Diefenbach Russell J, Davis April, Miranda-Saksena Monica, Fernandez Marian A, Kelly Barbara J, Jones Cheryl A, LaVail Jennifer H, Xue Jing, Lai Joey, Cunningham Anthony L
Centre for Virus Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW, Australia
Centre for Virus Research, The Westmead Institute for Medical Research, The University of Sydney, Westmead, NSW, Australia.
J Virol. 2015 Dec 9;90(4):2102-11. doi: 10.1128/JVI.03041-15. Print 2016 Feb 15.
The alphaherpesviral envelope protein pUS9 has been shown to play a role in the anterograde axonal transport of herpes simplex virus 1 (HSV-1), yet the molecular mechanism is unknown. To address this, we used an in vitro pulldown assay to define a series of five arginine residues within the conserved pUS9 basic domain that were essential for binding the molecular motor kinesin-1. The mutation of these pUS9 arginine residues to asparagine blocked the binding of both recombinant and native kinesin-1. We next generated HSV-1 with the same pUS9 arginine residues mutated to asparagine (HSV-1pUS9KBDM) and then restored them being to arginine (HSV-1pUS9KBDR). The two mutated viruses were analyzed initially in a zosteriform model of recurrent cutaneous infection. The primary skin lesion scores were identical in severity and kinetics, and there were no differences in viral load at dorsal root ganglionic (DRG) neurons at day 4 postinfection (p.i.) for both viruses. In contrast, HSV-1pUS9KBDM showed a partial reduction in secondary skin lesions at day 8 p.i. compared to the level for HSV-1pUS9KBDR. The use of rat DRG neuronal cultures in a microfluidic chamber system showed both a reduction in anterograde axonal transport and spread from axons to nonneuronal cells for HSV-1pUS9KBDM. Therefore, the basic domain of pUS9 contributes to anterograde axonal transport and spread of HSV-1 from neurons to the skin through recruitment of kinesin-1.
Herpes simplex virus 1 and 2 cause genital herpes, blindness, encephalitis, and occasionally neonatal deaths. There is also increasing evidence that sexually transmitted genital herpes increases HIV acquisition, and the reactivation of HSV increases HIV replication and transmission. New antiviral strategies are required to control resistant viruses and to block HSV spread, thereby reducing HIV acquisition and transmission. These aims will be facilitated through understanding how HSV is transported down nerves and into skin. In this study, we have defined how a key viral protein plays a role in both axonal transport and spread of the virus from nerve cells to the skin.
α疱疹病毒包膜蛋白pUS9已被证明在单纯疱疹病毒1（HSV-1）的顺行轴突运输中起作用，但其分子机制尚不清楚。为了解决这个问题，我们使用体外下拉试验来确定保守的pUS9碱性结构域内的一系列五个精氨酸残基，这些残基对于结合分子马达驱动蛋白-1至关重要。将这些pUS9精氨酸残基突变为天冬酰胺会阻断重组驱动蛋白-1和天然驱动蛋白-1的结合。接下来，我们构建了将相同的pUS9精氨酸残基突变为天冬酰胺的HSV-1（HSV-1pUS9KBDM），然后将它们恢复为精氨酸（HSV-1pUS9KBDR）。最初在复发性皮肤感染的带状疱疹模型中分析这两种突变病毒。原发性皮肤病变的严重程度和动力学相同，两种病毒在感染后第4天背根神经节（DRG）神经元中的病毒载量没有差异。相比之下，与HSV-1pUS9KBDR相比，HSV-1pUS9KBDM在感染后第8天的继发性皮肤病变出现部分减少。在微流控腔室系统中使用大鼠DRG神经元培养物显示，HSV-1pUS9KBDM的顺行轴突运输减少，并且从轴突扩散到非神经元细胞的情况也减少。因此，pUS9的碱性结构域通过募集驱动蛋白-1促进HSV-1的顺行轴突运输以及从神经元向皮肤的扩散。
单纯疱疹病毒1和2可引起生殖器疱疹、失明、脑炎，偶尔还会导致新生儿死亡。也有越来越多的证据表明，性传播的生殖器疱疹会增加感染HIV的风险，而HSV的重新激活会增加HIV的复制和传播。需要新的抗病毒策略来控制耐药病毒并阻断HSV的传播，从而减少HIV的感染和传播。通过了解HSV如何沿着神经向下运输并进入皮肤，将有助于实现这些目标。在这项研究中，我们已经确定了一种关键病毒蛋白如何在病毒的轴突运输以及从神经细胞向皮肤的扩散中发挥作用。
