Combinatorial Regulation of theSaccharomyces cerevisiae CAR1(Arginase) Promoter in Response to Multiple Environmental Signals†

Abstract

CAR1 (arginase) gene expression responds to multiple environmental signals; expression is induced in response to the intracellular accumulation of arginine and repressed when readily transported and catabolized nitrogen sources are available in the environment. Up to 14 cis-acting sites and 9 trans-acting factors have been implicated in regulated CAR1 transcription. In all but one case, the sites are redundant. To test whether these sites actually participate in CAR1 expression, each class of sites was inactivated by substitution mutations that retained the native spacing of the CAR1 cis-acting elements. Three types of sites function independently of the nitrogen source: two clusters of Abf1p-and Rap1p-binding sites, and a GC-rich sequence. Two different sets of nitrogen source-dependent sites are also required: the first consists of two GATAA-containing UAS NTR sites that mediate nitrogen catabolite repression-sensitive transcription, and the second is arginine dependent and consists of three UAS I elements that activate transcription only when arginine is present. A single URS1 site mediates repression of CAR1 arginine-independent upstream activator site (UAS) activity in the absence of arginine and the presence of a poor nitrogen source (a condition under which the inducer-independent Gln3p can function in association with the UAS NTR sites). When arginine is present, the combined activity of the UAS elements overcomes the negative effects mediated by URS1. Mutation of the classes of sites either singly or in combination markedly alters CAR1 promoter operation and control, supporting the idea that they function synergistically to regulate expression of the gene.One of the more complex and well-studied nitrogen catabolic genes in Saccharomyces cerevisiae is CAR1, which encodes arginase

Nitrogen Starvation and TorC1 Inhibition Differentially Affect Nuclear Localization of the Gln3 and Gat1 Transcription Factors Through the Rare Glutamine tRNACUG in Saccharomyces cerevisiae

“…To assess the effects of the sup70-65 mutation on the five identifiable modes of nitrogen-responsive regulation, we chose Gln3-GFP localization as the reporter because (1) it is the most comprehensively studied reporter across the entire spectrum of catabolic nitrogen conditions and (2) Gln3-GFP localization is a more specific probe of nitrogenresponsive regulation than NCR-sensitive gene expression in that it avoids the complication that nitrogen-responsive mRNA levels derive from the cumulative actions of multiple transcription factors whose actions are not coordinately regulated (Messenguy et al 1991(Messenguy et al , 2000Kovari et al 1993a, b;Smart et al 1996;Dubois and Messenguy 1997;Park et al 1999;van der Merwe et al 2001;Rai et al 2004).…”

Genome-wide coexpression dynamics: Theory and application

“…Intuitively we may expect a positive correlation between the two profiles, but this is not the case; very little similarity exists between the two expression profiles. Perhaps this merely reflects the complexity due to many cis elements and trans-acting factors for CAR1 (14), the localization of Gln3p, and its interactions with other factors (15). To apply the LA methodology, we take (GLN3,CAR1) as the pair (X,Y) in Fig.…”

Regulation of Amino Acid, Nucleotide, and Phosphate Metabolism in Saccharomyces cerevisiae

“…P CAR1 contains up to 14 different cis-acting promoter elements, of which at least 11 are functionally active and contribute to regulate CAR1 expression (Smart et al 1996;Dubois and Messenguy 1997). Only three of these promoter elements will be discussed, i.e., the repressing URS1 motif, the UAS i required for arginine induction, and the UAS NTR required for NCR-controlled transcription.…”

“…Upon nitrogen starvation, the repression at URS1 is released and Ume6 interacts with the ArgR/Mcm1 complex, presumably enhancing the binding of this complex to the three UAS i motifs in a manner that facilitates induced expression. To achieve full derepression of CAR1, Gln3 and Gat1 binding to GATA sequences in the UAS NTR elements is required (Smart et al 1996). Finally, CAR1 expression is nonspecifically induced by the addition of micromolar amounts of the amino acids (Dubois and Wiame 1976;Godard et al 2007).…”