Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 99, Issue 17, Pages 11061-11066Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.172197099
Keywords
SOS mutagenesis; error-prone DNA polymerase; DNA damage-induced mutation
Categories
Funding
- NIA NIH HHS [T32 AG000093, AG00093] Funding Source: Medline
- NIGMS NIH HHS [GM52725, GM21422, R37 GM021422, GM42554, R01 GM021422, R01 GM052725] Funding Source: Medline
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SOS mutagenesis in Escherichia coli requires DNA polymerase V (pol V) and RecA protein to copy damaged DNA templates. Here we show that two distinct biochemical modes for RecA protein are necessary for pol V-catalyzed translesion synthesis. One RecA mode is characterized by a strong stimulation in nucleotide incorporation either directly opposite a lesion or at undamaged template sites, but by the absence of lesion bypass. A separate RecA mode is necessary for translesion synthesis. The RecA1730 mutant protein, which was identified on the basis of its inability to promote pol V (UmuD'C-2)-dependent UV-mutagenesis, appears proficient for the first mode of RecA action but is deficient in the second mode. Data are presented suggesting that the two RecA modes are nonfilamentous. That is, contrary to current models for SOS mutagenesis, formation of a RecA nucleoprotein filament may not be required for copying damaged DNA templates. Instead, SOS mutagenesis occurs when pol V interacts with two RecA molecules, first at a 3' primer end, upstream of a template lesion, where RecA mode 1 stimulates pol V activity, and subsequently at a site immediately downstream of the lesion, where RecA mode 2 cocatalyzes lesion bypass. We posit that in vivo assembly of a RecA nucleoprotein filament may be required principally to target pol V to a site of DNA damage and to stabilize the pol V-RecA interaction at the lesion. However, it is only a RecA molecule located at the 3' filament tip, proximal to a damaged template base, that is directly responsible for translesion synthesis.
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