(A) Cell cycle analysis of α factor arrested
MATa cells treated with splitomicin.
Logarithmically growing MATa cells were treated
first with α factor for 90 min. At time 0, splitomicin (20 μM) or
DMSO was added to the culture. DNA content of the cells was determined
by flow cytometry at several time points after the addition of
splitomicin. (B) α2 mRNA expression from the silent HML
locus in G1-arrested cells treated with
splitomicin. A strain containing the galactose-inducible
CLN3 gene in which genomic G1 cyclin
genes were deleted [MATa cln1Δ,
cln2Δ, cln3Δ, GAL-CLN3 (35)], was
arrested in G1 by exposure to glucose for 90 min.
Splitomicin (20 μM) or DMSO was added to the culture of these
G1-arrested cells, and the expression of MATα
from the silent HML locus was assessed at several time points. Both
splitomicin- and DMSO-treated cells remained arrested in
G1 as judged by flow cytometry (data not shown).
The RNA from MATα and MATa
sir2Δ cells is included for comparison. The weak lower
molecular weight band is caused by cross hybridization to a2
mRNA. The blot was stripped and reprobed for the PDA1 mRNA
as a loading control.