Direct physical interactions between HNF-4 and Sp1 mediate synergistic transactivation of the apolipoprotein CIII promoter.
We have investigated the mechanism of functional cooperativity between specificity protein 1 (Sp1) and hepatocyte nuclear factor-4 (HNF-4) on the human apolipoprotein CIII (apoCIII) promoter. Cotransfections in Drosophila SL2 cells that lack endogenous Sp1 or Sp1-related activities showed that HNF-4 and Sp1 synergistically transactivate the -890/+24 apoCIII promoter up to 150-fold. Synergistic transactivation required the HNF-4 binding site of the apoCIII enhancer. Deletion of part of the Ser/Thr-rich and Gln-rich domain or the C-terminal domain of Sp1 decreased, and deletion of residues 501-610 of Sp1 increased, the functional cooperativity between Sp1 and HNF-4. Physical interactions between the two factors were demonstrated by glutathione S-transferase pull-down and co-immunoprecipitation assays. The amino terminal domain of both factors and the carboxy terminal domain of Sp1 contribute to these interactions. Antagonism between HNF-4 and Sp1 was demonstrated on homopolymeric promoters containing multiple binding sites for either factor, suggesting that the synergism between the two factors occurs only when both factors are bound simultaneously to the DNA. The observed physical interactions between Sp1 and HNF-4 in the context of the apoCIII promoter may explain in part their in vitro and in vivo synergism in the transcriptional activation of the apolipoprotein A-I/apoCIII/apolipoprotein A-IV gene cluster.
Kardassis
Dimitris
D
Department of Basic Sciences, University of Crete Medical School and Institute of Molecular Biology and Biotechnology, FORTH, Heraklion GR-71110, Crete, Greece.
Falvey
Eileen
E
Tsantili
Panayota
P
Hadzopoulou-Cladaras
Margarita
M
Zannis
Vassilis
V
D000818:Animals; D053305:Apolipoprotein C-III; D001056:Apolipoproteins C / Q000737:chemistry / Q000235:genetics*; D001483:Base Sequence; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D002478:Cells, Cultured; D017931:DNA Primers; D004268:DNA-Binding Proteins; D015536:Down-Regulation; D051557:Hepatocyte Nuclear Factor 4; D006801:Humans; D010750:Phosphoproteins / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D017384:Sequence Deletion; D016329:Sp1 Transcription Factor / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*; D015533:Transcriptional Activation
D053305:Apolipoprotein C-III; D001056:Apolipoproteins C; D051778:Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; D017931:DNA Primers; D004268:DNA-Binding Proteins; D051557:Hepatocyte Nuclear Factor 4; C403128:MLX protein, human; D010750:Phosphoproteins; D016329:Sp1 Transcription Factor; D014157:Transcription Factors