Effects of Novel Polymorphisms in the RAGE Gene on Transcriptional Regulation and Their Association With Diabetic Retinopathy

Abstract

Interactions between advanced glycation end products (AGEs) and the receptor for AGE (RAGE) are implicated in the vascular complications in diabetes. We have identified eight novel polymorphisms, of which the ؊1420 (GGT)n, ؊1393 G/T, ؊1390 G/T, and ؊1202 G/A were in the overlapping PBX2 3 untranslated region (UTR), and the ؊429 T/C (66.5% TT, 33.5% TC/CC), ؊407 to -345 deletion (99% I, 1% I/D, 0% D), ؊374 T/A (66.4% TT, 33.6% TA/AA), and ؉20 T/A were in the RAGE promoter. To evaluate the effects on transcriptional activity, we measured chloramphenicol acetyl transferase (CAT) reporter gene expression, driven by variants of the -738 to ؉49 RAGE gene fragment containing the four polymorphisms identified close to the transcriptional start site. The -429 C, ؊374 A, and 63-bp deletion alleles resulted in a mean increase of CAT expression of twofold (P < 0.0001), threefold (P < 0.001), and fourfold (P < 0.05), respectively, with the -374 T and A alleles yielding highly differential binding of nuclear protein extract from both monocyte-and hepatocyte-derived cell lines. The prevalence of the functional polymorphisms were investigated in subjects with type 2 diabetes (106 with and 109 without retinopathy), with the -429 C allele showing an increase in the retinopathy group (P < 0.05). These data suggest that the polymorphisms involved in differences in RAGE gene regulation may influence the pathogenesis of diabetic vascular complications. Diabetes 50:1505-1511, 2001

Rage gene promoter polymorphisms and diabetic retinopathy in a clinic-based population from South India

“…Given the presence of LD between these polymorphisms as has been reported in other studies as well, 18,19 we also performed haplotype tests for association which also yielded no evidence for association. In this study, we also compared the allele frequencies between the nondiabetics and the DM group, and also found them to be similar.…”

“…PCR-RFLP genotyping was carried out as described. 18 PCR consisted of 100-150 ng of DNA denaturing at 941C, annealing at 561C, extension at 721C for 30 cycles, followed by a final extension at 721C for 5 min in a 50 ml reaction. The PCR products were then digested with Alu I (New England Bio Labs) at 371C for 6 h for À429 T/C polymorphism and Tsp509 I (New England Bio Labs) at 651C for 16 h for À374 T/A polymorphism.…”

“…The PCR products were then digested with Alu I (New England Bio Labs) at 371C for 6 h for À429 T/C polymorphism and Tsp509 I (New England Bio Labs) at 651C for 16 h for À374 T/A polymorphism. 18 The restricted products were separated in a 3% agarose gel and visualized using ethidium bromide staining.…”

“…Among many candidate genes that have been found to be associated with retinopathy, the gene encoding RAGE is of particular significance, because the RAGE-mediated signaling induces key pathological changes in the microvasculature leading to DR. A functional polymorphism Gly82Ser in exon 3 has been previously implicated in DR of which the Ser allele of this Gly82Ser polymorphism has been shown to be protective in the development of DR in the Asian-Indian population. 17 Studies performed by Hudson et al 18 have shown that three polymorphisms located in the promoter region (À429TC and À374TA and 63 bp del) of the RAGE gene are involved in the upregulation of the gene and thereby could play a pivotal role in the pathogenesis leading to DR. The 63 bp deletion polymorphism is in the region encompassing the À374 region, and these promoter polymorphisms exert their effect by affecting the transcriptional regulation of the RAGE gene.…”

RAGE and arthritis: the G82S polymorphism amplifies the inflammatory response

“…In this context, recently identified polymorphisms within the promoter of the gene encoding RAGE may hold the key to unique elements controlling enhanced transcription of the RAGE gene and increased expression of cell surface RAGE. 54 As a number of studies have suggested that expression of RAGE is increased at sites of ligand accumulation such as diabetic atherosclerotic lesions, infected periodontium or Alzheimer disease brain, 13,18,55 polymorphisms within regulation elements and/or ligand-binding regions may, alone or in combination, orchestrate RAGE's availability and/or functional potential in a given milieu. 56 In conclusion, we propose that these observations in an animal model of inflammatory arthritis and in S100/calgranulin-stimulated activation of 82S RAGEbearing cells highlight ligand-RAGE engagement as an amplifier of proinflammatory mechanisms in immune/ inflammatory diseases.…”