FIGURE 6.

plc1Δ cells that display reduced levels of acetyl-CoA and histone hypoacetylation in plc1Δ cells can be suppressed by increasing the pool of cytosolic acetyl-CoA. A, low intracellular level of acetyl-CoA in plc1Δ cells is partially suppressed by reduced ACC1 expression. The indicated strains were grown in YPD medium containing 0.05 μg/ml of doxycycline to an A₆₀₀ = 0.8. The cells were harvested by centrifugation, lysed with glass beads in perchloric acid, and acetyl-CoA was determined in neutralized lysates. The experiment was repeated three times, and the results are shown as mean ± S.D. 100% wild-type levels of acetyl-CoA equals to 1.6 nmol/10⁷ cells. B, histone hypoacetylation in plc1Δ cells is partially suppressed by the tetO₇-ACC1 allele. The indicated strains were grown in YPD medium containing 0.05 μg/ml of doxycycline to an A₆₀₀ = 0.8. Samples were analyzed by Western blotting with antibodies against histone H3 acetylated at lysine 14 (acH3), hyperacetylated histone H4 (acH4), and total histone H3. Even loading of protein samples was confirmed with anti-Pgk1p antibody. The experiment was performed three times, and representative results are shown. C, temperature sensitivity of plc1Δ cells is partially suppressed by reduced ACC1 expression. 10-Fold serial dilutions of the indicated strains were spotted onto YPD plates without doxycycline and YPD plates containing 0.1 μg/ml of doxycycline and grown for 2 days at 30 and 35 °C. Typical results from three independent experiments are shown. D, histone hypoacetylation in the promoter regions of RPL18B and RPS22B of plc1Δ cells is partially suppressed by the tetO₇-ACC1 allele. Occupancies of H3, acH3, and acH4 were determined in the promoter regions of RPL18B and RPS22B and the acetylation per nucleosome was calculated as ratios of acH3 to total H3 and acH4 to total H3. The experiment was repeated three times, and the results are shown as mean ± S.D. E, low expression of ribosomal protein genes in plc1Δ cells is suppressed by the tetO₇-ACC1 allele. Wild-type, plc1Δ, tetO₇-ACC1, and plc1Δ tetO₇-ACC1 cells were grown in YPD medium containing 0.05 μg/ml of doxycycline at 30 °C to an A₆₀₀ = 1.0 and the total RNA was isolated and assayed for RPL18B and RPS22B transcripts by real-time RT-PCR. The results were normalized to ACT1 RNA and expressed relative to the value for the WT strain. The experiment was repeated three times, and the results are shown as mean ± S.D.