In vivo requirement for RecJ, ExoVII, ExoI, and ExoX in methyl-directed mismatch repair

Biochemical studies with model DNA heteroduplexes have implicated RecJ exonuclease, exonuclease VII, exonuclease I, and exonuclease X in Escherichia coli methyl-directed mismatch correction. However, strains deficient in the four exonucleases display only a modest increase in mutation rate, raising questions concerning involvement of these activities in mismatch repair in vivo. The quadruple mutant deficient in the four exonucleases, as well as the triple mutant deficient in RecJ exonuclease, exonuclease VII, and exonuclease I, grow poorly in the presence of the base analogue 2-aminopurine, and exposure to the base analogue results in filament formation, indicative of induction of SOS DNA damage response. The growth defect and filamentation phenotypes associated with 2-aminopurine exposure are effectively suppressed by null mutations in mutH, mutL, mutS, or uvrD/mutU, which encode activities that act upstream of the four exonucleases in the mechanism for the methyl-directed reaction that has been proposed based on in vitro studies. The quadruple exonuclease mutant is also cold-sensitive, having a severe growth defect at 30°C. This phenotype is suppressed by a uvrD/mutU defect, and partially suppressed by mutH, mutL, or mutS mutations. These observations confirm involvement of the four exonucleases in methyl-directed mismatch repair in vivo and suggest that the low mutability of exonuclease-deficient strains is a consequence of under recovery of mutants due to a reduction in viability and/or chromosome loss associated with activation of the mismatch repair system in the absence of RecJ exonuclease, exonuclease VII, exonuclease I, and exonuclease X.

Microsatellite instability in childhood rhabdomyosarcoma is locus specific and correlates with fractional allelic loss.

Replication errors (RERs) were initially identified in hereditary nonpolyposis colon cancer and other tumors of Lynch syndrome II. Mutations in genes involved in mismatch repair give rise to a mutator phenotype, resulting in RERs. The mutator phenotype is thought to predispose to malignant transformation. Here we show that in the embryonal form of childhood rhabdomyosarcoma, RERs also occur, but in contrast to hereditary nonpolyposis colon cancer, only a subset of the microsatellite loci analyzed show RERs. The occurrence of RERs is strongly correlated with increased fractional allelic loss (P < 0.001), suggesting that the occurrence of RERs is a secondary phenomenon in rhabdomyosarcoma. Coincidental loss of genes involved in mismatch repair, possibly due to their proximity to tumor suppressor genes involved in tumor progression of embryonal form of childhood rhabdomyosarcoma, could explain the observed phenomenon.