Global analysis of the RNA-RNA interactome inAcinetobacter baumanniiAB5075 uncovers a small regulatory RNA repressing the virulence-related outer membrane protein CarO

Abstract

ABSTRACTAcinetobacter baumanniiis an opportunistic Gram-negative pathogen that infects critically ill patients. The emergence of antimicrobial resistantA. baumanniihas exacerbated the need to functionally characterise environmental adaptation, antibiotic resistance and pathogenicity of this organism and their genetic regulators to inform intervention strategies. Critical to rapid adaptation to changing environments in bacteria are small regulatory RNAs (sRNAs), however, the role that sRNAs play in the biology ofA. baumanniiis poorly understood. To assess the regulatory function of sRNAs and to uncover their RNA interaction partners inA. baumannii, we employed an RNA proximity ligation and sequencing method (Hi-GRIL-seq) in three different environmental conditions. We found that 40 sRNA candidates were ligated to sRNA-RNA chimeric sequencing reads, suggesting that sRNA-mediated gene regulation is pervasive inA. baumanniiand that sRNAs act as direct regulators of mRNA molecules through antisense base-pairing. In-depth characterisation uncovered the sRNA Aar to be a post-transcriptional regulator of four mRNA targets including that of the outer membrane protein CarO and the siderophore receptor BfnH. We show that Aar initiates base-pairing with these mRNA molecules using a conserved seed region of nine nucleotides, sequestering the ribosome binding sites and inhibiting translation. Aar is differentially expressed in response to multiple stress stimuli suggesting a role in fine-tuning translation of the Aar-target molecules inA. baumanniiunder hostile conditions. Together, our study provides mechanistic insights into sRNA-mediated gene expression control inA. baumanniiand represents a valuable resource for future RNA-centric research endeavours in this ESKAPE pathogen.