Identification of RNase T as a high-copy suppressor of the UV sensitivity associated with single-strand DNA exonuclease deficiency in Escherichia coli.
There are three known single-strand DNA-specific exonucleases in Escherichia coli: RecJ, exonuclease I (ExoI), and exonuclease VII (ExoVII). E. coli that are deficient in all three exonucleases are abnormally sensitive to UV irradiation, most likely because of their inability to repair lesions that block replication. We have performed an iterative screen to uncover genes capable of ameliorating the UV repair defect of xonA (ExoI-) xseA (ExoVII-) recJ triple mutants. In this screen, exonuclease-deficient cells were transformed with a high-copy E. coli genomic library and then irradiated; plasmids harvested from surviving cells were used to seed subsequent rounds of transformation and selection. After several rounds of selection, multiple plasmids containing the rnt gene, which encodes RNase T, were found. An rnt plasmid increased the UV resistance of a xonA xseA recJ mutant and uvrA and uvrC mutants; however, it did not alter the survival of xseA recJ or recA mutants. RNase T also has amino acid sequence similarity to other 3' DNA exonucleases, including ExoI. These results suggest that RNase T may possess a 3' DNase activity capable of substituting for ExoI in the recombinational repair of UV-induced lesions.
Department of Biology and Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, Massachusetts 02454-9110, USA.
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Viswanathan
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Lanjuin
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Lovett
D001426:Bacterial Proteins / Q000235:genetics; D004277:DNA, Single-Stranded / Q000502:physiology*; D003851:Deoxyribonucleases / Q000502:physiology; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics* / Q000528:radiation effects*; D029968:Escherichia coli Proteins; D005090:Exodeoxyribonucleases / Q000235:genetics / Q000502:physiology*; D005095:Exoribonucleases / Q000502:physiology*; D015723:Gene Library; D005798:Genes, Bacterial; D005820:Genetic Testing; D005838:Genotype; D008954:Models, Biological; D009154:Mutation; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic; D013489:Suppression, Genetic; D014170:Transformation, Genetic; D014466:Ultraviolet Rays
D001426:Bacterial Proteins; D004277:DNA, Single-Stranded; D029968:Escherichia coli Proteins; D003851:Deoxyribonucleases; D005090:Exodeoxyribonucleases; D005095:Exoribonucleases; C489467:RecJ protein, E coli; C070765:recJ protein, Bacteria; C028450:exodeoxyribonuclease I; C026870:exodeoxyribonuclease VII; C042081:exoribonuclease T