FACT and the proteasome promote promoter chromatin disassembly and transcriptional initiation.

The packaging of the eukaryotic genome into chromatin represses gene expression by blocking access of the general transcription machinery to the underlying DNA sequences. Accordingly, eukaryotes have developed a variety of mechanisms to disrupt, alter, or disassemble nucleosomes from promoter regions and open reading frames to allow transcription to occur. Although we know that chromatin disassembly from the yeast PHO5 promoter is triggered by the Pho4 activator, the mechanism is far from clear. Here we show that the Pho4 activator can occupy its nucleosome-bound DNA binding site within the PHO5 promoter. In contrast to the role of Saccharomyces cerevisiae FACT (facilitates chromatin transcription) complex in assembling chromatin within open reading frames, we find that FACT is involved in the disassembly of histones H2A/H2B from the PHO5 promoter during transcriptional induction. We have also discovered that the proteasome is required for efficient chromatin disassembly and transcriptional induction from the PHO5 promoter. Mutants of the degradation function of the proteasome have a defect in recruitment of the Pho4 activator, whereas mutants of the ATPase cap of the proteasome do recruit Pho4 but are still delayed for chromatin assembly. Finally, we rule out the possibility that the proteasome or ATPase cap is driving chromatin disassembly via a potential ATP-dependent chromatin remodeling activity.

Acetylation in the globular core of histone H3 on lysine-56 promotes chromatin disassembly during transcriptional activation.

Promoter chromatin disassembly is a widely used mechanism to regulate eukaryotic transcriptional induction. Delaying histone H3/H4 removal from the yeast PHO5 promoter also leads to delayed removal of histones H2A/H2B, suggesting a constant equilibrium of assembly and disassembly of H2A/H2B, whereas H3/H4 disassembly is the highly regulated step. Toward understanding how H3/H4 disassembly is regulated, we observe a drastic increase in the levels of histone H3 acetylated on lysine-56 (K56ac) during promoter chromatin disassembly. Indeed, promoter chromatin disassembly is driven by Rtt109 and Asf1-dependent acetylation of H3 K56. Conversely, promoter chromatin reassembly during transcriptional repression is accompanied by decreased levels of histone H3 acetylated on lysine-56, and a mutation that prevents K56 acetylation increases the rate of transcriptional repression. As such, H3 K56 acetylation drives chromatin toward the disassembled state during transcriptional activation, whereas loss of H3 K56 acetylation drives the chromatin toward the assembled state.

Chromatin disassembly from the PHO5 promoter is essential for the recruitment of the general transcription machinery and coactivators.

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.

Transcriptional regulation by chromatin disassembly and reassembly.

The packaging of the eukaryotic genome into chromatin severely restricts the access of the transcriptional machinery to the DNA. Recent studies reveal that histones are removed and replaced to enable or restrict, respectively, access of the transcription machinery to regulate transcription. Chromatin disassembly at promoters enables transcriptional activation, whereas promoter chromatin reassembly represses transcription. Histone loss also occurs within transcription units to enable passage of the RNA polymerase, but in this case the histones are rapidly replaced, sometimes by 'variant' histones with specific properties that might serve as a memory of transcriptional competence. Furthermore, the ultimate goal of some epigenetic modifications might well turn out to be the regulation of histone occupancy on the DNA.