GCR1, a transcriptional activator in Saccharomyces cerevisiae, complexes with RAP1 and can function without its DNA binding domain.

Abstract

In Saccharomyces cerevisiae, efficient expression of glycolytic and translational component genes requires two DNA binding proteins, RAP1 (which binds to UASRPG) and GCR1 (which binds to the CT box). We generated deletions in GCR1 to test the validity of several different models for GCR1 function. We report here that the C‐terminal half of GCR1, which includes the domain required for DNA binding to the CT box in vitro, can be removed without affecting GCR1‐dependent transcription of either the glycolytic gene ADH1 or the translational component genes TEF1 and TEF2. We have also identified an activation domain within a segment of the GCR1 protein (the N‐terminal third) that is essential for in vivo function. RAP1 and GCR1 can be co‐immunoprecipitated from whole cell extracts, suggesting that they form a complex in vivo. The data are most consistent with a model in which GCR1 is attracted to DNA through contact with RAP1.

Regulation of Ribosome Biogenesis by the Rapamycin-sensitive TOR-signaling Pathway inSaccharomyces cerevisiae

“…Not depicted in the figure are Bmh1 and Bmh2, the yeast homologues of 14-3-3 proteins, which have been shown recently to be involved in TOR signaling (Bertram et al, 1998). of many r-protein genes, as well as Gcr1, a protein required for the activity of both r-protein genes as well as genes involved in glycolysis (Santangelo and Tornow, 1990;Tornow et al, 1993). In support of possible involvement of Gcr1 in TOR signaling, we have observed that rapamycin also severely inhibits expression of several glycolytic genes, including ADH1, ENO1, and PGK1 (our unpublished results).…”

Reverse recruitment: The Nup84 nuclear pore subcomplex mediates Rap1/Gcr1/Gcr2 transcriptional activation

“…We report here independent approaches demonstrating that the Rap1͞Gcr1͞Gcr2 activation assemblage (7)(8)(9)19), like its silencing counterpart, is anchored at the nuclear periphery. For example, synthetic genetic array (SGA) analysis identified a robust genetic network that connects the Rap1 coactivators Gcr1 and Gcr2 with the Nup84 subcomplex (20,21) and other nuclear envelope components; some of these same perinuclear factors coimmunoprecipitated with Gcr1 and Gcr2.…”

“…Numerous aspects of Rap1 activation have conformed poorly with the ''free diffusion'' aspect of the recruitment model for transcriptional activation. One such aspect is the presence of an unusually large activation domain that is easily inactivated by means of mutations throughout the N-terminal 280 residues of Gcr1, spanning four distinct hypomutable regions (8,17,18); two of these hypomutable regions overlap with putative transmembrane domains.We report here independent approaches demonstrating that the Rap1͞Gcr1͞Gcr2 activation assemblage (7-9, 19), like its silencing counterpart, is anchored at the nuclear periphery. For example, synthetic genetic array (SGA) analysis identified a robust genetic network that connects the Rap1 coactivators Gcr1 and Gcr2 with the Nup84 subcomplex (20, 21) and other nuclear envelope components; some of these same perinuclear factors coimmunoprecipitated with Gcr1 and Gcr2.…”

“…Numerous aspects of Rap1 activation have conformed poorly with the ''free diffusion'' aspect of the recruitment model for transcriptional activation. One such aspect is the presence of an unusually large activation domain that is easily inactivated by means of mutations throughout the N-terminal 280 residues of Gcr1, spanning four distinct hypomutable regions (8,17,18); two of these hypomutable regions overlap with putative transmembrane domains.…”

Rearrangements of the transcriptional regulatory networks of metabolic pathways in fungi

“…S. cerevisiae has a highly specialized strategy of sugar metabolism that is of great interest to humans, as yeast cells efficiently ferment glucose even in the presence of oxygen, sacrificing short-term energy production for rapid exploitation of the carbon source and favoring the production of ethanol as a byproduct [33]. The transcriptional circuit controlling hexose metabolism has been extensively investigated in the brewer's yeast and the key activators of the glycolytic pathway that directs the fermentation of hexose sugars are Gcr1 and Gcr2, which associate with the CT box motif upstream of the glycolytic genes along with the Rap1 general transcription factor [34,35]. As well, S. cerevisiae and its close relatives exhibit the Crabtree effect that results from repressors such as Mig1 and Rgt1 inhibiting the non-fermentative use of glucose [26,36].…”