Syntaxin-1a is a Direct Target of miR-29a in Insulin-producing β-Cells

Abstract

Downregulation of proteins involved in the ­exocytotic machinery has been implicated in the impairment of normal β-cell function in response to high glucose levels. Syntaxin-1a ­(Stx-1a) is one of two t-SNAREs involved in insulin exocytosis and decreased expression of Stx-1a protein impairs glucose-stimulated insulin secretion (GSIS) in isolated rat pancreatic islets. In diabetic patients Stx-1a protein levels are reduced, but the mechanism of this suppression is unknown.

MicroRNAs are small noncoding RNAs, which are important regulators of gene-expression at the post transcriptional level, partially binding to the 3′UTRs of their target gene transcripts either mediating transcript degradation or inhibiting translation. We have recently shown that miR-29a is upregulated in response to elevated glucose levels in β-cells and is involved in mediating the negative effect of high glucose levels on GSIS. Stx-1a has a predicted target site of miR-29a present in its 3′ untranslated region. The objective of this study was to evaluate whether miR-29a targets Stx-1a directly to decrease mRNA and/or protein levels in response to glucose. Stx-1a mRNA and protein levels decreased in β-cells treated with increased glucose levels. Overexpression of miR-29a decreased Stx-1a mRNA and protein levels. Furthermore, miR-29a decreases the response of a luciferase reporter construct containing the predicted target site normally present in the Stx-1a gene. When 2 nucleotides are mutated in this target site, responsiveness to miR-29a disappears, confirming miR-29a binding to this sequence. Collectively, these data implicate miR-29a as a mediator of glucose-induced downregulation of Stx-1a in β-cells.

• References

• 1 Ostenson CG, Gaisano H, Sheu L, Tibell A, Bartfai T. Diabetes 2006; 55: 435-440
• 2 Nagamatsu S, Nakamichi Y, Yamamura C, Matsushima S, Watanabe T, Ozawa S, Furukawa H, Ishida H. Diabetes 1999; 48: 2367-2373
• 3 Dubois M, Vacher P, Roger B, Huyghe D, Vandewalle B, Kerr-Conte J, Pattou F, Moustaid-Moussa N, Lang J. Endocrinology 2007; 148: 1605-1614
• 4 Bagge A, Clausen TR, Larsen S, Ladefoged M, Rosenstierne MW, Larsen L, Vang O, Nielsen JH, Dalgaard LT. Biochem Biophys Res Commun 2012; 426: 266-272
• 5 Larsen L, Rosenstierne MW, Gaarn LW, Bagge A, Pedersen L, Dahmcke CM, Nielsen JH, Dalgaard LT. PLoS One 2011; 6: e25997
• 6 Zuker M. Nucleic Acids Res 2003; 31: 3406-3415
• 7 Andersson SA, Olsson AH, Esguerra JL, Heimann E, Ladenvall C, Edlund A, Salehi A, Taneera J, Degerman E, Groop L, Ling C, Eliasson L. Mol Cell Endocrinol 2012; 364: 36-45
• 8 Leung YM, Kwan EP, Ng B, Kang Y, Gaisano HY. Endocr Rev 2007; 28: 653-663
• 9 Lam PP, Leung YM, Sheu L, Ellis J, Tsushima RG, Osborne LR, Gaisano HY. Diabetes 2005; 54: 2744-2754
• 10 Roggli E, Gattesco S, Caille D, Briet C, Boitard C, Meda P, Regazzi R. Diabetes 2012; 61: 1742-1751
• 11 Vikman J, Jimenez-Feltstrom J, Nyman P, Thelin J, Eliasson L. FASEB J 2009; 23: 58-67
• 12 Pullen TJ, da SX, Kelsey G, Rutter GA. Mol Cell Biol 2011; 31: 3182-3194
• 13 Landgraf P, Rusu M, Sheridan R, Sewer A, Iovino N, Aravin A, Pfeffer S, Rice A, Kamphorst AO, Landthaler M, Lin C, Socci ND, Hermida L, Fulci V, Chiaretti S, Foa R, Schliwka J, Fuchs U, Novosel A, Muller RU, Schermer B, Bissels U, Inman J, Phan Q, Chien M, Weir DB, Choksi R, De VG, Frezzetti D, Trompeter HI, Hornung V, Teng G, Hartmann G, Palkovits M, Di LR, Wernet P, Macino G, Rogler CE, Nagle JW, Ju J, Papavasiliou FN, Benzing T, Lichter P, Tam W, Brownstein MJ, Bosio A, Borkhardt A, Russo JJ, Sander C, Zavolan M, Tuschl T. Cell 2007; 129: 1401-1414