Keratinocyte transglutaminase promoter analysis. Identification of a functional response element.
Keratinocyte transglutaminase catalyzes isopeptide bond formation to yield the highly insoluble cross-linked envelope during terminal differentiation of epidermal cells. Transcriptional response elements were identified in the 5'-flanking DNA of the gene for this enzyme by a combination of transient transfection and electrophoretic mobility shift analyses. Since human keratinocytes transcribed ineffectively transfected transglutaminase flanking DNA, a key feature of these experiments was the use of rat bladder epithelial cells as recipients. Serial deletion experiments identified by transient transfection an important response region containing three putative AP2-like response elements approximately 0.5 kilobases from the transcription initiation site. Oligonucleotides, each containing a single one of the elements, formed specific complexes with keratinocyte nuclear proteins. Two of the response elements were found to be functional by transfection in site-specific deletion experiments. Of these one formed specific DNA-protein complexes with nuclear proteins only from cells exhibiting keratinocyte differentiation. UV cross-linking experiments estimated the protein component of the complex to be approximately 85 kDa. This response element alone increased substantially the transcription of a minimal transglutaminase promoter in transient transfections. Further characterization of the putative transcription factor binding to this response element may provide insight into the regulation of keratinocyte transglutaminase.
Department of Environmental Toxicology, University of California, Davis 95616-8588, USA.
L
L
Mariniello
Q
Q
Qin
B A
BA
Jessen
R H
RH
Rice
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D004591:Electrophoresis, Polyacrylamide Gel; D005347:Fibroblasts / Q000378:metabolism; D006801:Humans; D015603:Keratinocytes / Q000201:enzymology*; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D011401:Promoter Regions, Genetic; D051381:Rats; D014162:Transfection; D011503:Transglutaminases / Q000235:genetics*; D014407:Tumor Cells, Cultured
D009838:Oligodeoxyribonucleotides; D011503:Transglutaminases