Alfano M, Schmidtmayerova H, Amella C A, Pushkarsky T, Bukrinsky M
Picower Institute for Medical Research, Manhasset, New York 11030, USA.
J Exp Med. 1999 Sep 6;190(5):597-605. doi: 10.1084/jem.190.5.597.
Infection of target cells by HIV-1 requires initial binding interactions between the viral envelope glycoprotein gp120, the cell surface protein CD4, and one of the members of the seven-transmembrane G protein-coupled chemokine receptor family. Most primary isolates (R5 strains) use chemokine receptor CCR5, but some primary syncytium-inducing, as well as T cell line-adapted, strains (X4 strains) use the CXCR4 receptor. Signaling from both CCR5 and CXCR4 is mediated by pertussis toxin (PTX)-sensitive G(i) proteins and is not required for HIV-1 entry. Here, we show that the PTX holotoxin as well as its binding subunit, B-oligomer, which lacks G(i)-inhibitory activity, blocked entry of R5 but not X4 strains into primary T lymphocytes. Interestingly, B-oligomer inhibited virus production by peripheral blood mononuclear cell cultures infected with either R5 or X4 strains, indicating that it can affect HIV-1 replication at both entry and post-entry levels. T cells treated with B-oligomer did not initiate signal transduction in response to macrophage inflammatory protein (MIP)-1beta or RANTES (regulated upon activation, normal T cell expressed and secreted); however, cell surface expression of CCR5 and binding of MIP-1beta or HIV-1 to such cells were not impaired. The inhibitory effect of B-oligomer on signaling from CCR5 and on entry of R5 HIV-1 strains was reversed by protein kinase C (PKC) inhibitors, indicating that B-oligomer activity is mediated by signaling events that involve PKC. B-oligomer also blocked cocapping of CCR5 and CD4 induced by R5 HIV-1 in primary T cells, but did not affect cocapping of CXCR4 and CD4 after inoculation of the cultures with X4 HIV-1. These results suggest that the B-oligomer of PTX cross-deactivates CCR5 to impair its function as a coreceptor for HIV-1.
HIV-1对靶细胞的感染需要病毒包膜糖蛋白gp120、细胞表面蛋白CD4以及七跨膜G蛋白偶联趋化因子受体家族成员之一之间的初始结合相互作用。大多数原代分离株（R5毒株）使用趋化因子受体CCR5，但一些原代合胞体诱导株以及T细胞系适应株（X4毒株）使用CXCR4受体。CCR5和CXCR4的信号传导均由百日咳毒素（PTX）敏感的G(i)蛋白介导，且HIV-1进入细胞并不需要该信号传导。在此，我们表明PTX全毒素及其缺乏G(i)抑制活性的结合亚基B-寡聚体可阻断R5毒株而非X4毒株进入原代T淋巴细胞。有趣的是，B-寡聚体抑制外周血单核细胞培养物中感染R5或X4毒株后的病毒产生，表明其可在进入和进入后水平影响HIV-1复制。用B-寡聚体处理的T细胞对巨噬细胞炎性蛋白（MIP）-1β或RANTES（激活后调节，正常T细胞表达和分泌）无反应而启动信号转导；然而，CCR5的细胞表面表达以及MIP-1β或HIV-1与此类细胞的结合并未受损。蛋白激酶C（PKC）抑制剂可逆转B-寡聚体对CCR5信号传导和R5 HIV-1毒株进入的抑制作用，表明B-寡聚体活性由涉及PKC的信号事件介导。B-寡聚体还可阻断原代T细胞中由R5 HIV-1诱导的CCR5和CD4的共帽化，但在用X4 HIV-1接种培养物后不影响CXCR4和CD4的共帽化。这些结果表明PTX的B-寡聚体可交叉失活CCR5以损害其作为HIV-1共受体的功能。
