Hart T K, Kirsh R, Ellens H, Sweet R W, Lambert D M, Petteway S R, Leary J, Bugelski P J
Department of Experimental Pathology, SmithKline Beecham Pharmaceuticals, King of Prussia, PA 19406.
Proc Natl Acad Sci U S A. 1991 Mar 15;88(6):2189-93. doi: 10.1073/pnas.88.6.2189.
Human immunodeficiency virus (HIV) infects cells after binding of the viral envelope glycoprotein gp120 to the cell surface recognition marker CD4. gp120 is noncovalently associated with the HIV transmembrane envelope glycoprotein gp41, and this complex is believed responsible for the initial stages of HIV infection and cytopathic events in infected cells. Soluble constructs of CD4 that contain the gp120 binding site inhibit HIV infection in vitro. This is believed to occur by competitive inhibition of viral binding to cellular CD4. Here we suggest an alternative mechanism of viral inhibition by soluble CD4 proteins. We demonstrate biochemically and morphologically that following binding, the soluble CD4 proteins sT4, V1V2,DT, and V1[106] (amino acids 1-369, 1-183, and -2 to 106 of mature CD4) induced the release of gp120 from HIV-1 and HIV-1-infected cells. gp120 release was concentration-, time-, and temperature-dependent. The reaction was biphasic at 37 degrees C and did not take place at 4 degrees C, indicating that binding of soluble CD4 was not sufficient to release gp120. The appearance of free gp120 in the medium after incubation with sT4 correlated with a decrease in envelope glycoprotein spikes on virions and exposure of a previously cryptic epitope near the amino terminus of gp41 on virions and infected cells. The concentration of soluble CD4 proteins needed to induce the release of gp120 from virally infected cells also correlated with those required to inhibit HIV-mediated syncytium formation. These results suggest that soluble CD4 constructs may inactivate HIV by inducing the release of gp120. We propose that HIV envelope-mediated fusion is initiated following rearrangement and/or dissociation of gp120 from the gp120-gp41 complex upon binding to cellular CD4, thus exposing the fusion domain of gp41.
人类免疫缺陷病毒(HIV)在病毒包膜糖蛋白gp120与细胞表面识别标志物CD4结合后感染细胞。gp120与HIV跨膜包膜糖蛋白gp41非共价结合,并且这种复合物被认为是HIV感染初始阶段以及感染细胞中细胞病变事件的原因。含有gp120结合位点的可溶性CD4构建体在体外可抑制HIV感染。据信这是通过竞争性抑制病毒与细胞CD4的结合而发生的。在此,我们提出了可溶性CD4蛋白抑制病毒的另一种机制。我们通过生化和形态学方法证明,在结合后,可溶性CD4蛋白sT4、V1V2、DT和V1[106](成熟CD4的氨基酸1 - 369、1 - 183以及 - 2至106)可诱导gp120从HIV - 1及HIV - 1感染细胞中释放。gp120的释放具有浓度、时间和温度依赖性。该反应在37℃时呈双相性,在4℃时不发生,这表明可溶性CD4的结合不足以释放gp120。与sT4孵育后培养基中游离gp120的出现与病毒粒子上包膜糖蛋白刺突的减少以及病毒粒子和感染细胞上gp41氨基末端附近一个先前隐蔽的表位的暴露相关。从病毒感染细胞中诱导gp120释放所需的可溶性CD4蛋白浓度也与抑制HIV介导的合胞体形成所需的浓度相关。这些结果表明,可溶性CD4构建体可能通过诱导gp120的释放使HIV失活。我们提出,HIV包膜介导的融合是在gp120与细胞CD4结合后从gp120 - gp41复合物重排和/或解离后启动的,从而暴露gp41的融合结构域。