Glucose uptake, the first, rate-limiting step of its utilization, is facilitated 
by glucose transporters. Expression of several glucose transporter (HXT) genes 
in yeast is repressed by the Rgt1 repressor, which recruits the 
glucose-responsive transcription factor Mth1 and the general corepressor complex 
Ssn6-Tup1 in the absence of glucose; however, it is derepressed when Mth1 is 
inactivated by glucose. Here we show that Ssn6-Tup1 interferes with the 
DNA-binding ability of Rgt1 in the absence of Mth1 and that the Rgt1 function 
abrogated by Ssn6 overexpression is restored by co-overexpression of Mth1. Thus 
Mth1 likely regulates Rgt1 function not by modulating its DNA-binding activity 
directly but by functionally antagonizing Ssn6-Tup1. Mth1 does so by acting as a 
scaffold-like protein to recruit Ssn6-Tup1 to Rgt1. Supporting evidence shows 
that Mth1 blocks the protein kinase A-dependent phosphorylation of Rgt1 that 
impairs the ability of Rgt1 to interact with Ssn6-Tup1. Of note, Rgt1 can bind 
DNA in the absence of Ssn6-Tup1 but does not inhibit transcription, suggesting 
that dissociation of Rgt1 from Ssn6-Tup1, but not from DNA, is necessary and 
sufficient for the expression of its target genes. Taken together, these 
findings show that Mth1 is a transcriptional corepressor that facilitates the 
recruitment of Ssn6-Tup1 by Rgt1.