Sánchez-Tacuba Liliana, Rojas Margarito, Arias Carlos F, López Susana
Departamento de Génetica del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos, Mexico.
Departamento de Génetica del Desarrollo y Fisiología Molecular, Instituto de Biotecnología, Universidad Nacional Autónoma de México, Cuernavaca, Morelos, Mexico
J Virol. 2015 Dec;89(23):12145-53. doi: 10.1128/JVI.01874-15. Epub 2015 Sep 23.
The innate immune response is the first line of defense of the host cell against a viral infection. In turn, viruses have evolved a wide variety of strategies to hide from, and to directly antagonize, the host innate immune pathways. One of these pathways is the 2'-5'-oligoadenylate synthetase (OAS)/RNase L pathway. OAS is activated by double-stranded RNA (dsRNA) to produce 2'-5' oligoadenylates, which are the activators of RNase L; this enzyme degrades viral and cellular RNAs, restricting viral infection. It has been recently found that the carboxy-terminal domain (CTD) of rotavirus VP3 has a 2'-5'-phosphodiesterase (PDE) activity that is able to functionally substitute for the PDE activity of the mouse hepatitis virus ns2 protein. This particular phosphodiesterase cleaves the 2'-5'-phosphodiester bond of the oligoadenylates, antagonizing the OAS/RNase L pathway. However, whether this activity of VP3 is relevant during the replication cycle of rotavirus is not known. Here, we demonstrate that after rotavirus infection the OAS/RNase L complex becomes activated; however, the virus is able to control its activity using at least two distinct mechanisms. A virus-cell interaction that occurs during or before rotavirus endocytosis triggers a signal that prevents the early activation of RNase L, while later on the control is taken by the newly synthesized VP3. Cosilencing the expression of VP3 and RNase L in infected cells yields viral infectious particles at levels similar to those obtained in control infected cells, where no genes were silenced, suggesting that the capping activity of VP3 is not essential for the formation of infectious viral particles.
Rotaviruses represent an important cause of severe gastroenteritis in the young of many animal species, including humans. In this work, we have found that the OAS/RNase L pathway is activated during rotavirus infection, but the virus uses two different strategies to prevent the deleterious effects of this innate immune response of the cell. Early during virus entry, the initial interactions of the viral particle with the cell result in the inhibition of RNase L activity during the first hours of the infection. Later on, once viral proteins are synthesized, the phosphodiesterase activity of VP3 degrades the cellular 2'-5'-oligoadenylates, which are potent activators of RNase L, preventing its activation. This work demonstrates that the OAS/RNase L pathway plays an important role during infection and that the phosphodiesterase activity of VP3 is relevant during the replication cycle of the virus.
固有免疫反应是宿主细胞抵御病毒感染的第一道防线。反过来，病毒也进化出了多种策略来躲避和直接对抗宿主的固有免疫途径。其中一条途径是2'-5'-寡腺苷酸合成酶（OAS）/核糖核酸酶L途径。OAS被双链RNA（dsRNA）激活后产生2'-5'寡腺苷酸，后者是核糖核酸酶L的激活剂；这种酶可降解病毒和细胞RNA，从而限制病毒感染。最近发现，轮状病毒VP3的羧基末端结构域（CTD）具有2'-5'-磷酸二酯酶（PDE）活性，能够在功能上替代小鼠肝炎病毒ns2蛋白的PDE活性。这种特殊的磷酸二酯酶可切割寡腺苷酸的2'-5'-磷酸二酯键，对抗OAS/核糖核酸酶L途径。然而，VP3的这种活性在轮状病毒复制周期中是否相关尚不清楚。在此，我们证明，轮状病毒感染后，OAS/核糖核酸酶L复合物被激活；然而，病毒能够利用至少两种不同机制来控制其活性。在轮状病毒内吞作用期间或之前发生的病毒-细胞相互作用会触发一个信号，阻止核糖核酸酶L的早期激活，而后期则由新合成的VP3进行控制。在感染细胞中共沉默VP3和核糖核酸酶L的表达，所产生的病毒感染性颗粒水平与未沉默任何基因的对照感染细胞中获得的水平相似，这表明VP3的封端活性对于感染性病毒颗粒的形成并非必不可少。
轮状病毒是包括人类在内的许多动物幼崽严重胃肠炎的重要病因。在这项研究中，我们发现OAS/核糖核酸酶L途径在轮状病毒感染期间被激活，但病毒采用两种不同策略来防止细胞这种固有免疫反应的有害影响。在病毒进入早期，病毒颗粒与细胞的初始相互作用导致在感染的最初几个小时内核糖核酸酶L活性受到抑制。随后，一旦合成病毒蛋白，VP3的磷酸二酯酶活性就会降解细胞2'-5'-寡腺苷酸，后者是核糖核酸酶L的有效激活剂，从而阻止其激活。这项研究表明，OAS/核糖核酸酶L途径在感染过程中发挥重要作用，并且VP3的磷酸二酯酶活性在病毒复制周期中具有相关性。
