Molecular biology of the 230-kD bullous pemphigoid antigen. Cloning of the BPAG1 gene and its tissue-specific expression.
The 230-kD bullous pemphigoid antigen is a hemidesmosomal protein of the cutaneous basement membrane zone. We have previously cloned overlapping cDNAs corresponding to the human 230-kD bullous pemphigoid antigen gene (BPAG1), located at the human chromosomal locus 6p11-12. Utilizing the cDNA clones, a genomic DNA lambda FIX II phage library was screened. Seven over-lapping genomic clones, spanning approximately 20 kb, were isolated. These clones were shown to contain the entire approximately 9-kb coding sequence of BPAG1, and it consisted of 22 separate exons which varied from 78 to 2,810 bp in size. Elucidation of 2.6 kb of 5'-flanking DNA was found to contain several putative transcriptional response elements, and development of promoter chloramphenicol acetyltransferase (CAT) reporter gene constructs allowed identification of putative cis-elements which confer keratinocyte-specific expression to the gene. In particular, a putative AP2-binding sequence (KRE2) in the position -(1,786-1778) was shown to be responsible for marked enhancement of the endogenous promoter, as well as of a heterologous thymidine kinase/CAT construct, activity in normal human keratinocytes. Normal human keratinocyte nuclear extracts contained a protein, designated as KTP1, which complexed with the KRE2 oligomer by gel mobility shift assays. UV cross-linking and Southwestern analysis suggested that KTP1 is a DNA-binding protein clearly distinct from AP2, and this protein may be responsible for the basal keratinocyte-specific expression of the BPAG1 gene.
Tamai
K
K
Department of Dermatology, Jefferson Medical College, Thomas Jefferson University Philadelphia, Pa. 19107.
Sawamura
D
D
Choi Do
H Y
HY
Li
K
K
Uitto
J
J
D001324:Autoantigens / Q000235:genetics*; D002352:Carrier Proteins; D002478:Cells, Cultured; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D003094:Collagen; D003598:Cytoskeletal Proteins; D018076:DNA, Complementary / Q000235:genetics; D000071821:Dystonin; D005091:Exons / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D007438:Introns / Q000235:genetics; D015603:Keratinocytes / Q000276:immunology* / Q000378:metabolism; D008970:Molecular Weight; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D010391:Pemphigoid, Bullous; D011401:Promoter Regions, Genetic; D000097805:Collagen Type XVII
D001324:Autoantigens; D002352:Carrier Proteins; D003094:Collagen; D003598:Cytoskeletal Proteins; D018076:DNA, Complementary; D000071821:Dystonin; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D000097805:Collagen Type XVII; C486268:DST protein, human