Characterization of tRNACys processing in a conditional Bacillus subtilis CCase mutant reveals the participation of RNase R in its quality control

We generated a conditional CCase mutant of Bacillus subtilis to explore the participation in vivo of the tRNA nucleotidyltransferase (CCA transferase or CCase) in the maturation of the single-copy tRNA Cys , which lacks an encoded CCA 3′ end. We observed that shorter tRNA Cys species, presumably lacking CCA, only accumulated when the inducible Pspac : cca was introduced into an rnr mutant strain, but not in combination with pnp . We sequenced the tRNA 3′ ends produced in the various mutant tRNA Cys species to detect maturation and decay intermediates and observed that decay of the tRNA Cys occurs through the addition of poly(A) or heteropolymeric tails. A few clones corresponding to full-size tRNAs contained either CCA or other C and/or A sequences, suggesting that these are substrates for repair and/or decay. We also observed editing of tRNA Cys at position 21, which seems to occur preferentially in mature tRNAs. Altogether, our results provide in vivo evidence for the participation of the B. subtilis cca gene product in the maturation of tRNAs lacking CCA. We also suggest that RNase R exoRNase in B. subtilis participates in the quality control of tRNA.