Unité de Génétique Moléculaire des Bunyavirus, Institut Pasteur, Paris, France.
J Virol. 2013 Feb;87(3):1631-48. doi: 10.1128/JVI.02795-12. Epub 2012 Nov 21.
Rift Valley fever virus (RVFV) is a Phlebovirus (Bunyaviridae family) transmitted by mosquitoes. It infects humans and ruminants, causing dramatic epidemics and epizootics in Africa, Yemen, and Saudi Arabia. While recent studies demonstrated the importance of the nonstructural protein NSs as a major component of virulence in vertebrates, little is known about infection of mosquito vectors. Here we studied RVFV infection in three different mosquito cell lines, Aag2 cells from Aedes aegypti and U4.4 and C6/36 cells from Aedes albopictus. In contrast with mammalian cells, where NSs forms nuclear filaments, U4.4 and Aag2 cells downregulated NSs expression such that NSs filaments were never formed in nuclei of U4.4 cells and disappeared at an early time postinfection in the case of Aag2 cells. On the contrary, in C6/36 cells, NSs nuclear filaments were visible during the entire time course of infection. Analysis of virus-derived small interfering RNAs (viRNAs) by deep sequencing indicated that production of viRNAs was very low in C6/36 cells, which are known to be Dicer-2 deficient but expressed some viRNAs presenting a Piwi signature. In contrast, Aag2 and U4.4 cells produced large amounts of viRNAs predominantly matching the S segment and displaying Dicer-2 and Piwi signatures. Whereas 21-nucleotide (nt) Dicer-2 viRNAs were prominent during early infection, the population of 24- to 27-nt Piwi RNAs (piRNAs) increased progressively and became predominant later during the acute infection and during persistence. In Aag2 and U4.4 cells, the combined actions of the Dicer-2 and Piwi pathways triggered an efficient antiviral response permitting, among other actions, suppression of NSs filament formation and allowing establishment of persistence. In C6/36 cells, Piwi-mediated RNA interference (RNAi) appeared to be sufficient to mount an antiviral response against a secondary infection with a superinfecting virus. This study provides new insights into the role of Dicer and Piwi in mosquito antiviral defense and the development of the antiviral response in mosquitoes.
裂谷热病毒（RVFV）是一种布尼亚病毒（布尼亚病毒科），通过蚊子传播。它感染人类和反刍动物，在非洲、也门和沙特阿拉伯引发了大规模的流行和动物疫情。尽管最近的研究表明非结构蛋白 NSs 是脊椎动物毒力的主要组成部分，但对于蚊子载体的感染知之甚少。在这里，我们研究了三种不同的蚊子细胞系中的 RVFV 感染，即埃及伊蚊的 Aag2 细胞和白纹伊蚊的 U4.4 和 C6/36 细胞。与哺乳动物细胞不同，NSs 在哺乳动物细胞中形成核丝，U4.4 和 Aag2 细胞下调了 NSs 的表达，使得 NSs 丝从未在 U4.4 细胞的核中形成，并在 Aag2 细胞感染后早期消失。相反，在 C6/36 细胞中，在整个感染过程中都可以看到 NSs 核丝。通过深度测序分析病毒衍生的小干扰 RNA（siRNA）表明，在 C6/36 细胞中，siRNA 的产生非常低，已知 C6/36 细胞缺乏 Dicer-2，但表达了一些具有 Piwi 特征的 siRNA。相比之下，Aag2 和 U4.4 细胞产生大量的 siRNA，主要与 S 片段匹配，并显示出 Dicer-2 和 Piwi 的特征。虽然 21 核苷酸（nt）的 Dicer-2 siRNA 在早期感染时很突出，但 24-27nt 的 Piwi RNA（piRNA）的数量逐渐增加，并在急性感染和持续感染期间变得更为突出。在 Aag2 和 U4.4 细胞中，Dicer-2 和 Piwi 途径的联合作用触发了有效的抗病毒反应，除了其他作用外，还抑制了 NSs 丝的形成，并允许建立持续感染。在 C6/36 细胞中，Piwi 介导的 RNA 干扰（RNAi）似乎足以对继发感染的超级感染病毒产生抗病毒反应。本研究为 Dicer 和 Piwi 在蚊子抗病毒防御和蚊子抗病毒反应的发展中的作用提供了新的见解。
