Chromatin, TAFs, and a novel multiprotein coactivator are required for synergistic activation by Sp1 and SREBP-1a in vitro.
The promoter selectivity factor Sp1 often cooperates with other enhancer-binding proteins to activate transcription. To study the molecular underpinnings of these regulatory events, we have reconstituted in vitro the synergy observed in vivo between Sp1 and the sterol-regulated factor SREBP-1a at the low density lipoprotein receptor (LDLR) promoter. Using a highly purified human transcription system, we found that chromatin, TAFs, and a novel SREBP-binding coactivator activity, which includes CBP, are all required to mediate full synergistic activation by Sp1 and SREBP-1a. The SREBP-binding domain of CBP inhibits activation by SREBP-1a and Sp1 in a dominant-negative fashion that is both chromatin- and activator-specific. Whereas recombinant CBP alone is not sufficient to mediate activation, a human cellular fraction containing CBP can support high levels of chromatin-dependent synergistic activation. Purification of this activity to near homogeneity resulted in the identification of a multiprotein coactivator, including CBP, that selectively binds to the SREBP-1a activation domain and is capable of mediating high levels of synergistic activation by SREBP/Sp1 on chromatin templates. The development of a reconstituted chromatin transcription system has allowed us to isolate a novel coactivator that is recruited by the SREBP-1a activation domain and that functions in concert with TFIID to coordinate the action of multiple activators at complex promoters in the context of chromatin.
Howard Hughes Medical Institute, Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA.
A M
AM
Näär
P A
PA
Beaurang
K M
KM
Robinson
J D
JD
Oliner
D
D
Avizonis
S
S
Scheek
J
J
Zwicker
J T
JT
Kadonaga
R
R
Tjian
D000818:Animals; D022762:CCAAT-Enhancer-Binding Proteins; D002843:Chromatin / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000502:physiology*; D004330:Drosophila; D004357:Drug Synergism; D006367:HeLa Cells; D018257:Helix-Loop-Helix Motifs / Q000502:physiology*; D006801:Humans; D016350:Leucine Zippers / Q000502:physiology*; D008957:Models, Genetic; D009687:Nuclear Proteins / Q000502:physiology*; D011401:Promoter Regions, Genetic; D011973:Receptors, LDL / Q000235:genetics; D016329:Sp1 Transcription Factor / Q000502:physiology*; D051780:Sterol Regulatory Element Binding Protein 1; D035362:Transcription Factor TFIID; D014157:Transcription Factors; D019473:Transcription Factors, TFII / Q000502:physiology; D015533:Transcriptional Activation
D022762:CCAAT-Enhancer-Binding Proteins; D002843:Chromatin; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D011973:Receptors, LDL; C497387:SREBF1 protein, human; D016329:Sp1 Transcription Factor; D051780:Sterol Regulatory Element Binding Protein 1; D035362:Transcription Factor TFIID; D014157:Transcription Factors; D019473:Transcription Factors, TFII