Shahiduzzaman Md, Ezatti Peyman, Xin Gang, Coombs Kevin M
Department of Medical Microbiology, Faculty of Medicine, University of Manitoba , Winnipeg, Manitoba R3E 0J9, Canada.
J Proteome Res. 2014 May 2;13(5):2223-38. doi: 10.1021/pr5001779. Epub 2014 Apr 7.
Interactions between viruses and their host cells are important determinants of virus replication and of immune responses to the virus. However, these interactions and resulting consequences of these interactions remain poorly defined. Numerous recent quantitative proteomic approaches have measured host proteins affected by virus infection. Here, we used activity-based protein profiling (ABPP) to measure functional alterations in host serine hydrolases after influenza A virus infection of Madin-Darby canine kidney and human A549 lung cells. We identified 62 serine proteases. We then combined the ABPP approach with stable isotope labeling to directly measure how serine hydrolase activities were affected by virus infection. Differentially regulated SHs mapped into a few key cellular pathway systems, most notably the proteasomal system. The specific serine protease inhibitors Aprotinin and Pefablock and specific proteasomal inhibitors Bortezomib and MG132 significantly inhibited influenza virus growth. Some inhibitors also down-regulated activities of several proteasomal proteins, including PSMA1, PSMA2, and PMSB3. Genetic knockdown of PMSA2 also attenuated influenza virus replication. These findings further our understanding of enzymatic cellular processes affected by influenza virus and may be beneficial in the search for additional antiviral therapeutic targets.
病毒与其宿主细胞之间的相互作用是病毒复制以及机体对病毒免疫反应的重要决定因素。然而，这些相互作用及其产生的后果仍未得到充分阐明。最近有许多定量蛋白质组学方法用于检测受病毒感染影响的宿主蛋白。在此，我们运用基于活性的蛋白质谱分析（ABPP）技术，来检测甲型流感病毒感染麦迪逊-达比犬肾细胞和人A549肺细胞后宿主丝氨酸水解酶的功能变化。我们鉴定出了62种丝氨酸蛋白酶。随后，我们将ABPP方法与稳定同位素标记相结合，以直接测定病毒感染对丝氨酸水解酶活性的影响。差异调节的丝氨酸水解酶映射到一些关键的细胞信号通路系统中，最显著的是蛋白酶体系统。特异性丝氨酸蛋白酶抑制剂抑肽酶和苯甲磺酰氟以及特异性蛋白酶体抑制剂硼替佐米和MG132能显著抑制流感病毒的生长。一些抑制剂还下调了几种蛋白酶体蛋白的活性，包括蛋白酶体α亚基1、蛋白酶体α亚基2和蛋白酶体β亚基3。蛋白酶体α亚基2的基因敲低也减弱了流感病毒的复制。这些发现加深了我们对受流感病毒影响的细胞酶促过程的理解，可能有助于寻找更多的抗病毒治疗靶点。
