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Quantitative evaluation of recA gene expression in Escherichia coli

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Summary

A recA::lac operon fusion was constructed using the phage Mu d(Ap, lac) in Escherichia coli to obtain precise measurements of the level of recA gene expression in various genetic backgrounds. The RecA protein normally represents 0.02% of total protein. This value is known to increase dramatically after treatments interrupting DNA synthesis; kinetic experiments showed that the rate of recA expression increases 17-fold within 10 min after UV irradiation or thymine starvation. In mutants affected in SOS regulation or repair the following observations were made: (i) the tif-1 mutation in the recA gene does not alter the basal level of recA expression, suggesting that it improves the protease activity of RecA; (ii) the lexA3 mutation does not create a “super-repressor” of recA; (iii) the tsl-1 mutation in the lexA gene makes the LexA protein a poor repressor of recA at 30°C (2.5-fold derepression) and a poor substrate for RecA protease (3-fold stimulation of recA expression by UV); (iv) the spr-55 amber mutation in the lexA gene causes a 30-fold increase in recA expression, higher than all inducing treatments, and this level cannot be further increased by nalidixic acid; (v) the zab-53 mutation at the recA locus, known to abolish tsl-mediated induction of recA expression, is trans-recessive and thus probably affects a regulatory site on the DNA; (vi) uvrA, B and C, recB and recF mutations do not increase the basal level of recA expression, suggesting that there are not sufficient spontaneous lesions to cause induction even when any one of these three repair pathways is inoperative.

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Abbreviations

Ap:

ampicillin

Km:

kanamycin

Cm:

chloramphenicol

Tc:

terracycline

Sm:

streptomycin

Ts:

thermosensitive

Tr:

thermoresistant

Nal:

nalidixic acid

X-Gal:

5-bromo-4-chloro-3-indolyl-β-D-galactoside

mito C:

mitomycin C

LFT:

low frequency transducing

HFT:

high frequency transducing

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Author notes

  1. Serge Casaregola

    Present address: Department of Genetics, University of Leicester, Adrian Building, LE1 7RH, Leicester, England

Authors and Affiliations

  1. Institut de Recherche en Biologie Moléculaire, CNRS, Université Paris VII, 2, Place Jussieu, F-75251, Paris Cedex 05, France

    Serge Casaregola, Richard D'Ari & Olivier Huisman

Authors
  1. Serge Casaregola
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  2. Richard D'Ari
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  3. Olivier Huisman
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Communicated by B.A. Bridges

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Casaregola, S., D'Ari, R. & Huisman, O. Quantitative evaluation of recA gene expression in Escherichia coli . Mol Gen Genet 185, 430–439 (1982). https://doi.org/10.1007/BF00334135

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