RNA Binding Proteins KhpA and KhpB Interact with Small Regulatory RNAs and Affect Global Gene Expression in Deinococcus radiodurans

Abstract

Small regulatory RNAs (sRNAs) in bacteria often associate with RNA-binding proteins to gain intracellular stability and/or to enable regulatory efficiency. While much of the current knowledge about those sRNA binding proteins is derived from studies in Gram-negative organisms, the characterization of such proteins in Gram-positive species is still lagging behind. Here, we identified and characterized two sRNA binding proteins (KhpA and KhpB) in Deinococcus radiodurans, a Gram-positive bacterium that exhibits extreme resistance to radiation and other oxidative stressors. We demonstrate that KhpA and KhpB interact with key sRNAs in D. radiodurans and influence their stabilities. Although KhpA and KhpB interact with each other, they do not bind the sRNAs as a complex. Insightfully, KhpA and KhpB facilitate the interactions of the representative sRNAs PprS and Dsr9 in D. radiodurans with their respective mRNA targets, pprM and DR_1968. Through RNA-seq analysis, we further revealed that KhpA and KhpB have both overlapping and specific roles in a global gene regulation in D. radiodurans. Overall, this study expands our knowledge of posttranscriptional regulation in D. radiodurans and supports the growing consensus that KhpA and KhpB homologs constitute a new family of sRNA binding proteins in Gram-positive bacteria. IMPORTANCEThe bacterium Deinococcus radiodurans is the most radiation-resistant organism identified to date. Understanding the mechanisms of resistance of D. radiodurans is essential for leveraging this bacterium in biomedical and biomanufacturing applications. It was previously revealed that small regulatory RNAs (sRNAs) play crucial roles in the gene regulation of D. radiodurans. However, how these sRNAs are influenced by RNA binding proteins is poorly understood. Here we identified two conserved RNA binding proteins, KhpA and KhpB, as sRNA binding partners in D. radiodurans. These proteins affect the sRNA stability, sRNA-target interaction, and global gene regulation. Characterization of KhpA and KhpB will help us advance the understanding of how post-transcriptional network regulates the physiology and radioresistance of D. radiodurans.