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MBC in Press, published online ahead of print March 23, 2005
Mol. Biol. Cell 10.1091/mbc.E05-02-0126

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Submitted on February 15, 2005
Accepted on March 7, 2005

Genome-wide Analysis of the Functions of a Conserved Surface on the Corepressor Tup1

Sarah R. Green* and Alexander D. Johnson*{dagger}

Departments of *Biochemistry and Molecular Biology and {dagger}Microbiology and Immunology, University of California-San Francisco, San Francisco, CA 94143

Monitoring Editor: William Tansey

The general transcriptional repressor Tup1 is responsible for the regulation of a large, diverse set of genes in S. cerevisiae, and functional homologues of Tup1 have been identified in many metazoans. The crystal structure for the C-terminal portion of Tup1 has been solved and, when sequences of Tup1 homologues from fungi and metazoans were compared, a highly conserved surface was revealed. In this paper, we analyze five point mutations that lie on this conserved surface. A statistical analysis of expression microarrays demonstrates that the mutant alleles are deficient in the repression of different subsets of Tup1-regulated genes. We were able to rank the mutant alleles of TUP1 based on the severity of their repression defects measured both by the number of genes derepressed and the magnitude of that derepression. For one particular class of genes, the mutations on the conserved surface disrupted recruitment of Tup1 to the repressed promoters. However, for the majority of the genes derepressed by the Tup1 point mutants, recruitment of Tup1 to the regulated promoters is largely unaffected. These mutations affect the mechanism of repression subsequent to recruitment of the complex and likely represent a disruption of a mechanism that is conserved in fungi and metazoans. This work demonstrates that the evolutionarily conserved surface of Tup1 interacts with two separate types of proteins--sequence-specific DNA-binding proteins responsible for recruiting Tup1 to promoters as well as components that are likely to function in a conserved repression mechanism.

Address correspondence to: Alexander D. Johnson (ajohnson{at}cgl.ucsf.edu)




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