Multiple exoribonucleases are required for the 3' processing of Escherichia coli tRNA precursors in vivo

Our knowledge of the 3′ processing of tRNA precursors is severely limited. Although six exoribonucleases able to act on Escherichia coli tRNA precursors in vitro have been identified, their involvement in tRNA maturation in vivo has not been demonstrated. Here we show, using a wide range of multiple RNase-deficient strains and a quantitative suppression assay, that at least five of these enzymes -RNase II, RNase D, RNase BN, RNase T, and RNase PH-can participate in the synthesis of functional tRNA^Tyrsu⁺₃ in vivo. Moreover, any one of the five RNases is sufficient to allow tRNA processing to proceed although with varying effectiveness. Examination of the level of aminoacylation of tRNA isolated from RNase-deficient strains suggested that tRNA precursors accumulate in the most defective cells. These data indicate that exoribonucleases are required for tRNA maturation in vivo and that there is a high degree of functional overlap among the enzymes. These studies contribute to the identification of all the enzymes necessary for defining the complete processing pathway for E. coli tRNA precursors.