CIF, AN ESSENTIAL COFACTOR FOR TFIID-DEPENDENT INITIATOR FUNCTION

The core promoters for mammalian protein-coding genes often contain a TATA box, an initiator (Inr) element, or both of these control element s. The TFIID complex is essential both for TATA activity and for the a ctivity of a common class of Inr elements characterized by an approxim ate consensus sequence PyPyA(+1)NT/APyPy. Although the complete set of proteins required for basal TATA-mediated transcription has been esta blished, the requirements for TFIID-dependent Inr activity remain unde fined. In this study we set out to reconstitute Inr activity with puri fied and recombinant general transcription factors. for this analysis, Inr activity was measured as the ability of an Inr to enhance the str ength of a core promoter containing an upstream TATA box. Inr activity was not detected in reactions containing TFIIB, RAP30, RAP74, RNA pol ymerase II, and either TBP or TFIID, even though these factors were su fficient for TATA-mediated transcription from supercoiled templates. B y use of a complementation assay, a factor that imparts Inr activity w as identified. This factor, named CIF, stimulated Inr activity in reac tions containing the TFIID complex, but activity was not detected with TBP. Further characterization of CIF suggested that it contains multi ple components. functional and immunological experiments demonstrated that one of the CIF components is the mammalian homolog of Drosophila TAF(II)150, which is not tightly associated with mammalian TFIID. Thes e results reveal significant differences in the factor requirements fo r basal TATA and Inr activity. Further elucidation of these difference s is likely to explain the need for the core promoter heterogeneity fo und within protein-coding genes.