Redundancy of Chromatin Remodeling Pathways for the Induction of the Yeast PHO5 Promoter in Vivo

Induction of the yeast PHO5 and PHO8 genes leads to a prominent chromatin transition at their promoter regions as a prerequisite for transcription activation. Although induction of PHO8 is strictly dependent on Snf2 and Gcn5, there is no chromatin remodeler identified so far that would be essential for the opening of PHO5 promoter chromatin. Nonetheless, the nonessential but significant involvement of cofactors can be identified if the chromatin opening kinetics are delayed in the respective mutants. Using this approach, we have tested individually all 15 viable Snf2 type ATPase deletion mutants for their effect on PHO5 promoter induction and opening. Only the absence of Snf2 and Ino80 showed a strong delay in chromatin remodeling kinetics. The snf2 ino80 double mutation had a synthetic kinetic effect but eventually still allowed strong PHO5 induction. The same was true for the snf2 gcn5 and ino80 gcn5 double mutants. This strongly suggests a complex network of redundant and mutually independent parallel pathways that lead to the remodeling of the PHO5 promoter. Further, chromatin remodeling kinetics at a transcriptionally inactive TATA box-mutated PHO5 promoter were affected neither under wild type conditions nor in the absence of Snf2 or Gcn5. This demonstrates the complete independence of promoter chromatin opening from downstream PHO5 transcription processes. Finally, the histone variant Htz1 has no prominent role for the kinetics of PHO5 promoter chromatin remodeling.

Chromatin Disassembly from the PHO5 Promoter Is Essential for the Recruitment of the General Transcription Machinery and Coactivators

ABSTRACT The disassembly of promoter nucleosomes appears to be a general property of highly transcribed eukaryotic genes. We have previously shown that the disassembly of chromatin from the promoters of the Saccharomyces cerevisiae PHO5 and PHO8 genes, mediated by the histone chaperone anti-silencing function 1 (Asf1), is essential for transcriptional activation upon phosphate depletion. This mechanism of transcriptional regulation is shared with the ADY2 and ADH2 genes upon glucose removal. Promoter chromatin disassembly by Asf1 is required for recruitment of TBP and RNA polymerase II, but not the Pho4 and Pho2 activators. Furthermore, accumulation of SWI/SNF and SAGA at the PHO5 promoter requires promoter chromatin disassembly. By contrast, the requirement for SWI/SNF and SAGA to facilitate Pho4 activator recruitment to the nucleosome-buried binding site in the PHO5 promoter occurs prior to chromatin disassembly and is distinct from the stable recruitment of SWI/SNF and SAGA that occurs after chromatin disassembly.