Expansion of DNA repeats in Escherichia coli: effects of recombination and replication functions.
Duplication or expansion of directly repeated sequence elements is associated with a number of human genetic diseases. To study the mechanisms of repeat expansion, we have developed a plasmid assay in Escherichia coli. Our assay involves two simple repeats of 787 bp in length; expansion to three or more copies of the repeat can be selected by restoration of an intact tetracycline-resistance gene. Expansions occurred at relatively high rates, >10(-5), in the population. Both RecA-dependent recombination and RecA-independent slipped misalignments contributed to the observed expansion events. Mutations that impair DNA polymerase III (DnaE, DnaQ subunits) or the replication fork helicase, DnaB, stimulated both RecA-dependent and RecA-independent expansion events. In these respects, the properties of repeat expansion resemble repeat deletion and suggest that difficulties in DNA replication may trigger both classes of rearrangements. About 20% of the RecA-independent expansion events are accompanied by reciprocal sister-chromosome exchange, producing dimeric plasmids carrying one triplicated and one deleted locus. These products are explained by a model involving misaligned strands across the replication fork. This model predicts that the location of a replication stall site may govern the types of resulting rearrangements. The specific location of such a stall site can also, in theory, account for propensity towards expansion or deletion of repeat arrays. This may have relevance to trinucleotide repeat expansion in human genetic disease.
Morag
A S
AS
Department of Biology and Rosenstiel Basic Medical Sciences Research Center, Brandeis University, Waltham, MA, 02454-9110, USA.
Saveson
C J
CJ
Lovett
S T
ST
D004261:DNA Replication; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D030342:Genetic Diseases, Inborn / Q000235:genetics; D006801:Humans; D008957:Models, Genetic; D010957:Plasmids; D011938:Rec A Recombinases / Q000378:metabolism; D011995:Recombination, Genetic; D012091:Repetitive Sequences, Nucleic Acid; D017384:Sequence Deletion; D013753:Tetracycline Resistance / Q000235:genetics*
D004269:DNA, Bacterial; D011938:Rec A Recombinases