Amino Termini of Histones H3 and H4 Are Required for a1-α2 Repression in Yeast

Abstract

The Saccharomyces cerevisiae ␣2 repressor controls two classes of cell-type-specific genes in yeast through association with different partners. ␣2-Mcm1 complexes repress a cell-specific gene expression in haploid ␣ cells and diploid a/␣ cells, while a1-␣2 complexes repress haploid-specific genes in diploid cells. In both cases, repression is mediated through Ssn6-Tup1 corepressor complexes that are recruited via direct interactions with ␣2. We have previously shown that nucleosomes are positioned adjacent to the ␣2-Mcm1 operator under conditions of repression and that Tup1 interacts directly with histones H3 and H4. Here, we examine the role of chromatin in a1-␣2 repression to determine if chromatin is a general feature of repression by Ssn6-Tup1. We find that mutations in the amino terminus of histone H4 cause a 4-to 11-fold derepression of a reporter gene under a1-␣2 control, while truncation of the H3 amino terminus has a more modest (3-fold or less) effect. Strikingly, combination of the H3 truncation with an H4 mutation causes a 40-fold decrease in repression, clearly indicating a central role for these histones in a1-␣2-mediated repression. However, in contrast to the ordered positioning of nucleosomes adjacent to the ␣2-Mcm1 operator, nucleosomes are not positioned adjacent to the a1-␣2 operator in diploid cells. Our data indicate that chromatin is important to Ssn6-Tup1-mediated repression but that the degrees of chromatin organization directed by these proteins differ at different promoters.Saccharomyces cerevisiae can exist as two haploid cell types, a or ␣, which can mate to form a third cell type, the a/␣ diploid cell (11). Cell type is largely controlled by the ␣2 repressor protein encoded by the MAT␣ locus. In ␣ cells and a/␣ cells, dimers of ␣2 interact with dimers of the Mcm1 protein and these complexes bind a unique sequence upstream of the a cell-specific genes, thereby repressing their transcription (13,16). In diploid cells, ␣2 forms heterodimers with the a1 protein (4) and this complex binds to a distinct sequence upstream of the haploid-specific genes to confer repression (7). In haploid a cells, ␣2 is not expressed, and so neither the a cell-specific nor the haploid-specific genes are repressed.Although ␣2 is directed to different DNA sequences in different stoichiometries through association with either Mcm1 or a1, in both cases it recruits the Ssn6-Tup1 corepressor complex (17). Neither Ssn6 nor Tup1 binds directly to DNA, and these proteins are drawn to specific promoters through interactions with DNA binding factors such as ␣2 (18, 33). The need for the DNA binding factor can be bypassed by fusion of either SSN6 or TUP1 to a heterologous, LexA DNA binding domain, provided the target of repression contains a LexA binding site (17, 36). Interestingly, Ssn6-LexA repression requires the presence of TUP1 (17), but Tup1-LexA can confer repression in the absence of SSN6 (36). These findings suggest that Tup1 is the dominant repressor moiety in the Ssn6-Tup1 complex.In addition to the a cell-...

Redundant Mechanisms Are Used by Ssn6-Tup1 in Repressing Chromosomal Gene Transcription in Saccharomyces cerevisiae

“…We demonstrate that under this condition (⌬isw2) Tup1 represses transcription and TBP recruitment even though nucleosome positioning is disrupted over the promoter. Similar observations were made at haploid-specific genes and ANB1 (11,16), but it was unclear from those studies how Tup1 represses transcription in the absence of nucleosome positioning. The mutational analysis presented here argues that Tup1 continues to repress transcription in a disrupted nucleosomal context by recruiting HDACs to maintain low levels of histone acetylation and by interfering with Mediator function.…”

“…3, A and B, upper panels). In parallel, the ⌬rpd3 mutation was also examined and showed no further derepression in combination with ⌬med3, ⌬srb10, or 14) and MMS-treated cells (lanes [15][16][17][18][19][20][21][22][23][24][25][26][27][28] are presented in the upper and lower panels, respectively. B, Northern blot, as in A except double mutations in Mediator components and ⌬isw2 were analyzed.…”

“…Nucleosomes are well positioned in an Ssn6-Tup1-dependent manner at many repressed loci (7-14). Furthermore, the repression domain of Tup1 can directly bind hypoacetylated histone H3 and H4 tails (15), and H3 and H4 tails are required for repression of Tup1-dependent genes (15,16). At the mating type-specific gene STE6, Tup1 was shown to cross-link throughout the entire gene, and two Tup1 molecules per nucleosome were incorporated into the repressive chromatin structure of a minichromosome (17).…”

Cell-type-dependent repression of yeast a-specific genes requires Itc1p, a subunit of the Isw2p–Itc1p chromatin remodelling complex

“…This organized chromatin, that is disrupted in MATa cells, is implicated in the mechanism of repression of a-specific genes. In contrast, no nucleosome positioning is needed for repression of haploid-specific genes in diploid cells (Huang et al, 1997). Therefore, the fact that absence of Itc1p affects the a-specific but not the haploid-specific gene expression, points to a role of the Itc1p-Isw2p complex in Fig.…”

Tup1-Ssn6 Interacts with Multiple Class I Histone Deacetylases in Vivo

“…Certain repressed genes under Tup1-Ssn6 control are packaged into highly positioned nucleosomes during repression (6 -9). Tup1 binds preferentially to underacetylated H3 and H4 amino-terminal histone tails in vitro, and combined mutation of the H3 and H4 tails leads to a large derepression of Tup1-Ssn6-regulated genes in vivo (10,11). Chromatin immunoprecipitation experiments indicate that Tup1 binding in vivo is associated with decreased acetylation of H3 and H4 (12)(13)(14).…”