Activation of amylin gene transcription by LIM domain homeobox gene isl-1.
The amylin (IAPP) and insulin genes are coexpressed in the pancreatic beta-cell and share related promoter elements that may bind similar islet transcription factors. The observation that these promoter elements contain AT-rich subdomains suggests that homeobox proteins may be important for the regulation of both insulin and amylin gene transcription. We show here that the LIM domain homeobox protein isl-1 activates the rat amylin promoter in both fibroblast and islet cell lines. Mutation of the rAMY promoter TAAT motifs was associated with a marked reduction in both basal and isl-1 -dependent transcriptional activity. The isl-1 homeodomain binds to the AT-rich AMY element (-156 to -137) in the human amylin (hAMY) gene promoter, and electrophoretic mobility shift assay experiments using isl-1 specific antiserum detected the formation of an hAMY-isl-1 complex using nuclear extract from InR1 -G9 islet cells. Although isl-1 binds to both the insulin and amylin gene promoter elements in vitro, these sequences display marked differences in their relative transcriptional properties when ligated adjacent to a heterologous promoter and transfected into InR1 -G9 islet cells. The insulin gene E2 sequence that binds isl-1 (-230 to -208) functions as a negative element, whereas the hAMY sequence activates the thymidine kinase promoter in islet, but not nonislet, cell lines. Transfection of isl-1-depleted isl-1 (AS)InR1 -G9 cell lines demonstrated that the E2 element continued to repress thymidine kinase promoter activity, whereas the positive transcriptional activity mediated by the AMY element was considerably reduced in isl-1 (AS)-InR1-G9 cell lines. These dat2 demonstrate that highly similar elements in islet hormone gene promoters display differential functional properties and support a role for the isl-1 homeodomain protein in the regulation of amylin, but not insulin, gene transcription.
Wang
M
M
Departments of Medicine, Banting and Best Diabetes Centre, The Toronto Hospital, Ontario, Canada.
Drucker
D J
DJ
D000682:Amyloid / Q000235:genetics*; D000818:Animals; D001665:Binding Sites; D002460:Cell Line; D006224:Cricetinae; D016373:DNA, Antisense / Q000235:genetics; D004274:DNA, Recombinant / Q000235:genetics; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects*; D005813:Genes, Synthetic; D018398:Homeodomain Proteins / Q000494:pharmacology*; D006801:Humans; D007328:Insulin / Q000235:genetics; D058228:Islet Amyloid Polypeptide; D007515:Islets of Langerhans / Q000187:drug effects / Q000378:metabolism; D060850:LIM-Homeodomain Proteins; D008647:Mesocricetus; D009419:Nerve Tissue Proteins; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis* / Q000235:genetics; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics; D012045:Regulatory Sequences, Nucleic Acid; D014157:Transcription Factors; D014158:Transcription, Genetic / Q000187:drug effects*
D000682:Amyloid; D016373:DNA, Antisense; D004274:DNA, Recombinant; D018398:Homeodomain Proteins; D007328:Insulin; D058228:Islet Amyloid Polypeptide; D060850:LIM-Homeodomain Proteins; D009419:Nerve Tissue Proteins; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C063843:insulin gene enhancer binding protein Isl-1