Transcriptional regulation in the galactose regulon of yeast is determined by an interplay between a positive regulatory protein, GAL4, and a negative regulatory protein, GAL80. We show that derivatives of GAL4 missing as few as 28 carboxy-terminal amino acids are not responsive to GAL80 regulation, implying that the carboxyl terminus of GAL4 is required for interaction with GAL80. Furthermore, a lesion in GAL4 that genetically defines the GAL80-interactive region maps to the 3' end of the gene. Since the carboxyl terminus of GAL4 has also been implicated in interaction with a transcriptional factor, we propose a model for negative regulation in which GAL80 and this factor compete for binding a common region of GAL4, and the on/off state of transcription is determined by the relative affinities of the negative regulator and the transcriptional factor for this region.