Initiation of RNA Polymerization and Polymerase Encapsidation by a Small dsRNA Virus

PLoS Pathogens

Yusong R. Guo¹  Yizhi J. Tao¹  Yukimatsu Toh¹  Aaron M. Collier¹  Minna M. Poranen²  Outi L. Lyytinen² 

[1] Department of BioSciences, Rice University, Houston, Texas, United States of America;Department of Biosciences, University of Helsinki, Helsinki, Finland

关键词: Polymerases;    Viral packaging;    RNA synthesis;    dsRNA viruses;    Viral structure;    Sequence motif analysis;    RNA structure;    Viral genomics;   

DOI  :  10.1371/journal.ppat.1005523

学科分类：生物科学（综合）

来源: Public Library of Science

【 摘 要 】

During the replication cycle of double-stranded (ds) RNA viruses, the viral RNA-dependent RNA polymerase (RdRP) replicates and transcribes the viral genome from within the viral capsid. How the RdRP molecules are packaged within the virion and how they function within the confines of an intact capsid are intriguing questions with answers that most likely vary across the different dsRNA virus families. In this study, we have determined a 2.4 Å resolution structure of an RdRP from the human picobirnavirus (hPBV). In addition to the conserved polymerase fold, the hPBV RdRP possesses a highly flexible 24 amino acid loop structure located near the C-terminus of the protein that is inserted into its active site. In vitro RNA polymerization assays and site-directed mutagenesis showed that: (1) the hPBV RdRP is fully active using both ssRNA and dsRNA templates; (2) the insertion loop likely functions as an assembly platform for the priming nucleotide to allow de novo initiation; (3) RNA transcription by the hPBV RdRP proceeds in a semi-conservative manner; and (4) the preference of virus-specific RNA during transcription is dictated by the lower melting temperature associated with the terminal sequences. Co-expression of the hPBV RdRP and the capsid protein (CP) indicated that, under the conditions used, the RdRP could not be incorporated into the recombinant capsids in the absence of the viral genome. Additionally, the hPBV RdRP exhibited higher affinity towards the conserved 5’-terminal sequence of the viral RNA, suggesting that the RdRP molecules may be encapsidated through their specific binding to the viral RNAs during assembly.

【 授权许可】

CC BY