Identification of a Glucose Response Element in the Promoter of the Rat Glucagon Receptor Gene

Abstract

We cloned the 5 upstream region of the rat glucagon receptor gene, demonstrating that the 5 noncoding domain of the glucagon receptor mRNA contained two untranslated exons of 131 and 166 nucleotides (nt), respectively, separated by two introns of 0.6 and 3.2 kilobase pairs. We also observed an alternative splicing involving the 166-base pair exon. Cloning of up to 2 kilobase pairs of the newly identified genomic domain and transfection of various constructs driving a reporter gene, in pancreatic islet cell line INS-1, uncovered a strong glucose regulation of the promoter activity of plasmids containing up to nucleotide ؊868, or more, upstream from the transcriptional start point. This promoter activity displayed threshold-like behavior, with low activity of the promoter below 5 mM glucose, and maximal activation as of 10 mM glucose. This glucose regulation was mapped to a highly palindromic 19-nucleotide region between nt ؊545 and ؊527. Indeed, deletion or mutation of this sequence abolished the glucose regulation. This domain contained two palindromic "E-boxes" CACGTG and CAGCTG separated by 3 nt, a feature similar to the "L4 box" found in the pyruvate kinase L gene promoter. This is the first description of a G protein-coupled receptor gene promoter regulated by glucose.The primary physiological role of glucagon, together with insulin, is maintenance of normal glycemia. The liver has a central role in handling absorbed nutrients and in the regulation of hepatic glycogen disposability; this requires high density of glucagon receptors in the liver (for review see Refs. 1 and 2). The glucagon receptor mRNA has also been detected at variable levels in other tissues such as heart, kidney, adrenal gland, and adipose tissue (3-5), as well as in pancreatic islets, especially in B cells (6, 7). The expression of the glucagon receptor mRNA is stimulated by glucose and inhibited by cyclic AMP, both in liver (8) and in cultured endocrine cells (9). Consequently, the promoter of the glucagon receptor gene could contain regulatory elements for these factors. It has been recently suggested by Burcelin et al. (10) that the glucose regulation of the glucagon receptor mRNA level differs from the glucose regulation of other genes such as Glut2, and might be mediated by triose metabolites, suggesting the existence of new enhancer sequences. To address this question, a prerequisite is the precise knowledge of the complete 5Ј mRNA sequence. For the glucagon receptor mRNA, there was an ambiguity; the glucagon receptor cDNA has been first cloned (from rat) simultaneously by us (11, 12) and by Jelinek et al. (13). Our sequence (GenBank accession number L04796) was almost identical to that of Jelinek (GenBank accession number M96674) except for the 5Ј end; the first 25 nt 1 in our sequence (corresponding to positions Ϫ105 to Ϫ80 from the ATG codon) differed from the sequence published by Jelinek (13). The coding domain of the rat glucagon receptor gene is highly fragmented; it contains 12 introns with uneven splicing maturation (14) t...

International Union of Pharmacology. XXXV. The Glucagon Receptor Family

“…Glucagon receptor expression in islet cells is up-regulated by glucose, and glucose-responsive sequences have been identified in the glucagon receptor gene promoter Yamato et al, 1997;Portois et al, 1999). Agents such as forskolin and 3-isobutyl-1-methylxanthine, which increase levels of cAMP, and the glucocorticoid dexamethasone, inhibit glucagon receptor expression in rat islets, whereas somatostatin, which reduces cAMP, increases levels of islet glucagon receptor RNA transcripts .…”

Glucose-stimulated and self-limiting insulin production by glucose 6-phosphatase promoter driven insulin expression in hepatoma cells

“…A glucose responsive element (GlRE or ChoRE) has been identified in promoters of the liver-type pyruvate kinase gene (L-PK), the spot 14 (S14) gene, and the glucagon receptor gene. [36][37][38] This element contains two E-boxes separated by a few nucleotides. A similar consensus sequence does not exist in the G6Pase gene promoter.…”

Recent progress on the role of ChREBP in glucose and lipid metabolism [Review]

“…Glucose induces rat Gcgr mRNA expression; however, the precise mechanism remains unknown [64,65]. The G-box has previously been reported to be responsible for glucose regulation of Gcgr mRNA expression [65]. In the rat Gcgr promoter, a putative ChoRE (−554 bp/−538 bp) is localized near the G-box (−543 bp/−529 bp) [16,65].…”

“…Glucose induces rat Gcgr mRNA expression; however, the precise mechanism remains unknown [64,65]. The G-box has previously been reported to be responsible for glucose regulation of Gcgr mRNA expression [65].…”

“…The G-box has previously been reported to be responsible for glucose regulation of Gcgr mRNA expression [65]. In the rat Gcgr promoter, a putative ChoRE (−554 bp/−538 bp) is localized near the G-box (−543 bp/−529 bp) [16,65]. In rat INS-1E insulinoma cells, deletion studies of the rat Gcgr promoter showed that ChoRE is a minimal glucose response element early stage of metabolic syndrome, and ChREBP transactivity is probably rather suppressed during the late stage of metabolic syndrome with severe insulin resistance and insufficient insulin secretion.…”