Transcription factor eUSF is an essential component of isolated transcription complexes on the duck histone H5 gene and it mediates the interaction of TFIID with a TATA-deficient promoter.
We analysed the formation of transcription complexes on the H5 gene of the duck which is efficiently transcribed in HeLa cell extracts in vitro. Upon deletion of its TATA-box, the fidelity of transcription of the H5 gene is maintained, although the efficiency of this process is significantly reduced. Selective inactivation of TFIID in whole cell extracts and reconstitution experiments either with human recombinant TFIID or a protein fraction from duck erythrocytes enriched in TFIID show that transcription of the TATA-less H5 promoter nevertheless requires the protein TFIID. Screening of promoter elements which could indirectly mediate the interaction of TFIID with a TATA-less H5 promoter led to the identification of a sequence element located about 40 base-pairs downstream from the H5 initiation site that shows partial homology to the USF consensus sequence. In electrophoretic mobility shift and footprinting studies we demonstrated a specific interaction of the erythroid factor USF (eUSF) with this downstream element. By isolating active transcription complexes we found that all components required for correct initiation remain stably associated with the H5 promoter irrespective of the presence or absence of the TATA box. Moreover, the reconstitution of eUSF and TFIID-depleted transcription complexes with purified protein fractions demonstrate that not only TFIID but also eUSF essentially participates in complex formation even on H5 promoter mutations lacking the TATA-box. Mutual interactions between eUSF and TFIID appear to stabilize the binding of TFIID in the presence or absence of its proper binding site.
Bungert
J
J
Institut für Molekularbiologie und Tumorforschung, Marburg/Lahnberge, Germany.
Kober
I
I
Düring
F
F
Seifart
K H
KH
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D004268:DNA-Binding Proteins / Q000502:physiology*; D004372:Ducks; D005786:Gene Expression Regulation; D006657:Histones / Q000235:genetics*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012045:Regulatory Sequences, Nucleic Acid; D035362:Transcription Factor TFIID; D014157:Transcription Factors / Q000378:metabolism* / Q000502:physiology*; D014158:Transcription, Genetic; D051720:Upstream Stimulatory Factors
D004268:DNA-Binding Proteins; D006657:Histones; D046911:Macromolecular Substances; D035362:Transcription Factor TFIID; D014157:Transcription Factors; D051720:Upstream Stimulatory Factors