL33的抗体抑制了病毒感染的事实证明，传代病毒已经适应了使用STRL33。我们从适应的MAL病毒中克隆了12个包膜基因，从适应的SIVmac239中克隆了6个env基因。我们目前正在将这些基因返回到传染性分子克隆中，以评估它们的生物活性。基于许多病毒（包括SIV和HIV-1）的细胞趋向性，我们预测在SUP-T1细胞上存在另一种辅助受体。我们克隆了这个辅助受体，并将其鉴定为TECK的趋化因子受体CCR9 （CCL25）。CCR9以A和B两种形式存在，它们在N端相差12个氨基酸。CCR9B对包括SIVagmSAB和SIVagmTAN在内的几种SIV毒株具有活性，但迄今为止尚未发现使用该分子作为进入辅因子的HIV-1。正在筛选更多的HIV-1分离株，包括初级分离株。由于CCR9A/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疾病和艾滋病发病机制的明显重要性，我们正在扩大我们的研究范围，包括在这些细胞中表达的共受体，以及这些细胞和树突状细胞的感染，这些细胞分泌ccr25，因此可能招募T细胞。

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 STRL33 is expressed in all lymphoid tissues, in collaboration with Dr J. Farber, NIAID, we tested it for coreceptor activity with HIV. STRL33 expression in Jurkat cells conferred increased permissivity to infection by the ELI1 isolate of HIV-1. Thus, STRL33 can act as an HIV-1 coreceptor in vitro. As well as testing the coreceptor activity of STRL33 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 STRL33 but not as well as they use CCR5. The appearance of virus only after about 30 days in culture is indicative of adaptation. To confirm this, virus emerging after about 35 days was used to infect fresh Jurkat- STRL33 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 STRL33 more efficiently. Importantly, these passaged viruses were still unable to infect Jurkat cells. That the passaged virus had adapted to use STRL33 was demonstrated by the fact that an antibody raised to STRL33 inhibited virus infection. We have cloned 12 envelope genes from the adapted MAL virus and 6 env genes from the adapted SIVmac239. We are currently returning these genes to the infectious molecular clones to assess their biological activities. 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.