Nanopore sequencing-based measurement of paramyxovirus RNA editing reveals virus-specific differences in editing efficiency of mRNA, antigenome and genome

Abstract

Paramyxovirus polymerase recognizes an RNA editing signal on the viral genome and transcribes mRNA in which guanine nucleotides are inserted in a template-independent manner. This enables the synthesis of multiple proteins from a single gene, which is important for viral growth. We developed a method to quantify RNA editing efficiency using Oxford Nanopore Technologies’ MinION platform. We performed sequence analysis of reverse transcription-PCR amplicons with the RNA editing sites in cells infected with Sendai virus (SeV) and canine distemper virus (CDV). By modifying reverse transcription primers, we simultaneously assessed RNA editing efficiency in mRNA, antigenome and genome. We observed distinct differences in mRNA editing efficiency between SeV and CDV. Notably, while RNA editing in SeV is confined to mRNA, in CDV it is also observed in antigenome/genome. (Anti)genomes harboring extra nucleotides may deviate from a multiple-of-six sequence, suggesting that RNAs not following the “Rule of Six” are produced in CDV-infected cells.