Replication restart: a pathway for (CTG).(CAG) repeat deletion in Escherichia coli.
(CTG)n.(CAG)n repeats undergo deletion at a high rate in plasmids in Escherichia coli in a process that involves RecA and RecB. In addition, DNA replication fork progression can be blocked during synthesis of (CTG)n.(CAG)n repeats. Replication forks stalled at (CTG)n.(CAG)n repeats may be rescued by replication restart that involves recombination as well as enzymes involved in replication and DNA repair, and this process may be responsible for the high rate of repeat deletion in E. coli. To test this hypothesis (CAG)n.(CTG)n deletion rates were measured in several E. coli strains carrying mutations involved in replication restart. (CAG)n.(CTG)n deletion rates were decreased, relative to the rates in wild type cells, in strains containing mutations in priA, recG, ruvAB, and recO. Mutations in priB and priC resulted in small reductions in deletion rates. In a recF strain, rates were decreased when (CAG)n comprised the leading template strand, but rates were increased when (CTG)n comprised the leading template. Deletion rates were increased slightly in a recJ strain. The mutational spectra for most mutant strains were altered relative to those in parental strains. In addition, purified PriA and RecG proteins showed unexpected binding to single-stranded, duplex, and forked DNAs containing (CAG)n and/or (CTG)n loop-outs in various positions. The results presented are consistent with an interpretation that the high rates of trinucleotide repeat instability observed in E. coli result from the attempted restart of replication forks stalled at (CAG)n.(CTG)n repeats.
Laboratory of DNA Structure and Mutagenesis, Center for Genome Research, Institute of Biosciences and Technology, Texas A&M University System Health Science Center, 2121 West Holcombe Blvd., Houston, TX 77030-3303, USA.
Seung-Hwan
SH
Kim
Małgorzata J
MJ
Pytlos
Richard R
RR
Sinden
D000251:Adenosine Triphosphatases / Q000378:metabolism; D001483:Base Sequence; D004265:DNA Helicases / Q000378:metabolism; D004261:DNA Replication / Q000235:genetics*; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000378:metabolism; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D017384:Sequence Deletion / Q000235:genetics*; D013379:Substrate Specificity; D019680:Trinucleotide Repeat Expansion / Q000235:genetics*
D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C081006:RecO protein, E coli; C489340:recF protein, E coli; C071446:RecG protein, E coli; D000251:Adenosine Triphosphatases; C506743:priA protein, E coli; D004265:DNA Helicases