Functional overlap of tRNA nucleotidyltransferase, poly(A) polymerase I, and polynucleotide phosphorylase

Repair of the 3'-terminal -CCA sequence of tRNA generally requires the action of the enzyme tRNA nucleotidyltransferase. However, in Escherichia coli in the absence of this enzyme, a decreased level of tRNA end repair continues. To ascertain the enzymes responsible for this residual repair, mutant strains were constructed lacking tRNA nucleotidyltransferase and other enzymes potentially involved in the process, poly(A) polymerase I and polynucleotide phosphorylase (PNPase). Strains lacking tRNA nucleotidyltransferase and either one of the other enzymes displayed decreased growth rates and increased levels of defective tRNA compared with the single cca mutant. Triple mutants lacking all three enzymes grew very slowly, had even more defective tRNA, and were devoid of activity incorporating AMP into tRNA-C-C. Overexpression of poly(A) polymerase I, but not PNPase, partially compensated for the absence of tRNA nucleotidyltransferase. These data show that poly(A) polymerase I and PNPase participate in the end repair process and are required to maintain functional tRNA levels when tRNA nucleotidyltransferase is absent.