RNase Z Oxidative Degradation Impedes tRNA Maturation and is Involved in Streptococcal Translation Regulation in Response to Oxidative Stress

When encountering oxidative stress, organisms selectively upregulate antioxidant genes and simultaneously suppress the translation of most other proteins. Eukaryotes employ multiple strategies to adjust translation at both the initiation and elongation stages; however, how prokaryotes modulate translation under oxidative stress remains unclear. Here, we report that upon hydrogen peroxide (H2O2) challenge, Streptococcus oligofermentans reduced translation via RNase Z (So-RNaseZ) oxidative degradation, thus hindering tRNA maturation. S. oligofermentans encodes all CCA-less tRNAs that require So-RNaseZ for 3' end maturation. A combination of nonreducing SDS-PAGE and liquid chromatography/tandem mass spectrometry (LC/MS-MS) assays demonstrated that H2O2 oxidation induced Cys38-Cys149 disulfide linkages in recombinant So-RNaseZ protein, and serine substitution of Cys38 or Cys149 abolished these disulfide linkages. Consistently, redox Western blotting also determined intramolecular disulfide-linked So-RNaseZ in H2O2-treated S. oligofermentans cells. The disulfide-linked So-RNaseZ and monomer were both subject to proteolysis, whereas C149S mutation alleviated oxidative degradation of So-RNaseZ, suggesting that H2O2-mediated disulfide linkages substantially contributed to So-RNaseZ degradation. Accordingly, Northern blotting determined that tRNA precursor accumulation and mature tRNA species decrease in H2O2-treated S. oligofermentans. Moreover, reduced overall protein synthesis, as indicated by puromycin incorporation, and retarded growth of S. oligofermentans occurred in an H2O2 concentration-dependent manner. Overexpression of So-RNaseZ not only elevated tRNA precursor processing and protein synthesis but also partly rescued H2O2-suppressed S. oligofermentans growth. Moreover, So-RNaseZ oxidative degradation-mediated translation repression elevated S. oligofermentans survival under high H2O2 stress. Therefore, this work found that So-RNaseZ oxidative degradation-impeded tRNA maturation contributes to streptococcal translation repression and provides the oxidative stress adaptability for S. oligofermentans. IMPORTANCE Translation regulation is a common strategy used by organisms to reduce oxidative damage. Catalase-negative streptococci produce as well as tolerate high levels of H2O2. This work reports a novel translation regulation mechanism employed by Streptococcus oligofermentans in response to H2O2 challenge, in which the key tRNA endonuclease So-RNaseZ is oxidized to form Cys38-Cys149 disulfide linkages and both the disulfide-linked So-RNaseZ and monomers are subject to proteolysis; thus, tRNA maturation, protein translation, and growth are all suppressed. Notably, So-RNaseZ oxidative degradation-mediated translation repression offers oxidative adaptability to S. oligofermentans and enhances its survival against high H2O2 challenge. So-RNaseZ orthologs and H2O2-sensitive cysteines (Cys38 and Cys149) are widely distributed in Streptococcus and Lactococcus species genomes, which also encode all CCA-less tRNAs and lack catalase. Therefore, RNase Z oxidative degradation-based translation regulation could be widely employed by these lactic acid bacteria, including pathogenic streptococci, to cope with H2O2.