Biochemical and Biophysical Research Communications
Nuclear factor 1 regulates adipose tissue-specific expression in the mouse GLUT4 gene☆
Section snippets
Materials and methods
Plasmid construction. The MutNF1:O/E−3K GLUT4 minigene construct was derived from the −3K GLUT4 minigene containing 3237 bp of GLUT4 5′-flanking DNA with a “tag” consisting of 281 bp of foreign DNA [10]. In vitro mutagenesis (Quick change site-directed mutagenesis kit, Stratagene, La Jolla, CA) was used to change the 5′ half-site of the NF1-binding site TGG to TTT (thus also mutating the 3′ O/E half-site). The oligonucleotides used were: 5′-TGAGCACCTGTCCCTTTTGTCCCCTCCAAGAACC-3′ and
Mutations of the binding sites of both NF1 and O/E-1 in the 5′-flanking sequence abolish expression of GLUT4 in WAT
RNase protection analysis was used to examine the expression patterns of minigene and endogenous GLUT4 from MutNF1:O/E−3K GLUT4 minigene transgenic mice. MutNF1:O/E−3K GLUT4 had as a backbone the −3K GLUT4 minigene construct, but contained point-mutations at two nucleotides in the overlapping NF1- and O/E-binding sites at nucleotides −699 and −698 (Fig. 1). Mutation of these two bases was previously shown in gel shift assays to disrupt both NF1 and O/E binding [15], [16]. Two independent
Discussion
We have found that point mutations at bases −699 and −698, or at bases −690 and −689 of the mouse GLUT4 promoter result in a loss of expression of GLUT4 in WAT. Three proteins have been shown to bind to this region. We have demonstrated that NF1 and O/E proteins bind to this region of the mouse GLUT4 promoter [14], [15], [16], while Oshel et al. identified a novel transcription factor which they designated GEF (GLUT4 enhancer factor) which binds to the corresponding region of the human GLUT4
Acknowledgments
This work was supported in part by Grants-in-Aid for scientific research KAKENHI 14770030, 16700481, and 16300216 from the Japanese Ministry of Education, Culture, Sports, Science and Technology (MEXT, Tokyo), by research grants from the Japanese Ministry of Health, Labor and Welfare (Tokyo), by a grant from the Promotion of Fundamental Studies in Health Sciences of Organization for Pharmaceutical Safety and Research (OPSR), and by a grant from the National Institutes of Health (to D.W.C.).
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2021, Biochemical and Biophysical Research CommunicationsCitation Excerpt :At transcriptional level, it had been reported that GLUT8 promoter has binding sequences for the Nuclear Factor 1 (NF1) and Sex-determining Region Y (SRY). NF1 is involved in insulin and cyclic adenosine monophosphate-dependent expression of GLUT4 in adipocytes and muscle cells, and SRY regulates GLUT5 expression in the testis [33,34]. The results of DOG uptake and fructose competence presented reveal that the mammary GLUT8 transporter isolated from murine mammary glands on day 2 of lactation is fully functional.
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Abbreviations: ASE, adipose tissue-specific element(s); HFRE, high-fat responsive element(s); WAT, white adipose tissue; BAT, brown adipose tissue.
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Present address: Department of Clinical Dietetics and Human Nutrition, Faculty of Pharmaceutical Sciences, Josai University, Saitama, Japan.