Mg. Hanford et al., MICROSATELLITE MUTATION-RATES IN CANCER CELL-LINES DEFICIENT OR PROFICIENT IN MISMATCH REPAIR, Oncogene, 16(18), 1998, pp. 2389-2393
A selectable system has been used to determine mutation rates within a
microsatellite sequence in human cancer cell lines with or without de
fects in mismatch repair. A sequence consisting of 17 repeats of poly
(dC-dA).poly(dT-dG) [abbreviated as (Ca)(17)] was inserted near the 5'
end of the bacterial neomycin-resistance gene in a plasmid vector, su
ch that the reading frame of the neo gene is disrupted. This plasmid w
as introduced into cancer cell lines, where it became integrated into
the cellular genome. Clones with insertions or deletions of CA-repeats
that restored the normal reading frame of the neo gene were selected
in G418, and mutation rates were determined by fluctuation analysis. T
he rates of reversion in LoVo cells, which are deficient for hMSH2, we
re about one in a thousand per generation, which is approximately two
orders of magnitude higher than in the repair-proficient HT-1080 human
fibrosarcoma cell line. The mutation rates in H6 cells, which are der
ived from the hMLH1-deficient HCT116 line, were more heterogeneous tha
n in LoVo, but all were considerably higher than in the repair-profici
ent line. Nearly all of the revertants of the repair-deficient lines h
ad deletions of a single CA-repeat from the microsatellite sequence, w
hereas repair-proficient cells had a broader spectrum of mutations.