RNA methylation by NSUN4 marks mitochondrial double-stranded RNAs for degradation

Abstract

Human mitochondria have a compact circular genome that encodes 13 protein-coding genes as well as mitochondrial rRNAs and tRNAs. While most protein-coding genes are located in the heavy strand of the genome, the mitochondrial transcription occurs bidirectionally, resulting in the generation of long noncoding RNAs (ncRNAs) from the light strand. Recent studies show that heavy and light strand RNAs interact and generate double-stranded RNAs (mt-dsRNAs) that can activate antiviral signaling. Yet, the post-transcriptional regulation of these mt-dsRNAs remains largely unknown. Here, we employ CRISPR screening on RNA-binding proteins residing in the mitochondrial matrix and identify 5-methylcytosine rRNA methyltransferase NSUN4 as a key regulator of mt-dsRNAs. Methylation by NSUN4 readily occurs on mRNAs and ncRNAs from both heavy and light strands of mitochondria. Cells depleted of NSUN4 show decreased methylated RNA, but increased overall mt-dsRNA levels. We further find that methylated mitochondrial RNAs are recognized by Complement C1q Binding Protein (C1QBP), which degrades the RNA by recruiting PNPase. In line with this, C1QBP-deficient cells show elevated mt-dsRNA expression and increased interferon signature. Our study unveils the post-transcriptional regulation of mt-dsRNAs and establishes RNA methylation as a molecular mark for mitochondrial RNA decay.