Identification of functional regulatory regions of the connexin32 gene promoter.
Connexin32 (Cx32) is the predominant gap junction protein expressed in adult rat hepatocytes. This study investigated transcriptional regulation of the rat Cx32 gene in MH(1)C(1) rat hepatoma cells using transient expression assays in conjunction with promoter mutagenesis and 5' nested deletion analysis. Site-directed mutagenesis of the -736 and -187 hepatocyte nuclear factor-1 (HNF-1) sites, the -196 and -116 Sp1 sites, and the -729 and -329 Yin Yang 1 (YY1) sites all significantly reduced promoter activity. We have defined the contribution of each individual site to promoter activity in the intact cell. A novel upstream region of the Cx32 promoter (-1042 to -758) was cloned and shown to contain negative regulatory elements. The transcription factors HNF-1 and Sp1 have important functional roles in the transcriptional regulation of basal and cell-specific Cx32 expression. The multifunctional transcription factor YY1 is also implicated.
Field
Joanne M L
JM
School of Biosciences, The University of Birmingham, Edgbaston, B15 2TT, Birmingham, UK.
Tate
Laura A
LA
Chipman
J Kevin
JK
Minchin
Stephen D
SD
D000818:Animals; D001665:Binding Sites; D017630:Connexins / Q000235:genetics* / Q000502:physiology*; D004268:DNA-Binding Proteins; D017353:Gene Deletion; D005786:Gene Expression Regulation; D051537:Hepatocyte Nuclear Factor 1; D051538:Hepatocyte Nuclear Factor 1-alpha; D051539:Hepatocyte Nuclear Factor 1-beta; D022781:Hepatocytes / Q000378:metabolism; D008156:Luciferases / Q000378:metabolism; D008957:Models, Genetic; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009687:Nuclear Proteins; D010957:Plasmids / Q000378:metabolism; D011401:Promoter Regions, Genetic; D051381:Rats; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016329:Sp1 Transcription Factor / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured; D000097002:Gap Junction beta-1 Protein
D017630:Connexins; D004268:DNA-Binding Proteins; D051537:Hepatocyte Nuclear Factor 1; D051538:Hepatocyte Nuclear Factor 1-alpha; D051539:Hepatocyte Nuclear Factor 1-beta; D008156:Luciferases; D009687:Nuclear Proteins; D016329:Sp1 Transcription Factor; D014157:Transcription Factors; D000097002:Gap Junction beta-1 Protein; C496489:Hnf1a protein, rat