Liver cell-specific transcriptional regulation of connexin32.
Gap junctional intercellular communication facilitates liver homeostasis and growth control in the liver. The major gap junction protein expressed by hepatocytes is connexin32 (Cx32) and non-parenchymal hepatic cells do not express this gene. We investigated the regulation of Cx32 transcription by trans-activating factors in liver cells. Transient transfection assays using deletions of the rat Cx32 promoter (nt -753 to -33) linked to the luciferase gene were performed in MH1C1 rat hepatoma cells that express endogenous Cx32 compared with WB-F344 rat liver epithelial cells that do not. The basal promoter element was located within nt -134 to -33 and was 1.4-fold more active in MH1C1 cells than WB-F344 cells whereas the entire promoter fragment (nt -754 to -33) was four-fold more active in MH1C1 cells. Specific nuclear protein-DNA complexes that bound to Sp1 consensus sites within the basal promoter were formed using nuclear extracts from both types of cells. Additional promoter sequences increased promoter activity more strongly in MH1C1 cells than WB-F344 cells and this was correlated with the binding of hepatocyte nuclear factor-1 (HNF-1) to two HNF-1 consensus sites centered at -187 and -736. Expression of HNF-1 and binding to these elements was only observed with MH1C1 cells. Other specific protein-DNA complexes were formed, however, that included YY-1- and NF-1-containing complexes, but these were not related to promoter activity. Dexamethasone increased Cx32 promoter activity and expression in MH1C1 cells, but had little effect in WB-F344 cells and did not alter protein-DNA complex formation. These data suggest that Sp1 is responsible for Cx32 promoter basal activity, that HNF-1 determines the cell-specific expression of Cx32, and that dexamethasone increases Cx32 expression through other mechanisms.
Piechocki
M P
MP
Department of Pathology, Medical College of Ohio, 3055 Arlington Avenue, Toledo, OH, USA.
Toti
R M
RM
Fernstrom
M J
MJ
Burk
R D
RD
Ruch
R J
RJ
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D018031:Connexin 43 / Q000235:genetics / Q000378:metabolism; D017630:Connexins / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins; D003907:Dexamethasone / Q000494:pharmacology; D005786:Gene Expression Regulation; D051537:Hepatocyte Nuclear Factor 1; D051538:Hepatocyte Nuclear Factor 1-alpha; D051539:Hepatocyte Nuclear Factor 1-beta; D008099:Liver / Q000378:metabolism*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000378:metabolism; D011401:Promoter Regions, Genetic; D051381:Rats; D017384:Sequence Deletion; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D000097002:Gap Junction beta-1 Protein
D018031:Connexin 43; D017630:Connexins; D004268:DNA-Binding Proteins; D003907:Dexamethasone; D051537:Hepatocyte Nuclear Factor 1; D051538:Hepatocyte Nuclear Factor 1-alpha; D051539:Hepatocyte Nuclear Factor 1-beta; D009687:Nuclear Proteins; D014157:Transcription Factors; D000097002:Gap Junction beta-1 Protein; C496489:Hnf1a protein, rat