The mating-type switching of the
fission yeast, S. pombe, is highly regulated. Two consecutive
asymmetric divisions are required to produce one mating-type switched
cell among the four progeny. It is assumed that the switching pattern
is controlled by a site and strand-specific break (SSB) at the mat1
locus, preparing the switching event during the next DNA
replication. By tracking with a polymerase chain reaction assay, we
identified a novel DNA intermediate of mating-type switching. The time
and rate of appearance and disappearance of this DNA intermediate
reveals that the mating-type switching process occurs during the
S-phase. High-resolution mapping by fluorescence in situ
hybridization on combed DNA was used in order to observed the physical
folding of the mating-type region, bridging together the active locus
with one of the opposite silent cassette during S-phase. Using DNA
density-gradient centrifugation we demonstrate that one fourth
of the mat1 DNA is not replicated by the conventional
semi-conservative mode, but instead both DNA strands are
synthesized de novo. We further demonstrate that the virgin
switched mat1 containing chromatid no longer contained the SSB, while
it is reintroduced during the lagging strand synthesis of the mat1
locus on the sister chromatid. This finding reveals the molecular
basis of the gene conversion event responsible of the asymmetric
mating-type switching during mitotic divisions in this organism.
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