Figure 4

Maintenance of the nucleosome-depleted PHO5 promoter does not require persistent transcription. (a) Schematic of constructs used for removal of the analyses below. (b) JKT0010 (WT), ROY008 (spt6), JQS002 (WT TATA mutant), and ROY009 (spt6 TATA mutant) previously growing in phosphate containing media (-Pi) were subject to growth in the absence of phosphate (-Pi) to stimulate removal of nucleosomes from the PHO5 promoter, followed by a temperature shift to 39°C to inactivate Spt6, followed by addition of phosphate to stimulate removal of the activators from the promoter. Samples at the indicated times after phosphate addition, or in the indicated media were taken for phosphatase activity assays. (c) Samples of strains JKT0010 (WT), ROY008 (spt6), JQS002 (WT TATA mutant), and ROY009 (spt6 TATA mutant) taken from the same time course in (b) were assessed for histone occupancy over the TATA box of the PHO5 promoter. Data were normalized to the GAL1 promoter region as a control region, and to the input samples. The average and standard deviation of three independent experiments are plotted. (d) Samples of strains JKT0010 (WT), ROY008 (spt6), JQS002 (WT TATA mutant), and ROY009 (spt6 TATA mutant) taken from the same time course in (b) were assessed for histone occupancy at the UASp2 binding site of the PHO5 promoter. Data were normalized to the GAL1 promoter region as a control region, and to the input samples. The average and standard deviation of three independent experiments are plotted.