DNA nicks inflicted by restriction endonucleases are repaired by a RecA- and RecB-dependent pathway in Escherichia coli

Joseph Heitman, Tatyana Ivanenko, A. Kiss

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

Two mutants of the EcoRI endonuclease (R200K and E144C) predominantly nick only one strand of the DNA substrate. Temperature sensitivity of the mutant enzymes allowed us to study the consequences of inflicting DNA nicks at EcoRI sites in vivo. Expression of the EcoRI endonuclease mutants in the absence of the EcoRI methyltransferase induces the SOS DNA repair response and greatly reduces viability of recA56, recB21 and lexA3 mutant strains of Escherichia coli. In parallel studies, overexpression of the EcoRV endonuclease in cells also expressing the EcoRV methyltransferase was used to introduce nicks at non-cognate EcoRV sites in the bacterial genome. EcoRV overproduction was lethal in recA56 and recB21 mutant strains and moderately toxic in a lexA3 mutant strain. The toxic effect of EcoRV overproduction could be partially alleviated by introduction into the cells of multiple copies of the E. coli DNA ligase gene. These observations suggest that an increased number of DNA nicks can overwhelm the repair capacity of DNA ligase, resulting in the conversion of a proportion of DNA nicks into DNA lesions that require recombination for repair.

Original languageEnglish
Pages (from-to)1141-1151
Number of pages11
JournalMolecular Microbiology
Volume33
Issue number6
DOIs
Publication statusPublished - 1999

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Single-Stranded DNA Breaks
DNA Restriction Enzymes
Deoxyribonuclease EcoRI
DNA Ligases
Poisons
Methyltransferases
Escherichia coli
Bacterial Genomes
Recombinational DNA Repair
DNA
DNA Repair
Temperature
Enzymes
Genes

ASJC Scopus subject areas

  • Molecular Biology
  • Microbiology

Cite this

DNA nicks inflicted by restriction endonucleases are repaired by a RecA- and RecB-dependent pathway in Escherichia coli. / Heitman, Joseph; Ivanenko, Tatyana; Kiss, A.

In: Molecular Microbiology, Vol. 33, No. 6, 1999, p. 1141-1151.

Research output: Contribution to journalArticle

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