Glucose regulates islet amyloid polypeptide gene transcription in a PDX1- and calcium-dependent manner.
Islet amyloid polypeptide (IAPP) and insulin are expressed in the beta-cells of the islets of Langerhans. They are co-secreted in response to changes in glucose concentration, and their mRNA levels are also regulated by glucose. The promoters of both genes share similar cis-acting sequence elements, and both bind the homeodomain transcription factor PDX1, which plays an important role in the regulation of the insulin promoter and insulin mRNA levels by glucose. Here we examine the role of PDX1 in the regulation of the human IAPP promoter by glucose. The experiments were facilitated by the availability of a human beta-cell line (NES2Y) that lacks PDX1. NES2Y cells also lack operational K(ATP) channels, resulting in a loss of control of calcium signaling. We have previously used these cells to show that glucose regulation of the insulin gene is dependent on PDX1, but not calcium. In the mouse beta-cell line Min6, glucose (16 mm) stimulated a 3.5-4-fold increase in the activity of a -222 to +450 IAPP promoter construct compared with values observed in 0.5 mm glucose. In NES2Y cells, glucose failed to stimulate transcriptional activation of the IAPP promoter. Overexpression of PDX1 in NES2Y cells failed to reinstate glucose-responsive control of the IAPP promoter. Glucose effects on the IAPP promoter were observed only in the presence of PDX1 when normal calcium signaling was restored by overexpression of the two K(ATP) channel subunits SUR1 and Kir6.2. The importance of calcium was further emphasized by an experiment in which glucose-stimulated IAPP promoter activity was inhibited by the calcium channel blocker verapamil (50 microm). Verapamil was further shown to inhibit the stimulatory effect of glucose on IAPP mRNA levels. These results demonstrate that like the insulin promoter, glucose regulation of the IAPP promoter is dependent on the activity of PDX1, but unlike the insulin promoter, it additionally requires the activity of another, as yet uncharacterized factor(s), the activity of which is calcium-dependent.
Macfarlane
W M
WM
Department of Molecular and Cell Biology, University of Aberdeen, Institute of Medical Sciences, Foresterhill, Aberdeen AB25 2ZD, United Kingdom.
Campbell
S C
SC
Elrick
L J
LJ
Oates
V
V
Bermano
G
G
Lindley
K J
KJ
Aynsley-Green
A
A
Dunne
M J
MJ
James
R F
RF
Docherty
K
K
D000682:Amyloid / Q000235:genetics*; D000818:Animals; D002118:Calcium / Q000494:pharmacology / Q000502:physiology*; D002460:Cell Line; D005786:Gene Expression Regulation / Q000187:drug effects; D005947:Glucose / Q000494:pharmacology*; D018398:Homeodomain Proteins / Q000378:metabolism; D006801:Humans; D007328:Insulin / Q000235:genetics; D058228:Islet Amyloid Polypeptide; D007515:Islets of Langerhans / Q000166:cytology / Q000378:metabolism*; D051379:Mice; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D015534:Trans-Activators / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects*; D014162:Transfection; D014700:Verapamil / Q000494:pharmacology
D000682:Amyloid; D018398:Homeodomain Proteins; D007328:Insulin; D058228:Islet Amyloid Polypeptide; D012333:RNA, Messenger; D011994:Recombinant Proteins; D015534:Trans-Activators; C084010:pancreatic and duodenal homeobox 1 protein; D014700:Verapamil; D005947:Glucose; D002118:Calcium