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The interaction of Rotavirus A pig/China/NMTL/2008/G9P[23] VP6 with cellular beta-actin is required for optimal RV replication and infectivity

Author:
Yuan, Jing, Zhang, Xin, Shi, Hongyan, Chen, Jianfei, Han, Xiao, Wei, Ping, Feng, Li
Source:
Veterinary microbiology 2016 v.197 pp. 111-121
ISSN:
0378-1135
Subject:
Rotavirus, actin, capsid, endoplasmic reticulum, glutathione transferase, gold, mass spectrometry, microfilaments, microscopy, mitochondria, pathogenicity, polymerization, precipitin tests, ribosomes, small interfering RNA, swine, tropomyosins, virion, China
Abstract:
VP6 forms the intermediate layer of the rotavirus (RV) capsid, and it plays important roles after RV penetration and uncoating. These functions rely on its ability to interact with host cell proteins. To gain further insights into the role of VP6 in porcine RV (PoRV) infection, a glutathione S-transferase pull-down assay was utilized to find unknown cellular factors that interact with VP6. In this study, beta-actin, tropomyosin 1, and 40S ribosomal protein S16 were identified as interaction partners of VP6 by mass spectrometry and co-immunoprecipitation. The interaction with beta-actin was further studied. By immunoelectron microscopy, we observed VP6 proteins that labeled with colloidal gold localized on the actin microfilaments at the early stage of PoRV infection, we also found VP6 distributed in the ribosome, mitochondria, endoplasmic reticulum and nucleus in the infected cells. Actin binding protein spin-down assays verified PoRV double-layered particles (DLPs) bound to F-actin in vitro, but didn’t have actin polymerization enhancement activity. After a small interfering RNA (siACTB) was used to knock down beta-actin expression, PoRV VP6 expression and the infection rates of newly synthesized virions releasing into culture supernatants decreased dramatically. Our results confirm and extend previous reports indicating that the interaction between PoRV VP6 and beta-actin plays vital roles in the PoRV lifecycle.
Agid:
5585589