Endonucleolytic cleavages by RNase E generate the mature 3′ termini of the three proline tRNAs in Escherichia coli

We demonstrate here for the first time that proline tRNA 3′ end maturation in Escherichia coli employs a one-step endonucleolytic pathway that does not involve any of the six 3′ → 5′ exonucleases (RNase T, RNase PH, RNase D, RNase BN, RNase II and polynucleotide phosphorylase [PNPase]) to generate the mature CCA terminus. Rather, RNase E is primarily responsible for the endonucleolytic removal of the entire Rho-independent transcription terminator associated with the proK, proL and proM primary transcripts by cleaving immediately downstream of the CCA determinant. In the absence of RNase E, RNase G and RNase Z are weakly able to process the proK and proM transcripts, while PNPase and RNase P are utilized in the processing of proL. The terminator fragment derived from the endonucleolytic cleavage of proL transcript is degraded through a PNPase-dependent pathway. It is not clear which enzymes degrade the proK and proM terminator fragments. Our data also suggest that the mature 5′ nucleotide of the proline tRNAs may be responsible for the cleavage specificity of RNase E at the 3′ terminus.