Tissue-specific expression of the 230-kDa bullous pemphigoid antigen gene (BPAG1). Identification of a novel keratinocyte regulatory cis-element KRE3.
The 230-kDa bullous pemphigoid antigen gene (BPAG1) is expressed exclusively in basal keratinocytes of epidermis. In this study, we have identified a novel cis-element, keratinocyte responsive element 3 (KRE3), at position -216 to -197 of the human BPAG1 gene. A promoter-CAT construct containing this element had approximately 50-fold higher expression than a similar construct devoid of this sequence when tested in transient transfections of cultured human keratinocytes. However, there was no effect on the low base-line level of expression in cultured skin fibroblasts. KRE3 contains a palindromic sequence 5'-CAAATATTTG-3', and mutations in this sequence significantly reduced the promoter activity. Gel mobility shift assays with an oligomer containing KRE3 sequence demonstrated binding activity with nuclear proteins isolated from keratinocytes. One of the DNA/protein complexes was clearly specific, since competition with > 12.5-fold excess of the unlabeled oligomer resulted in disappearance of this band. No specific binding activity was noted with nuclear proteins extracted from fibroblasts. Thus, KRE3 appears to serve as the binding site for keratinocyte-specific trans-activating factor(s), and KRE3 may thus confer the tissue-specific expression to the BPAG1 gene.
Tamai
K
K
Department of Dermatology, Jefferson Medical College, Thomas Jefferson University, Philadelphia, Pennsylvania 19107, USA.
Silos
S A
SA
Li
K
K
Korkeela
E
E
Ishikawa
H
H
Uitto
J
J
D001324:Autoantigens / Q000096:biosynthesis* / Q000235:genetics; D001483:Base Sequence; D002352:Carrier Proteins; D002478:Cells, Cultured; D015500:Chloramphenicol O-Acetyltransferase / Q000096:biosynthesis; D003094:Collagen / Q000096:biosynthesis / Q000235:genetics; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D015870:Gene Expression; D006801:Humans; D015603:Keratinocytes / Q000378:metabolism*; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D009687:Nuclear Proteins / Q000302:isolation & purification / Q000378:metabolism; D009928:Organ Specificity; D010391:Pemphigoid, Bullous / Q000235:genetics; D011401:Promoter Regions, Genetic; D011994:Recombinant Proteins / Q000096:biosynthesis; D012045:Regulatory Sequences, Nucleic Acid; D012867:Skin / Q000378:metabolism*; D014162:Transfection; D000097805:Collagen Type XVII
D001324:Autoantigens; D002352:Carrier Proteins; D015500:Chloramphenicol O-Acetyltransferase; D003094:Collagen; D003598:Cytoskeletal Proteins; D000071821:Dystonin; D009419:Nerve Tissue Proteins; D024041:Non-Fibrillar Collagens; D009687:Nuclear Proteins; D011994:Recombinant Proteins; D000097805:Collagen Type XVII; C486268:DST protein, human