Herpesvirus Interactions With Cell Surface Receptors

单纯疱疹病毒(HSV)进入细胞需要病毒糖蛋白gD与其受体之一结合，从而导致病毒与细胞融合。已知HSV的穿透是通过用pH为3的缓冲液处理与病毒结合的细胞来阻止的。为了进一步了解HSV进入的分子机制，我们用β-半乳糖苷酶报告试验检测了酸灭活的效果。HSV-1无细胞病毒粒子(Kos)经弱酸性pH处理后，不能进入Vero细胞。因此，低pH值直接影响病毒粒子本身。具体地说，当病毒粒子用pH&lt；6.5处理时，进入活性被抑制。失活迅速、不可逆，且与温度有关。经酸处理的病毒粒子也不能进入表达gD结合受体疱疹病毒进入介体A(HveA)、Nectin-1或Nectin-2的CHO细胞。我们证明，这些受体在4℃下介导稳定的、抗肝素的病毒与细胞的结合，这种结合可被可溶性gD或可溶性受体阻断。这些发现表明，受体对于进入细胞是必不可少的，部分原因是它们在病毒-细胞融合之前加强了HSV与细胞的黏附。低pH处理不影响HSV与HveA、Nectin-1或Nectin-2的稳定结合，也不改变gD的受体结合构象。综上所述，这些数据表明，低pH值扰乱了HSV进入过程中的一个步骤，而不是gD受体的相互作用。我们认为低pH预处理使HSV失去与细胞膜融合的能力，是解偶联受体结合和融合进入过程的有用工具。

Entry of herpes simplex virus (HSV) into cells requires binding of viral glycoprotein gD to one of its receptors which then leads to virus-cell fusion. It is known that HSV penetration is blocked by treating cells bound with virus with a buffer at pH 3. To gain further insights into the molecular mechanism of HSV entry, the effects of acid inactivation were examined using a beta-galactosidase reporter assay. Cell-free virions of HSV-1 (KOS) that were pretreated with mildly acidic pH were incompetent for entry into Vero cells. Thus, low pH directly affects the virions themselves. Specifically, entry activity was inhibited when virions was treated with pH < 6.5. Inactivation was rapid, irreversible and temperature-dependent. Acid-treated virions also failed to enter CHO cells expressing the gD-binding receptors herpesvirus entry mediator A (HveA), nectin-1 or nectin-2. We demonstrated that these receptors mediate stable, heparin-resistant binding of virus to cells at 4?C, and that binding was blocked by addition of soluble gD or soluble receptors. These findings suggest that receptors are essential for entry partly due to their role in strengthening HSV adhesion to cells prior to virus-cell fusion. Low pH treatment did not affect stable binding of HSV to HveA, nectin-1 or nectin-2, nor did it alter the receptor-binding conformation of gD. Taken together, the data indicate low pH perturbs a step in the HSV entry process other than the gD-receptor interaction. We propose that low pH pretreatment incapacitates HSV for fusion with cell membranes and is a useful tool to uncouple the entry processes of receptor binding and fusion.