DEAH BOX HELICASE 9 (DHX9) AS A BIDIRECTIONAL REGULATOR IN ADENOSINE-TO-INOSINE (A-TO-I) EDITING

Abstract

Adenosine-to-Inosine (A-to-I) editing entails the enzymatic deamination of adenosines to inosines by Adenosine DeAminases acting on RNA (ADARs). Substrate structures play a central role in substrate selectivity and specificity of ADARs. RNA helicases, a family of ATP-driven enzymes remodelling RNA structures and ribonucleoprotein complexes may therefore have a regulatory role in A-to-I editing. In our study, we elucidated the regulatory roles of DEAH-box helicase 9 (DHX9) in A-to-I editing. ADARs and DHX9 form a complex in the nucleus, the primary site of A-to-I editing. To investigate the functional role of DHX9 in editing, we examined the global editome upon DHX9 knockdown using RNA sequencing. Intriguingly, we uncovered that DHX9 functions as a bidirectional regulator and the direction of change is largely dependent on the ADAR specificity of the sites. DHX9 knockdown represses editing of ADAR1-specific substrates and augments editing of ADAR2-specific substrates. The failure to rescue the knockdown phenotype with DHX9 helicase mutant (DHX9K417R) indicates that the helicase activity is essential for A-to-I editing regulation. In sum, DHX9 constitutes a bidirectional mode of regulation in A-to-I editing. Functionally, DHX9 is an essential gene as DHX9 suppression has deleterious effects on foci formation, anchorage-independent growth and in vivo xenograft tumour formation.