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Figure 1 | BMC Molecular Biology

Figure 1

From: Multiple histone modifications in euchromatin promote heterochromatin formation by redundant mechanisms in Saccharomyces cerevisiae

Figure 1

Suppression of the silencing defects of dot1Δ strains. (A) Reporter genes used for telomeric silencing. Cells in which the ADE2 gene is silenced accumulate a red pigment whereas cells that express ADE2 are white. Cells in which URA3 is silenced are resistant to 5-FOA, whereas cells in which URA3 is expressed convert 5-FOA into a toxic product and are sensitive to 5-FOA. (B) Wild-type (WT) and dot1Δ strains were transformed with empty vector (p) or a Sir3 overexpression plasmid (pSir3) and were spotted in 10-fold dilution series on media (YC) with and without 5-FOA. (C) Immunoblot analysis of Sir3 expression in sir3Δ and WT cells, and cells containing the Sir3 overexpression plasmid. Ctrl indicates a non-specific band recognized by the Sir3 antibody that was used as a loading control. (D) Telomeric silencing in WT and dot1Δ strains lacking RIF1 or RPD3; sir2Δ and sir3Δ strains are shown as no-silencing controls (E) mRNA expression levels of ADE2 and URA3 relative to ACT1 were determined by RT-qPCR. mRNA was isolated and quantified in duplicate with the difference as the standard error. (F) Sir3 binding at ADE2-TEL-VR, URA3-TEL-VIIL and 3500 bp from telomere VIR (VIR3500) relative to binding at control locus ACT1 was determined by ChIP combined with real-time qPCR. Each clone was analyzed in duplicate with the difference as the standard error. (G) Silencing in strains lacking DEP1 (Rpd3L complex) or RCO1 (RPD3S complex).

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