Mar ThiThi, Liu Wenwen, Wang Xifeng
State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
J Proteomics. 2014 May 6;102:83-97. doi: 10.1016/j.jprot.2014.03.004. Epub 2014 Mar 17.
Southern rice black-streaked dwarf virus (SRBSDV), transmitted by the white-backed planthopper (Sogatella furcifera) in a persistent-propagative manner, has caused serious yield losses in Asia. Here in a yeast two-hybrid system, protein interactions between SRBSDV P7-1 as a bait protein and a cDNA library of S. furcifera as prey protein were assessed. Of 153 proteins identified as putative interactors, 24 were selected for further analysis. Of the 24 proteins, 18 were further confirmed in a chemiluminescent coimmunoprecipitation (Co-IP) assay as true positive interactors with different strengths of interactions. Six potential candidate proteins (neuroglian, myosin light chain 2 [MLC2], polyubiquitin, E3 ubiquitin ligase, ribophorin ii, and profilin) were analyzed for gene expression in five organs by qRT-PCR; mRNA levels were highest in the gut for neuroglian, MLC2, polyubiquitin and profilin, in the salivary glands for ribophorin ii, and in the haemolymph for E3 ubiquitin ligase. A virus-host protein interaction network was constructed using SRBSDV P7-1 and 18 prey positive protein homologs of Drosophila melanogaster. Our findings suggest that these proteins are involved in the complex host reaction to infection by SRBSDV and provide new insights into the molecular basis of transmission.
Southern rice black-streaked dwarf virus (SRBSDV), transmitted by S. furcifera in a persistent-propagative manner, is a new found virus and a tentative member of the genus Fijivirus in the family Reoviridae. It was widely noted by plant virologist, government officials and the farmers in Asia in recent years because of its epidemic outbreak and causing serious yield losses after 2009. However, the molecular mechanism by which SRBSDV successfully infects and replicates in both plant and insect hosts remains unclear, and much less is known about how the virus spreads from initially infected cells to adjacent cells in the insect vector. In the present study, we examined protein interactions between SRBSDV P7-1 as the bait and cDNA library of WBPH as the prey by using yeast two-hybrid system, 153 proteins were identified as putative interactors and 24 putative proteins were selected for chemiluminescent coimmunoprecipitation (Co-IP) assay, and then constructed a viral protein-host protein interaction network with homologs of D. melanogaster. Six WBPH proteins were confirmed as potential P7-1 partners that take part in a pivotal role for viral movement in insect vector. These findings will greatly facilitate the understanding of the transmission mechanisms of SRBSDV by its insect vector. This is the first to study the molecular interaction between SRBSDV and its insect vector.
南方水稻黑条矮缩病毒（SRBSDV）由白背飞虱（Sogatella furcifera）以持久性增殖方式传播，已在亚洲造成严重的产量损失。在此，利用酵母双杂交系统评估了作为诱饵蛋白的SRBSDV P7-1与作为猎物蛋白的白背飞虱cDNA文库之间的蛋白质相互作用。在鉴定出的153个推定相互作用蛋白中，选择了24个进行进一步分析。在这24个蛋白中，有18个在化学发光免疫共沉淀（Co-IP）试验中被进一步确认为具有不同相互作用强度的真正阳性相互作用蛋白。通过qRT-PCR分析了6个潜在候选蛋白（神经黏连蛋白、肌球蛋白轻链2 [MLC2]、多聚泛素、E3泛素连接酶、核糖体结合蛋白ii和肌动蛋白结合蛋白）在五个器官中的基因表达；神经黏连蛋白、MLC2、多聚泛素和肌动蛋白结合蛋白的mRNA水平在肠道中最高，核糖体结合蛋白ii在唾液腺中最高，E3泛素连接酶在血淋巴中最高。利用SRBSDV P7-1和果蝇的18个猎物阳性蛋白同源物构建了病毒-宿主蛋白相互作用网络。我们的研究结果表明，这些蛋白参与了宿主对SRBSDV感染的复杂反应，并为传播的分子基础提供了新的见解。
南方水稻黑条矮缩病毒（SRBSDV）由白背飞虱以持久性增殖方式传播，是一种新发现的病毒，是呼肠孤病毒科斐济病毒属的暂定成员。近年来，由于其疫情爆发并在2009年后造成严重产量损失，受到了亚洲植物病毒学家、政府官员和农民的广泛关注。然而，SRBSDV在植物和昆虫宿主中成功感染和复制的分子机制仍不清楚，关于病毒如何从最初感染的细胞传播到昆虫介体中的相邻细胞知之甚少。在本研究中，我们利用酵母双杂交系统检测了作为诱饵的SRBSDV P7-1与白背飞虱cDNA文库作为猎物之间的蛋白质相互作用，鉴定出153个推定相互作用蛋白，选择24个推定蛋白进行化学发光免疫共沉淀（Co-IP）试验，然后与果蝇同源物构建了病毒蛋白-宿主蛋白相互作用网络。6个白背飞虱蛋白被确认为潜在的P7-1伙伴，它们在昆虫介体中的病毒移动中起关键作用。这些发现将极大地促进对SRBSDV通过其昆虫介体传播机制的理解。这是首次研究SRBSDV与其昆虫介体之间的分子相互作用。
