Distinct rat proteins can recognize CCAAT-homologous sequences of the metallothionein promoter and trans-activate this promoter.
We have previously purified and characterized a rat liver protein C'BP-1 that is either identical or closely related to C/EBPdelta (Aniskovitch and Jacob, 1997). The mouse metallothionein-I (MT-I) promoter contains two C'BP-1 binding sites, one of which includes the MRE-c' region (-135 to -110). The C'BP-1 binding activity was detected by EMSA as a major activity for MRE-c' in nonproliferating adult liver cells but not in rat hepatoma cells. In this study, we purified and characterized a factor, C'BP-2, which had a dominant binding activity for MRE-c' in Morris hepatoma 3924A, a poorly differentiated, fast-growing tissue. C'BP-2 is a 28 kDa protein which exists in solution as a monomer. As observed for C'BP-1, affinity-purified C'BP-2 stimulated transcription from the mMT-I gene promoter. DNase I footprinting revealed two C'BP-2 binding sites in the regions that overlap with the CCAAT homologies of the C'BP-1 binding sites on the mMT-I promoter. C'BP-2 made essential contacts with the CCAAT homology and in the region upstream of this sequence. Competition electrophoretic mobility shift assay and methylation interference analysis revealed that C'BP-2 is a protein closely related, but not identical, to CP2. These data suggest that C'BP-1 and C'BP-2 may play a role in hepatocyte proliferation and/or differentiation.
Aniskovitch
L P
LP
Department of Pharmacology and Molecular Biology, The Chicago Medical School, North Chicago, Illinois 60064, USA.
Jacob
S T
ST
D000818:Animals; D022762:CCAAT-Enhancer-Binding Proteins; D002848:Chromatography, DEAE-Cellulose; D004268:DNA-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D008114:Liver Neoplasms, Experimental / Q000378:metabolism / Q000473:pathology; D008668:Metallothionein / Q000235:genetics*; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D051381:Rats; D015533:Transcriptional Activation
D022762:CCAAT-Enhancer-Binding Proteins; D004268:DNA-Binding Proteins; D009687:Nuclear Proteins; D008668:Metallothionein