Two opposing effects of mismatch repair on CTG repeat instability in Escherichia coli

The expansion of normally polymorphic CTG microsatellites in certain human genes has been identified as the causative mutation of a number of heredita ry neurological disorders, including Huntington's disease and myotonic dyst rophy, Here, we have investigated the effect of methyl-directed mismatch re pair (MMR) on the stability of a (CTG)(43) repeat in Escherichia coli over 140 generations and find two opposing effects. In contrast to orientation-d ependent repeat instability in wild-type E. coli and yeast, we observed no orientation dependence in MMR- E. coli cells and suggest that, for the repe at that we have studied, orientation dependence in wild-type cells is mainl y caused by functional mismatch repair genes. Our results imply that slippe d structures are generated during replication, causing single triplet expan sions and contractions in MMR- cells, because they are left unrepaired. On the other hand, we find that the repair of such slipped structures by the M MR system can go awry, resulting in large contractions. We show that these mutS-dependent contractions arise preferentially when the CTG sequence is e ncoded by the lagging strand. The nature of this orientation dependence arg ues that the small slipped structures that are recognized by the MMR system are formed primarily on the lagging strand of the replication fork. It als o suggests that, in the presence of functional MMR, removal of 3 bp slipped structures causes the formation of larger contractions that are probably t he result of secondary structure formation by the CTG sequence. We rational ize the opposing effects of MMR on repeat tract stability with a model that accounts for CTG repeat instability and loss of orientation dependence in MMR- cells. Our work resolves a contradiction between opposing claims in th e literature of both stabilizing and destabilizing effects of MMR on CTG re peat instability in E. coli.