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.Asf1 ͉ K56 acetylation ͉ transcription T he packaging of DNA together with histone proteins into chromatin is essential for regulating all of the activities of the eukaryotic genome, including repair, replication, and gene expression. The chromatin structure dynamically changes to facilitate these processes and regulate access to the DNA sequence. For example, nucleosomes are disassembled from many eukaryotic promoters during transcriptional activation to provide access to the general transcription machinery (1). The histone H3/H4 chaperone known as Asf1 contributes to the disassembly of histones H3/H4 from multiple budding yeast promoters during transcriptional activation (2-4).Histone posttranslational modifications may also play an important role in regulating chromatin disassembly. Many of the best studied sites of histone posttranslational modifications map to the N-terminal tails of the histones that extend out from the globular core of the nucleosome and are unlikely to have a direct influence on the structure of the nucleosome. By contrast, the newly identified acetylation of lysine-56 within the globular core of histone H3 (H3 K56ac) (5, 6) is unique because it is predicted to break a DNA:histone interaction, potentially destabilizing the nucleosome. H3 K56 acetylation occurs predominantly on newly synthesized histones that are assembled into chromatin after DNA replication (6) and are rapidly deacetylated after S phase (7,8). Functionally, K56 acetylation promotes survival after the exposure of cells to genotoxic agents because of its role in stabilizing the replisome (6, 9). The histone acetyltransferase (HAT) responsible for acetylation of K56 on newly synthesized histones is Rtt109 (10-12) and needs either of two histone chaperones, Asf1 or Vps75, for enzymatic activity (13).Given the requirement for the histone chaperone Asf1 for H3 K56 acetylation and its role in chromatin disassembly from promoters during transcriptional activation, we investi...

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.

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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.Eukaryotic chromatin is made up of a fundamental repeating unit, termed the nucleosome, which consists of 147 bp of DNA wrapped around the outside of an octamer of histone proteins (1). The histone octamer in turn comprises a heterotetramer of histone proteins H3/H4 and two heterodimers of histones H2A/H2B. In order to allow the transcription machinery to gain access to the DNA, the chromatin structure is altered by the concerted action of three processes (2): (i) post-translational modifications on the histones, (ii) the breakage of histone-DNA contacts by ATP-dependent chromatin remodeling machines, and (iii) the ultimate removal of histones from the DNA by histone chaperones. The sequence-specific transcriptional activators trigger these chromatin alterations occurring at promoters during transcriptional induction, but it is unclear whether transcriptional activators can first bind to their nucleosome-buried DNA recognition sequences to start this cascade of chromatin dynamics. Similarly, once transcription is complete, transcriptional activators leave the DNA, and promoters are repackaged with histones via the process of chromatin assembly.The assembly and disassembly of nucleosomes appear to occur in a stepwise manner (3). This is due to the peripheral positions of the H2A/H2B dimers within the nucleosome (1), necessitating the removal of H2A/H2B prior to removal of the central H3/H4 tetramer. Conversely, H3/H4 must be deposited onto the DNA prior to H2A/H2B in order to assemble a nucleosome. Accordingly, histone chaperones exist that bind to either H2A/H2B or H3/H4 to mediate chromatin assembly and disassembly...

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