Chaturvedi Sonali, Seo Jang-Kyun, Rao A L N
Department of Plant Pathology & Microbiology, University of California, Riverside, CA 92521-0122, United States.
Department of Plant Pathology & Microbiology, University of California, Riverside, CA 92521-0122, United States.
Virology. 2016 Jul;494:47-55. doi: 10.1016/j.virol.2016.04.001. Epub 2016 Apr 11.
Here, we evaluated the role of two host proteins, a Bromo domain containing RNA binding protein (BRP1) and Glyceraldehyde 3-phosphate dehydrogenase (GAPDH), in the replication of Cucumber mosaic virus (CMV). LC-MS/MS analysis of host/viral proteins pull down against BRP1 from CMV-infected plants co-infiltrated with BRP1-FLAG agroconstruct identified that BRP1 specifically interacts with a ten amino acid motif (843-SPQDVVPLVR-852) encompassing the helicase domain of replicase protein p1a. The interaction between BRP1 and p1a was subsequently confirmed using a BiFC assay. Among fourteen other host proteins identified to interact with BRP1 during CMV infection, six were found to block accumulation of viral progeny in Arabidopsis thaliana lines defective in each of these host proteins. Additional BiFC assays followed by trans-complementation assays identified that plant lines defective in the expression of GAPDH blocked CMV replication by interfering with p1a:p2a interaction. Distinct roles of BRP1 and GAPDH in the replication of CMV are discussed.
在此，我们评估了两种宿主蛋白，即含溴结构域的RNA结合蛋白（BRP1）和甘油醛-3-磷酸脱氢酶（GAPDH）在黄瓜花叶病毒（CMV）复制中的作用。对用BRP1-FLAG农杆菌构建体共浸润的CMV感染植物中与BRP1相互作用的宿主/病毒蛋白进行的液相色谱-串联质谱（LC-MS/MS）分析表明，BRP1与一个包含复制酶蛋白p1a解旋酶结构域的十个氨基酸基序（843-SPQDVVPLVR-852）特异性相互作用。随后使用双分子荧光互补（BiFC）分析证实了BRP1与p1a之间的相互作用。在CMV感染期间鉴定出的与BRP1相互作用的其他十四个宿主蛋白中，发现有六个在这些宿主蛋白各自缺陷的拟南芥品系中阻断了病毒后代的积累。随后进行的补充实验的额外BiFC分析表明，GAPDH表达缺陷的植物品系通过干扰p1a:p2a相互作用来阻断CMV复制。本文讨论了BRP1和GAPDH在CMV复制中的不同作用。
