Cell Cycle-Dependent Binding of Yeast Heat Shock Factor to Nucleosomes

Abstract

In the nucleus, transcription factors must contend with the presence of chromatin in order to gain access to their cognate regulatory sequences. As most nuclear DNA is assembled into nucleosomes, activators must either invade a stable, preassembled nucleosome or preempt the formation of nucleosomes on newly replicated DNA, which is transiently free of histones. We have investigated the mechanism by which heat shock factor (HSF) binds to target nucleosomal heat shock elements (HSEs), using as our model a dinucleosomal heat shock promoter (hsp82-⌬HSE1). We find that activated HSF cannot bind a stable, sequence-positioned nucleosome in G 1 -arrested cells. It can do so readily, however, following release from G 1 arrest or after the imposition of either an early S-or late G 2 -phase arrest. Surprisingly, despite the S-phase requirement, HSF nucleosomal binding activity is restored in the absence of hsp82 replication. These results contrast with the prevailing paradigm for activator-nucleosome interactions and implicate a nonreplicative, S-phase-specific event as a prerequisite for HSF binding to nucleosomal sites in vivo.In the eukaryotic cell, nuclear DNA is packaged into a compact structure known as chromatin, a complex of DNA and histone proteins. The packaging of DNA into chromatin not only serves to confine the genome within the boundaries of the nucleus but also regulates the transcriptional activation of genes. The presence of nucleosomes, the individual subunits of chromatin, can inhibit the binding of sequence-specific activators to upstream elements, as well as impede access of the general transcription machinery to the core promoter (reviewed in reference 35). As a consequence, nucleosomes inhibit transcription, both in vitro (34,43,72) and in vivo (27,29,61). Thus, an important function of activators is to overcome nucleosomal repression, either by blocking nucleosome formation over promoters, thereby presetting genes for activation, or by remodeling preassembled nucleosomes as the initial step in transcriptional activation.The mechanisms by which activators recognize and bind their cognate sites within chromatin are varied. Certain factors, like NF-1 and GCN4, can access their sites only within nuclease-hypersensitive, nucleosome-rearranged regions (5,15,66,75). Others, such as PHO4, bind to accessible regions but, once DNA bound, actively remodel neighboring nucleosomes (1,19). Still others are capable of invading a stable nucleosome and binding to target sites wrapped around the nucleosome core. Glucocorticoid receptor, for example, invades a sequence-positioned nucleosome within the mouse mammary tumor virus promoter following exposure to hormone (5). Receptor binding to its cognate sequence leads to a dramatic reconfiguring of the underlying nucleosome (66) and occurs in the absence of replication (55). The ability of activators to invade a preassembled nucleosome may require the involvement of ATP-dependent chromatin remodeling complexes and/or histone acetyltransferases (reviewed in references 36...

Role of Mediator in Regulating Pol II Elongation and Nucleosome Displacement in Saccharomyces cerevisiae

“…As shown in Figure 2A, the spontaneous ewe mutations have a modest effect on basal HSP82 transcription, and may even decrease it. This contrasts with the stimulatory effect of these mutations on the hsp82-DHSE1/lacZ reporter, whose promoter is assembled into a repressive dinucleosome Venturi et al 2000) and whose coding region is of bacterial origin. Therefore, the basal expression state of hsp82-DHSE1/lacZ may be sensitized to recessive mutations in genes encoding specific subunits of Mediator, possibly as a consequence of the reporter's unique chromatin structure (see Discussion).…”

“…It is possible that enhanced basal expression arises from some attribute of the chromatin structure of the fusion gene, whose promoter is assembled into a repressive dinucleosome Venturi et al 2000) and whose coding region is derived from a heterologous, prokaryotic gene. Others have observed that in the context of Mediator mutations (either med16D alone or in a med5D nut2-1 double mutant), basal expression of HO/lacZ and PHO5/lacZ fusion genes is enhanced, while expression of the intact HO and PHO5 genes remains unaffected (Tabtiang and Herskowitz 1998).…”

A Functional Module of Yeast Mediator That Governs the Dynamic Range of Heat-Shock Gene Expression

“…Deletion of HSE1 also obviates formation of the DNase I hypersensitive (DH), nucleosome-free region characteristic of the wild-type promoter . In place of the DH site-which, despite its accessibility, is occupied by histones under noninducing conditions (Zhao et al 2005)-are two stably positioned nucleosomes, one centered over the mutated UAS and the other centered over the core promoter (Nuc-2 and Nuc-1, respectively) Venturi et al 2000). Accompanying this structural transformation is a 100-fold drop in noninduced transcription .…”

The tumor suppressor p53 associates with gene coding regions and co-traverses with elongating RNA polymerase II in an in vivo model

“…We also constructed a p53-responsive reporter gene by chromosomally integrating two high-affinity PREs in place of the natural heat-shock-responsive enhancer of HSP82. The HSP82 promoter region has an intrinsically high affinity for histones Venturi et al, 2000;Zhao et al, 2005) and in the absence of ectopic p53, the PREs are assembled into a stably positioned nucleosome (schematically depicted in Figure 1b) and transcription of p53-hsp82 is virtually undetectable (Supplementary Figure 1).…”