Data availabilityAll data presented in this manuscript are available from the corresponding author upon reasonable request. Bulk tumour cell RNA sequencing has been deposited at the Gene Expression Omnibus (GEO) under accession number https://www.ncbi.nlm.nih.gov/geo/ query/acc.cgi?acc=GSE110708. Single-cell RNA sequencing of tumour cells were also deposited at the GEO under accession numberhttps://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE110746.

Genomic mutations in key genes are known to drive tumorigenesis and have been the focus of much attention in recent years. However, genetic content also may change farther downstream. RNA editing alters the mRNA sequence from its genomic blueprint in a dynamic and flexible way. A few isolated cases of editing alterations in cancer have been reported previously. Here, we provide a transcriptome-wide characterization of RNA editing across hundreds of cancer samples from multiple cancer tissues, and we show that A-to-I editing and the enzymes mediating this modification are significantly altered, usually elevated, in most cancer types. Increased editing activity is found to be associated with patient survival. As is the case with somatic mutations in DNA, most of these newly introduced RNA mutations are likely passengers, but a few may serve as drivers that may be novel candidates for therapeutic and diagnostic purposes.

Systematic exploration of cancer cell vulnerabilities can inform the development of novel cancer therapeutics. Here, through analysis of genome-scale loss-of-function datasets, we identify adenosine deaminase acting on RNA (ADAR or ADAR1) as an essential gene for the survival of a subset of cancer cell lines. ADAR1-dependent cell lines display increased expression of interferon-stimulated genes. Activation of type I interferon signaling in the context of ADAR1 deficiency can induce cell lethality in non-ADAR1-dependent cell lines. ADAR deletion causes activation of the double-stranded RNA sensor, protein kinase R (PKR). Disruption of PKR signaling, through inactivation of PKR or overexpression of either a wildtype or catalytically inactive mutant version of the p150 isoform of ADAR1, partially rescues cell lethality after ADAR1 loss, suggesting that both catalytic and non-enzymatic functions of ADAR1 may contribute to preventing PKR-mediated cell lethality. Together, these data nominate ADAR1 as a potential therapeutic target in a subset of cancers.

A-to-I RNA editing is an important post-transcriptional modification, known to be altered in tumors. It targets dozens of sites within miRNAs, some of which impact miRNA biogenesis and function, as well as many miRNA recognition sites. However, the full extent of the effect of editing on regulation by miRNAs and its behavior in human cancers is still unknown. Here we systematically characterized miRNA editing in 10 593 human samples across 32 cancer types and normal controls. We find that the majority of previously reported sites show little to no evidence for editing in this dataset, compile a list of 58 reliable miRNA editing sites, and study them across normal and cancer samples. Edited miRNA versions tend to suppress expression of known oncogenes, and, consistently, we observe a clear global tendency for hypo-editing in tumors, in strike contrast to the behavior for mRNA editing, allowing an accurate classification of normal/tumor samples based on their miRNA editing profile. In many cancers this profile correlates with patients' survival. Finally, thousands of miRNA binding sites are differentially edited in cancer. Our study thus establishes the important effect of RNA editing on miRNA-regulation in the tumor cell, with prospects for diagnostic and prognostic applications.

In Drosophila, A-to-I editing is prevalent in the brain, and mutations in the editing enzyme ADAR correlate with specific behavioral defects. Here we demonstrate a role for ADAR in behavioral temperature adaptation in Drosophila. Although there is a higher level of editing at lower temperatures, at 29°C more sites are edited. These sites are less evolutionarily conserved, more disperse, less likely to be involved in secondary structures, and more likely to be located in exons. Interestingly, hypomorph mutants for ADAR display a weaker transcriptional response to temperature changes than wild-type flies and a highly abnormal behavioral response upon temperature increase. In sum, our data shows that ADAR is essential for proper temperature adaptation, a key behavior trait that is essential for survival of flies in the wild. Moreover, our results suggest a more general role of ADAR in regulating RNA secondary structures in vivo.