Abstract
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.