The LIM domain homeobox gene isl-1 is a positive regulator of islet cell-specific proglucagon gene transcription.
The LIM domain homeobox gene islet 1 (isl-1) is expressed in the embryonic nervous system and may be an early marker of motor neuron specification. isl-1 is expressed in all 4 islet cell types, but a role for isl-1 in the regulation of insulin gene expression has not been demonstrated, and the genetic targets for isl-1 in the pancreas remain unknown. We show here that the proximal rat proglucagon gene promoter binds an amino-terminally truncated Trp-E-isl-1 fusion protein that lacks the LIM domains. The proglucagon gene promoter also binds full-length in vitro translated isl-1 containing the intact LIM domains. isl-1 antisera detects binding of proglucagon gene sequences to isl-1 present in a slowly-migrating complex in nuclear extracts from InR1-G9 islet cells. The transcriptional properties of the proglucagon gene promoter sequences that bind isl-1 (designated Ga, Gb, and Gc) were assessed after transfection of reporter genes into wild type and isl-1-antisense (isl-1(AS)) InR1-G9 islet cells. The proximal proglucagon gene (Ga) promoter sequence reduced TK-CAT activity by approximately 50%, but no change in the activity of the Ga-TK-CAT plasmid was seen after transfection of isl-1(AS) InR1-G9 cells. In contrast, the Gb/Gc sites activated transcription 2-3 fold in wild type InR1-G9 cells, and the isl-1-dependent activation of gene transcription through the Gb/Gc element was eliminated following transfection of isl-1(AS) InR1-G9 cells. These data demonstrate that the LIM domain homeobox gene isl-1 1) is not constrained from DNA binding by its LIM domains and 2) functions as a positive regulator of proglucagon gene transcription in the endocrine pancreas.
Wang
M
M
Banting and Best Diabetes Centre, Toronto Hospital, University of Toronto, Canada.
Drucker
D J
DJ
D000818:Animals; D016375:Antisense Elements (Genetics); D001483:Base Sequence; D002467:Cell Nucleus / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D004742:Enhancer Elements, Genetic / Q000235:genetics; D005786:Gene Expression Regulation; D005801:Genes, Homeobox / Q000235:genetics*; D005809:Genes, Regulator / Q000235:genetics; D005934:Glucagon / Q000096:biosynthesis* / Q000235:genetics; D018398:Homeodomain Proteins; D007515:Islets of Langerhans / Q000502:physiology*; D060850:LIM-Homeodomain Proteins; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins; D052336:Proglucagon; D011485:Protein Binding; D011498:Protein Precursors / Q000096:biosynthesis* / Q000235:genetics; D051381:Rats; D011993:Recombinant Fusion Proteins; D013329:Structure-Activity Relationship; D014157:Transcription Factors; D014158:Transcription, Genetic
D016375:Antisense Elements (Genetics); D004268:DNA-Binding Proteins; D018398:Homeodomain Proteins; D060850:LIM-Homeodomain Proteins; D009419:Nerve Tissue Proteins; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; C063843:insulin gene enhancer binding protein Isl-1; D052336:Proglucagon; D005934:Glucagon