Silencers, Sir proteins, and the cell cycle.
Scott G.
Holmes, Mirela Matecic, Mary Lou Dula, Adrienne Woike
Molecular
Biology, Wesleyan University, Lawn Avenue, Middletown, CT 06762, USA
Silencing at the HM loci and telomeres in yeast exhibits
epigenetic inheritance, in which the repressed state is stably
propagated through DNA replication and mitosis. To examine this
epigenetic mechanism we have conducted two types of experiments. First,
we have created temperature sensitive (ts) alleles of the SIR2
gene and used them to determine the requirement for Sir2p at specific
points in the cell cycle. These alleles establish two distinct silencing
functions of Sir2p: when shifted to the non-permissive temperature, one
class of ts alleles shows a loss of silencing at all phases of the cell
cycle, while a second class defines a function that is dispensable at G1
phase, but is essential to maintain silencing through mitosis. We are
currently correlating these in vivo defined phenotypes to biochemical
defects caused by the mutations. In vivo deletion of the silencer
sequences at HML in G1 phase does not cause a loss of silencing,
but progression through a single cell cycle in the absence of silencers
disrupts silencing. We are determining the cell cycle intervals that
cause loss of silencing following in vivo deletion of the silencer
sequences. Thus far our experiments suggest that silencers are not
required to propagate the silenced state through S-phase, but are
required in the G2/M to G1 interval. Our results point to a crucial
event in the silencing mechanism that is coincident with mitosis.
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