Adenosine triphosphate-dependent chromatin remodeling machines play a central role in gene regulation by manipulating chromatin structure. Most genes have a nucleosome-depleted region at the promoter and an array of regularly spaced nucleosomes phased relative to the transcription start site. In vitro, the three known yeast nucleosome spacing enzymes (CHD1, ISW1 and ISW2) form arrays with different spacing. We used genome-wide nucleosome sequencing to determine whether these enzymes space nucleosomes differently in vivo. We find that CHD1 and ISW1 compete to set the spacing on most genes, such that CHD1 dominates genes with shorter spacing and ISW1 dominates genes with longer spacing. In contrast, ISW2 plays a minor role, limited to transcriptionally inactive genes. Heavily transcribed genes show weak phasing and extreme spacing, either very short or very long, and are depleted of linker histone (H1). Genes with longer spacing are enriched in H1, which directs chromatin folding. We propose that CHD1 directs short spacing, resulting in eviction of H1 and chromatin unfolding, whereas ISW1 directs longer spacing, allowing H1 to bind and condense the chromatin. Thus, competition between the two remodelers to set the spacing on each gene may result in a highly dynamic chromatin structure.

The yeast SPT10 gene encodes a putative histone acetyltransferase (HAT) implicated as a global transcription regulator acting through basal promoters. Here we address the mechanism of this global regulation. Although microarray analysis confirmed that Spt10p is a global regulator, Spt10p was not detected at any of the most strongly affected genes in vivo. In contrast, the presence of Spt10p at the core histone gene promoters in vivo was confirmed. Since Spt10p activates the core histone genes, a shortage of histones could occur in spt10⌬ cells, resulting in defective chromatin structure and a consequent activation of basal promoters. Consistent with this hypothesis, the spt10⌬ phenotype can be rescued by extra copies of the histone genes and chromatin is poorly assembled in spt10⌬ cells, as shown by irregular nucleosome spacing and reduced negative supercoiling of the endogenous 2m plasmid. Chromatin structure plays an essential role in gene regulation. The structural unit of chromatin is the nucleosome, which is composed of 147 bp of DNA wrapped in a negative superhelix around a central octamer of core histones (composed of two molecules each of H2A, H2B, H3, and H4) (25). Nucleosomes are separated by linker DNA, forming a "beads on a string" structure. A fifth histone, H1, binds to both the nucleosome and the linker DNA to drive the coiling of the nucleosomal filament to form the 30-nm fiber.The nucleosome presents a problem for regulatory proteins seeking access to DNA because so much of the DNA is protected by histones: the inner surface of the DNA is completely occluded by the central core, the external surface is at least partly protected by the core histone tail domains, and the DNA coils are so close together that their apposed surfaces are also unavailable. To cope with the intrinsically repressive nature of the nucleosome structure, regulatory proteins recruit two types of chromatin remodeling complex to promoters: (i) chromatin remodeling machines, which use ATP to move nucleosomes, effect nucleosomal conformational changes, and exchange core histones with variants (33); and (ii) chromatin modifying enzymes, which catalyze posttranslational modifications of the histones, mostly in their tail domains. These modifications are proposed to represent a "histone code" which is read by regulatory proteins that recognize particular combinations of modifications, resulting in activation or silencing of chromatin (41). Histone acetylation is generally associated with gene activation and is catalyzed by histone acetyltransferases (HATs). The identification of the Gcn5p coactivator as a HAT led to a breakthrough in the field, connecting transcription factors with chromatin (3). The current paradigm is that histone modifying complexes are cofactors recruited to promoters by sequencespecific activators or repressors.Our studies have focused on the CUP1 gene of Saccharomyces cerevisiae as a model for the role of chromatin in gene regulation (35)(36)(37). CUP1 encodes a metallothionein responsible for protecting cell...

We discuss the regulation of the histone genes of the budding yeast Saccharomyces cerevisiae. These include genes encoding the major core histones (H3, H4, H2A, and H2B), histone H1 (HHO1), H2AZ (HTZ1), and centromeric H3 (CSE4). Histone production is regulated during the cell cycle because the cell must replicate both its DNA during S phase and its chromatin. Consequently, the histone genes are activated in late G1 to provide sufficient core histones to assemble the replicated genome into chromatin. The major core histone genes are subject to both positive and negative regulation. The primary control system is positive, mediated by the histone gene-specific transcription activator, Spt10, through the histone upstream activating sequences (UAS) elements, with help from the major G1/S-phase activators, SBF (Swi4 cell cycle box binding factor) and perhaps MBF (MluI cell cycle box binding factor). Spt10 binds specifically to the histone UAS elements and contains a putative histone acetyltransferase domain. The negative system involves negative regulatory elements in the histone promoters, the RSC chromatin-remodeling complex, various histone chaperones [the histone regulatory (HIR) complex, Asf1, and Rtt106], and putative sequence-specific factors. The SWI/SNF chromatin-remodeling complex links the positive and negative systems. We propose that the negative system is a damping system that modulates the amount of transcription activated by Spt10 and SBF. We hypothesize that the negative system mediates negative feedback on the histone genes by histone proteins through the level of saturation of histone chaperones with histone. Thus, the negative system could communicate the degree of nucleosome assembly during DNA replication and the need to shut down the activating system under replication-stress conditions. We also discuss post-transcriptional regulation and dosage compensation of the histone genes.

Most yeast genes have a nucleosome-depleted region (NDR) at the promoter and an array of regularly spaced nucleosomes phased relative to the transcription start site. We have examined the interplay between RSC (a conserved essential SWI/SNFtype complex that determines NDR size) and the ISW1, CHD1, and ISW2 nucleosome spacing enzymes in chromatin organization and transcription, using isogenic strains lacking all combinations of these enzymes. The contributions of these remodelers to chromatin organization are largely combinatorial, distinct, and nonredundant, supporting a model in which the +1 nucleosome is positioned by RSC and then used as a reference nucleosome by the spacing enzymes. Defective chromatin organization correlates with altered RNA polymerase II (Pol II) distribution. RSC-depleted cells exhibit low levels of elongating Pol II and high levels of terminating Pol II, consistent with defects in both termination and initiation, suggesting that RSC facilitates both. Cells lacking both ISW1 and CHD1 show the opposite Pol II distribution, suggesting elongation and termination defects. These cells have extremely disrupted chromatin, with high levels of closely packed dinucleosomes involving the second (+2) nucleosome. We propose that ISW1 and CHD1 facilitate Pol II elongation by separating closely packed nucleosomes.