In situ structures of polymerase complex of mammalian reovirus illuminate RdRp activation and transcription regulation

Author:

Bao Keyan12,Zhang Xueli12ORCID,Li Dongyu12ORCID,Sun Wei3,Sun Zhenzhao3,Wang Jingfei3ORCID,Zhu Ping12ORCID

Affiliation:

1. National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China

2. University of Chinese Academy of Sciences, Beijing 100049, China

3. State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150069, China

Abstract

Mammalian reovirus (reovirus) is a multilayered, turreted member of Reoviridae characterized by transcription of dsRNA genome within the innermost capsid shell. Here, we present high-resolution in situ structures of reovirus transcriptase complex in an intact double-layered virion, and in the uncoated single-layered core particles in the unloaded, reloaded, pre-elongation, and elongation states, respectively, obtained by cryo-electron microscopy and sub-particle reconstructions. At the template entry of RNA-dependent RNA polymerase (RdRp), the RNA-loading region gets flexible after uncoating resulting in the unloading of terminal genomic RNA and inactivity of transcription. However, upon adding transcriptional substrates, the RNA-loading region is recovered leading the RNAs loaded again. The priming loop in RdRp was found to play a critical role in regulating transcription, which hinders the elongation of transcript in virion and triggers the rearrangement of RdRp C-terminal domain (CTD) during elongation, resulting in splitting of template-transcript hybrid and opening of transcript exit. With the integration of these structures, a transcriptional model of reovirus with five states is proposed. Our structures illuminate the RdRp activation and regulation of the multilayered turreted reovirus.

Funder

National Natural Science Foundation of China

Chinese Ministry of Science and Technology

Chinese Academy of Sciences

State Key Laboratory of Veterinary Biotechnology Foundation

National Laboratory of Biomacromolecules of China

Publisher

Proceedings of the National Academy of Sciences

Subject

Multidisciplinary

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