Cryo-EM structure of the respiratory syncytial virus RNA polymerase

Dongdong Cao,Anna Antonova,Yunrong Gao,Julia Slack,Lisa Zhuang,Gabriela Forero,Mason Domke,Puneet Juneja,Sarah Romanelli,Samantha Rice,Bo Liang,Paul D’Cunha,Claire Roesler,Shayon Keating

doi:10.1038/s41467-019-14246-3

Dongdong Cao, Anna Antonova + Show 12 more

Open Access

https://doi.org/10.1038/s41467-019-14246-3

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Journal: Nature Communications	Publication Date: Jan 17, 2020
Citations: 61	License type: open-access

Affiliation: Emory University

Abstract

The respiratory syncytial virus (RSV) RNA polymerase, constituted of a 250 kDa large (L) protein and tetrameric phosphoprotein (P), catalyzes three distinct enzymatic activities — nucleotide polymerization, cap addition, and cap methylation. How RSV L and P coordinate these activities is poorly understood. Here, we present a 3.67 Å cryo-EM structure of the RSV polymerase (L:P) complex. The structure reveals that the RNA dependent RNA polymerase (RdRp) and capping (Cap) domains of L interact with the oligomerization domain (POD) and C-terminal domain (PCTD) of a tetramer of P. The density of the methyltransferase (MT) domain of L and the N-terminal domain of P (PNTD) is missing. Further analysis and comparison with other RNA polymerases at different stages suggest the structure we obtained is likely to be at an elongation-compatible stage. Together, these data provide enriched insights into the interrelationship, the inhibitors, and the evolutionary implications of the RSV polymerase.

Highlights

The respiratory syncytial virus (RSV) RNA polymerase, constituted of a 250 kDa large (L) protein and tetrameric phosphoprotein (P), catalyzes three distinct enzymatic activities — nucleotide polymerization, cap addition, and cap methylation
Our studies reveal that the RNA dependent RNA polymerase (RdRp) and Cap domains of the RSV L shares similar architectures of that of the vesicular stomatitis virus (VSV) L40, uncover a previously unknown basis of how P interacts with L, and provide molecular insights into RNA synthesis by the
The interactions between L and a tetrameric P can be divided into three parts: (1) two of four-helix bundles of POD and the RdRp; (2) two flexible PCTD chains wrap around the surface of RdRp and stabilize the base of the tetrameric POD; (3) one flexible PCTD chain extends to the positively charged “palm” motif of RdRp and the edge of the putative NTP entrance channel (Fig. 2b–d)

Summary

Introduction

The respiratory syncytial virus (RSV) RNA polymerase, constituted of a 250 kDa large (L) protein and tetrameric phosphoprotein (P), catalyzes three distinct enzymatic activities — nucleotide polymerization, cap addition, and cap methylation. RSV initiates viral infection by delivering into the host cell a virus-specific RNA synthesis machine required for both genome replication and gene transcription[4,5]. This machine comprises the nucleoprotein (N) coated genomic RNA (N:RNA) and the RNA polymerase[6]. The catalytic core is a 250 kDa large (L) protein that catalyzes the RNA polymerization in both replication and transcription, the cap addition, and cap methylation of nascent viral mRNAs. A tetrameric phosphoprotein (P) is essential to modulate and constitute an active RNA polymerase with L4,5. The RSV cap addition and cap methylation assays have not been described yet and are speculated to share similar mechanisms with vesicular stomatitis virus (VSV), of which the assays were reported[16,17,18,24,25]

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