Analysis of polymorphism of the GLUT2 promoter in NIDDM patients and its functional consequence to the promoter activity.
Glucose transporter type 2 (GLUT2), along with glucokinase, has been implicated to participate in glucose-induced insulin secretion in pancreatic beta-cells. Recently, several sequence variations in the promoter of GLUT2 have been identified in patients with non-insulin dependent diabetes mellitus (NIDDM), but the functional effects of these polymorphisms on promoter activity have not previously been studied. We compared the incidence of sequence variations in the GLUT2 promoter in 100 normal subjects and 100 NIDDM patients. Sequencing of the promoter region (-294 to +301) revealed that an A --> G variant at position -44 was found in 45 of 100 NIDDM patients, but only in 23 of 100 normal subjects. In addition, -269 A --> C and + 103 A --> G mutations were identified in a single diabetic patient. Electrophoretic mobility shift assays using double-stranded oligonucleotide containing -44A as a probe showed a clearly shifted band of DNA-protein. To examine whether the sequence variation at position -44 affects the promoter activity, we carried out in vitro transfection experiments. Site-specific mutagenesis at position -44 region from A to C, T, or G resulted in reductions of CAT activity by 52.3%, 63.8%, and 63.6%, respectively. The -269 A --> C and + 103 A --> G mutations also decreased the promoter activity. These results suggest that polymorphisms at positions -269, -44, or + 103 may affect GLUT2 gene transcription, possibly associated with reduced expression of the GLUT2 gene in NIDDM patients.
Cha
Ji-Young
JY
Department of Biochemistry and Molecular Biology, Institute of Genetic Science, Yonsei University College of Medicine, Seoul, Korea.
Kim
Hyon-Suk
HS
Kim
Ha-Il
HI
Im
Seung-Soon
SS
Kim
So-Youn
SY
Kim
Jae-Woo
JW
Yeh
Byung-Il
BI
Ahn
Yong-Ho
YH
D002460:Cell Line; D003924:Diabetes Mellitus, Type 2 / Q000235:genetics*; D024202:Electrophoretic Mobility Shift Assay; D051273:Glucose Transporter Type 2; D006801:Humans; D007515:Islets of Langerhans / Q000166:cytology / Q000378:metabolism / Q000502:physiology; D009004:Monosaccharide Transport Proteins / Q000235:genetics*; D016297:Mutagenesis, Site-Directed; D011110:Polymorphism, Genetic; D011401:Promoter Regions, Genetic / Q000235:genetics*; D015533:Transcriptional Activation / Q000235:genetics; D014162:Transfection
D051273:Glucose Transporter Type 2; D009004:Monosaccharide Transport Proteins