Role of Negative Regulation in Promoter Specificity of the Homologous Transcriptional Activators Ace2p and Swi5p

Abstract

The Ace2p and Swi5p zinc finger proteins have nearly identical DNA-binding domains, yet in vivo they activate transcription of different genes, CTS1 and HO. We now demonstrate that Ace2p and Swi5p recognize sites in the CTS1 and HO promoters with the same affinities, raising the question of how promoter specificity is achieved by these proteins with similar DNA-binding domains. It has been previously shown that Swi5p binds to the HO promoter cooperatively with the Pho2p (Base2p/Grf10p) homeodomain protein, and we now show that Ace2p does not interact with Pho2p. Analysis of CTS1 promoter fragments inserted into a heterologous promoter identify a sequence 90 bp away from the Ace2p binding sites which is required to prevent activation by Swi5p through these binding sites. These results suggest that a regulatory protein bound to the CTS1 promoter is needed to prevent Swi5p from activating CT1S expression. A genetic screen was conducted to identify suppressor mutations which allow CTS1 expression in the absence of the Ace2p activator. The nce3 mutation suppresses the ace2 defect in CTS1 expression only if the strain contains a functional SWI5 gene, suggesting that NCE3 normally functions to prevent Swi5p from activating CTS1. The role of negative regulators such as NCE3, as well as the previously described SIN5 gene, in determining the promoter specificity of homologous activators is discussed.

Stochastic expression and epigenetic memory at the yeast HO promoter

“…Swi5 and Ace2 bind to the same DNA motif in vitro, but they target different genes in vivo (16,17). Only Swi5 effectively binds to and activates HO.…”

Ace2p Controls the Expression of Genes Required for Cell Separation inSchizosaccharomyces pombe

“…Analysis of the promoter region of the seven target genes indicated the existence of two or three copies of the hexanucleotide CCAGCC separated by 100 -200 nucleotides in the promoter of each gene. This sequence has been described as the consensus binding site for S. cerevisiae Ace2p and is necessary for correct transcriptional regulation of the target genes (Dohrmann et al, 1992(Dohrmann et al, , 1996. Based on the sequence similarity of the S. cerevisiae and S. pombe Ace2p factors, especially in the zinc-finger region (43.2% identity in 148 aa region), it is very likely that these regions would also be important for the induction of expression mediated by Ace2p in fission yeast.…”

Cdk10, a Cdc2-related kinase, associates with the Ets2 transcription factor and modulates its transactivation activity

“…Blue/white assays for yeast colony color were performed using the chromogenic substrate X-Gal on Whatman ®lters (Dohrmann et al, 1996). Extracts were prepared and quantitative assays for b-galactosidase activity were performed on three independent transformants using the chromogenic reagent ONPG as described (Breeden and Nasmyth, 1987).…”