<p>(a) <i>YFR057w</i> mRNA levels in <i>SIR<sup>+</sup></i> and <i>sir2Δ</i> cells (BY4741 background) exposed to 200 µg/ml cycloheximide (CX) for the indicated times. <i>YFR057w</i> transcripts were quantified by RT-qPCR, and normalized to those of <i>SCR1</i>. Depicted are means ± S.D. (N = 3; qPCR = 6). (b) ChIP-qPCR analysis of Pol II within the <i>YFR057w</i> promoter and ORF in <i>sir2Δ</i> and <i>SIR<sup>+</sup></i> cells exposed to cycloheximide for the indicated times as in A. Pol II occupancy was determined as in <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1004202#pgen-1004202-g003" target="_blank">Figure 3A</a>. Occupancy of the non-induced promoter (<i>sir2Δ</i>) was set to 1.0; all other occupancies (both promoter and ORF) are scaled relative to it. Depicted are means ± S.D. (N = 4; qPCR = 8). (c) ChIP-qPCR analysis of histone PTMs within the <i>YFR057w</i> promoter (H3K18ac, H4K16ac, H3K4me2,3) or ORF (H3K79me2) in <i>sir2Δ</i> and <i>SIR<sup>+</sup></i> cells exposed for the indicated times to cycloheximide as in A. Shown are normalized PTM/histone H3 quotients. PTM-specific and H3 signals at <i>YFR057w</i> were normalized to those measured at <i>PMA1</i> and <i>ARS504</i>, respectively. The PTM/histone H3 quotient of the non-induced <i>sir2Δ</i> sample was set to 1.0 in each case. Depicted are means ± S.D. (N = 2; qPCR = 4). (d) ChIP-qPCR analysis of H3K36me3 enrichment within the ORF and Htz1 enrichment within the promoter as in C, for the indicated times following addition of cycloheximide (N = 2; qPCR = 4). (e) As in D, except H3K56ac enrichment over <i>YFR057w</i> promoter and ORF was assayed.</p
