Involvement of negative cofactor NC2 in active repression by zinc finger-homeodomain transcription factor AREB6.
The transcription factor AREB6 contains a homeodomain flanked by two clusters of Krüppel type C2H2 zinc fingers. AREB6 binds to the E-box consensus sequence, CACCTGT, through either the N- or the C-terminal zinc finger cluster. To gain insights into the molecular mechanism by which AREB6 activates and represses gene expression, we analyzed the domain structure of AREB6 in the context of a heterologous DNA-binding domain by transient-transfection assays. The C-terminal region spanning amino acids 1011 to 1124 was identified as a conventional acidic activation domain. The region containing amino acids 754 to 901, which was identified as a repression domain, consists of 40% hydrophobic amino acids displaying no sequence similarities to other known repression domains. This region repressed transcription in vitro in a HeLa nuclear extract but not in reconstituted transcription systems consisting of transcription factor IID (TFIID), TFIIB, TFIIE, TFIIH/F, and RNA polymerase II. The addition of recombinant negative cofactor NC2 (NC2alpha/DRAP1 and NC2beta/Dr1) to the reconstituted transcription system restored the activity of the AREB6 repression domain. We further demonstrated interactions between the AREB6 repression domain and NC2alpha in yeast two-hybrid assay. Our findings suggest a mechanism of transcriptional repression that is mediated by the general cofactor NC2.
Ikeda
K
K
Department of Biology, Jichi Medical School, Kawachi, Tochigi, Japan.
Halle
J P
JP
Stelzer
G
G
Meisterernst
M
M
Kawakami
K
K
D000818:Animals; D002460:Cell Line; D005786:Gene Expression Regulation; D006367:HeLa Cells; D018398:Homeodomain Proteins / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D051379:Mice; D010750:Phosphoproteins / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids; D011485:Protein Binding; D012097:Repressor Proteins; D014157:Transcription Factors / Q000235:genetics* / Q000378:metabolism; D014158:Transcription, Genetic; D000071799:Zinc Finger E-box-Binding Homeobox 1; D016335:Zinc Fingers / Q000235:genetics
D018398:Homeodomain Proteins; D010750:Phosphoproteins; D012097:Repressor Proteins; D014157:Transcription Factors; C486307:ZEB1 protein, human; D000071799:Zinc Finger E-box-Binding Homeobox 1; C075995:down-regulator of transcription 1