Diribonuclease activity eliminates toxic diribonucleotide accumulation

RNA degradation is a central process in cells that is vital for transcriptional regulation. The degradation process is initiated by endoribonucleolytic fragmentation of long RNA polymers, followed by exoribonucleolytic cleavage to release mononucleotides from the fragment ends. In this process, the degradation of short oligonucleotides, especially diribonucleotides, into mononucleotides requires specific enzymatic activity, which in Gram-negative bacteria is provided by oligoribonuclease (Orn). Orn is unique in that it is the only essential exoribonuclease in Escherichia coli. Recent studies have shown that Orn cleaves only diribonucleotides under physiological conditions, suggesting that the accumulation of dinucleotides poisons cells. Yet, related organisms, such as Pseudomonas aeruginosa, display a growth defect but are viable without Orn, contesting its essentiality. Here, we took advantage of P. aeruginosa orn mutants to elucidate the mechanisms of their survival. Genetic screening for suppressors that restored colony morphology identified yciV. Purified YciV exhibited diribonuclease activity. YciV is present in all g-proteobacteria, suggesting that YciV from P. aeruginosa is distinct from its E. coli ortholog. Phylogenetic analysis revealed differences between the two orthologs that were mapped to the active site of the enzyme and correlated with differences in substrate profiles. The expression of P. aeruginosa YciV in E. coli eliminated the necessity of orn. Similarly, the deletion of yciV from P. aeruginosa caused orn to become essential. Together, these results show that diribonuclease activity is required in g-proteobacteria and that diribonucleotides are utilized to monitor the efficacy of RNA degradation. Because Orn is found in higher eukaryotes, these observations indicate a conserved mechanism for monitoring RNA degradation.