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.
Näär
A M
AM
Howard Hughes Medical Institute, Department of Molecular and Cell Biology, University of California, Berkeley, California 94720, USA.
Beaurang
P A
PA
Robinson
K M
KM
Oliner
J D
JD
Avizonis
D
D
Scheek
S
S
Zwicker
J
J
Kadonaga
J T
JT
Tjian
R
R
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