RNA editing modulates pathways critical to B cell lymphomagenesis

Diffuse large B cell lymphoma (DLBCL) is one of the most common and aggressive types of B cell lymphoma. Subtype heterogeneity cannot be completely explained by genomic alterations and/or gene expression differences, suggesting that additional mechanisms are involved. Here, we explore the contribution of an epitranscriptomic modification to B cell lymphomagenesis. Our data demonstrate that ADAR1-mediated RNA editing (deamination of adenosine to inosine) targets novel candidate driver genes within well-known disease-associated pathways in DLBCL. Within these pathways, DNA mutations and RNA editing events are often mutually exclusive at the gene level, suggesting that tumours can modulate pathway outcomes by altering sequence at either the genomic or the transcriptomic level. In contrast to what has been found in solid tumours, in DLBCL low ADAR1 expression correlates with low interferon-stimulated gene (ISG) scores and with a microenvironment that is not inflamed. To investigate the molecular mechanism behind the inability of DLBCL to mount an ISG response upon loss of ADAR1, we focused on one candidate MAVS, a central regulator of ISG (as well as NFkB) signaling. MAVS is never mutated but robustly edited by ADAR1 at its 3’UTR, and editing correlates with increased MAVS protein levels and increased downstream activation of the ISG as well as NFkB pathways. Using targeted base editing tools to specifically restore MAVS editing in ADAR1-deficient cells, we demonstrate that MAVS editing is directly causal to an increase in expression of genes downstream this signaling pathway. Our data imply that DLBCL tumours edit MAVS to strengthen NFkB signaling, to which they are dependent on survival; thus, targeted loss of MAVS editing might sensitize tumours to cell death. Overall, ADAR1-mediated RNA editing represents a new tumour evasion mechanism underlying DLBCL lymphomagenesis, with features distinct from those derived from solid tumours to date. ADAR1 KO 3 replicates were generated for RC-K8 and A549 cell lines. 3 WT replicates from A549 cell line, and 3 ctrl clones from RC-K8 cell line (here called WT), were employed as controls. Total RNA was extracted from ADAR1 KO and WT replicates with the RNAeasy Plus mini kit from Qiagen (Cat No./ID: 74134). The total RNA extracted was treated with DNase (TURBO, Thermofisher, Cat No. AM1907). Libraries for total RNA-seq were generated according to Illumina instructions for 125bp paired-end sequencing with universal adapters.