NOT1(CDC39), NOT2(CDC36), NOT3, AND NOT4 ENCODE A GLOBAL-NEGATIVE REGULATOR OF TRANSCRIPTION THAT DIFFERENTIALLY AFFECTS TATA-ELEMENT UTILIZATION

The yeast HIS3 T-R and T-C TATA elements support basal transcription, but only T-R can respond to transcriptional activators. Four genes, NOT1(CDC39), NOT2(CDC36), NOT3, NOT4, act as general negative regulators and preferentially affect T-C-dependent transcription. Allele-specific suppression, a two-hybrid interaction, and biochemical cofractionation suggest that NOT1 and NOT2 are nuclear proteins associated in a discrete, 500-kD complex. NOT4 interacts with NOT1 and NOT3 in the two-hybrid assay, and overexpression of NOT3 or NOT4 suppresses not1 and not2 mutations. Repression by the NOT proteins is not attributable to inhibition of transcriptional activators, does not involve the CYC8/TUP1 negative regulatory complex, and is distinct from repression by nucleosomes or by the SPT4,5,6 proteins that affect chromatin structure. We propose that the NOT proteins inhibit the basic RNA polymerase II transcription machinery, possibly by affecting TFIID function.