Eukaryotic transcription is regulated by repressors as well as activators. For example, geneticists have identified mutations in yeast that result in continuously high expression of certain genes. This type of unregulated, abnormally high expression, called constitutive expression, results from the inactivation of a repressor that normally inhibits the transcription of these genes. Similarly, mutants of Drosophila melanogaster and Caenorhabditis elegans have been isolated that are defective in embryonic development because they express genes in embryonic cells where those genes are normally repressed. The mutations in these mutants inactivate repressors, leading to abnormal development.

Repressor-binding sites in DNA have been identified by systematic linker scanning mutation analyses similar to the one depicted in Figure 9-22. In this type of analysis, whereas mutation of an activator-binding site leads to decreased expression of the linked reporter gene, mutation of a repressor-binding site leads to increased expression of a reporter gene. The repressor proteins that bind such sites can be purified and assayed using the same biochemical techniques described earlier for activator proteins.

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Eukaryotic transcription repressors are the functional converse of activators. They can inhibit transcription of a gene they do not normally regulate when their cognate binding sites are placed within tens of base pairs to many kilobases of the gene’s transcription start site. Like activators, most eukaryotic repressors are modular proteins that have two functional domains: a DNA-binding domain and a repression domain. Like activation domains, repression domains continue to function when fused to another type of DNA-binding domain. If binding sites for this second DNA-binding domain are inserted within a few hundred base pairs of a promoter, expression of the fusion protein inhibits transcription from the promoter. Also like activation domains, repression domains function by interacting with other proteins, as discussed later in this chapter.