Respiratory syncytial virus M2-1 protein associates non-specifically with viral messenger RNA and with specific cellular messenger RNA transcripts

• Type of Material: Article
• Authors: Molly R Braun, Boston University; Sarah L Noton, Boston University; Emmeline L Blanchard, Georgia Institute of Technology; Afzaal Shareef, Boston University; Philip Santangelo, Emory University; Evan W Johnson, Boston University; Rachel Fearns, Boston University
• Language: English
• Date: 2021-05-01
• Publisher: PUBLIC LIBRARY SCIENCE
• Publication Version: Final Published Version
• Copyright Statement: © 2021 Braun et al
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1371/journal.ppat.1009589
• Title of Journal or Parent Work: PLOS PATHOGENS
• Volume: 17
• Issue: 5
• Start Page: e1009589
• End Page: e1009589
• Grant/Funding Information: This work was supported by the National Institute of Allergy and Infectious Diseases of the National Institutes of Health in a grant to RF under award number R01AI113321 (https://www.niaid.nih.gov). EB was also funded by NSF GFRP under award number DGE-1650044 (https://www.nsfgrfp.org). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
• Keywords: EBOLA-VIRUS; ANTITERMINATOR M2-1; YOUNG-CHILDREN; Parasitology; TRAILER REGION; HUMAN METAPNEUMOVIRUS; INCLUSION-BODIES; ELONGATION-FACTOR; CRYSTAL-STRUCTURE; Virology; Science & Technology; Life Sciences & Biomedicine; Microbiology; PHOSPHOPROTEIN-P; MARBURG-VIRUS
• Research Categories: Biology, Biostatistics; Engineering, Biomedical

Abstract

Respiratory syncytial virus (RSV) is a major cause of respiratory disease in infants and the elderly. RSV is a non-segmented negative strand RNA virus. The viral M2-1 protein plays a key role in viral transcription, serving as an elongation factor to enable synthesis of full-length mRNAs. M2-1 contains an unusual CCCH zinc-finger motif that is conserved in the related human metapneumovirus M2-1 protein and filovirus VP30 proteins. Previous biochemical studies have suggested that RSV M2-1 might bind to specific virus RNA sequences, such as the transcription gene end signals or poly A tails, but there was no clear consensus on what RSV sequences it binds. To determine if M2-1 binds to specific RSV RNA sequences during infection, we mapped points of M2-1:RNA interactions in RSV-infected cells at 8 and 18 hours post infection using crosslinking immunoprecipitation with RNA sequencing (CLIP-Seq). This analysis revealed that M2-1 interacts specifically with positive sense RSV RNA, but not negative sense genome RNA. It also showed that M2-1 makes contacts along the length of each viral mRNA, indicating that M2-1 functions as a component of the transcriptase complex, transiently associating with nascent mRNA being extruded from the polymerase. In addition, we found that M2-1 binds specific cellular mRNAs. In contrast to the situation with RSV mRNA, M2-1 binds discrete sites within cellular mRNAs, with a preference for A/U rich sequences. These results suggest that in addition to its previously described role in transcription elongation, M2-1 might have an additional role involving cellular RNA interactions.