Reassessment of the in vivo functions of DNA polymerase I and RNase H in bacterial cell growth.
A major factor in removing RNA primers during the processing of Okazaki fragments is DNA polymerase I (Pol I). Pol I is thought to remove the RNA primers and to fill the resulting gaps simultaneously. RNase H, encoded by rnh genes, is another factor in removing the RNA primers, and there is disagreement with respect to the essentiality of both the polA and rnh genes. In a previous study, we looked for the synthetic lethality of paralogs in Bacillus subtilis and detected several essential doublet paralogs, including the polA ypcP pair. YpcP consists of only the 5'-3' exonuclease domain. In the current study, we first confirmed that the polA genes of both Escherichia coli and B. subtilis could be completely deleted. We found that the 5'-3' exonuclease activity encoded by either polA or ypcP xni was required for the growth of B. subtilis and E. coli. Also, the 5'-3' exonuclease activity of Pol I was indispensable in the cyanobacterium Synechococcus elongatus. These results suggest that a 5'-3' exonuclease activity is essential in these organisms. Our success in constructing a B. subtilis strain that lacked all RNase H genes indicates that the enzymatic activity is dispensable, at least in the wild type. Increasing the 5'-3' exonuclease activity partially compensated for a defective phenotype of an RNase H-deficient mutant, suggesting cooperative functions for the two enzyme systems. Our search for the distribution of the 5'-3' exonuclease domain among 250 bacterial genomes resulted in the finding that all eubacteria, but not archaea, possess this domain.
Fukushima
Sanae
S
Department of Bioscience, Tokyo University of Agriculture, Sakuragaoka 1-1-1 Setagaya-ku, Tokyo 156-8502, Japan.
Itaya
Mitsuhiro
M
Kato
Hiroaki
H
Ogasawara
Naotake
N
Yoshikawa
Hirofumi
H
D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000166:cytology / Q000201:enzymology* / Q000254:growth & development*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D004256:DNA Polymerase I / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000166:cytology / Q000201:enzymology* / Q000254:growth & development; D017353:Gene Deletion; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D016914:Ribonuclease H / Q000235:genetics / Q000378:metabolism*; D046940:Synechococcus / Q000201:enzymology
D001426:Bacterial Proteins; D004256:DNA Polymerase I; D016914:Ribonuclease H