FoxA2, Nkx2.2, and PDX-1 Regulate Islet β-Cell-Specific mafA Expression through Conserved Sequences Located between Base Pairs −8118 and −7750 Upstream from the Transcription Start Site

Abstract

The MafA transcription factor is both critical to islet ␤-cell function and has a unique pancreatic cell-typespecific expression pattern. To localize the potential transcriptional regulatory region(s) involved in directing expression to the ␤ cell, areas of identity within the 5 flanking region of the mouse, human, and rat mafA genes were found between nucleotides ؊9389 and ؊9194, ؊8426 and ؊8293, ؊8118 and ؊7750, ؊6622 and ؊6441, ؊6217 and ؊6031, and ؊250 and ؉56 relative to the transcription start site. The identity between species was greater than 75%, with the highest found between bp ؊8118 and ؊7750 (ϳ94%, termed region 3). Region 3 was the only upstream mammalian conserved region found in chicken mafA (88% identity). In addition, region 3 uniquely displayed ␤-cell-specific activity in cell-line-based reporter assays. Important regulators of ␤-cell formation and function, PDX-1, FoxA2, and Nkx2.2, were shown to specifically bind to region 3 in vivo using the chromatin immunoprecipitation assay. Mutational and functional analyses demonstrated that FoxA2 (bp ؊7943 to ؊7910), Nkx2.2 (bp ؊7771 to ؊7746), and PDX-1 (bp ؊8087 to ؊8063) mediated region 3 activation. Consistent with a role in transcription, small interfering RNA-mediated knockdown of PDX-1 led to decreased mafA mRNA production in INS-1-derived ␤-cell lines (832/13 and 832/3), while MafA expression was undetected in the pancreatic epithelium of Nkx2.2 null animals. These results suggest that ␤-cell-type-specific mafA transcription is principally controlled by region 3-acting transcription factors that are essential in the formation of functional ␤ cells.

Pancreas organogenesis: From bud to plexus to gland

“…The bZIP factors MafA/B are expressed relatively later in development and are essential to acquire and maintain the mature state of hormone-expressing cells by activating genes important for b-cell function. These genes include insulin, Pdx1, GLUT2, Nkx6.1, Slc30a8 (zinc transporter) and G6pc2 (Glucose-6-phosphatase catalytic sub-unit 2 protein) (Aramata et al, 2005;Zhao et al, 2005, Raum et al, 2006Artner et al, 2007). During development, b-cells move from a MafB þ immature state to MafA þ / MafB þ , and finally, to a MafA þ mature state.…”

A feat of metabolic proportions: Pdx1 orchestrates islet development and function in the maintenance of glucose homeostasis

“…Based upon these studies, several mechanisms have been postulated: (a) increased β cell apoptosis via and Bcl-2), with resulting loss of downregulation of anti-apoptotic genes (Bcl XL functional cell mass [56,67], (b) loss of activity of key Pdx1 target genes whose products are involved in glucose-stimulated insulin transcription and secretion (including Glut2, glucokinase, MafA, Nkx6.1, insulin) [57,63,[68][69][70][71][72][73][74][75][76], and (d) loss of new β cell formation/regeneration [64,77]; in this regard, studies suggest that the action of glucagon-like peptide 1 (GLP1) in enhancing β cell growth and formation in the adult may rely upon activation of Pdx1 in β cells and potential precursor cell types, such as those residing within ducts [74,[78][79][80][81][82].…”

“…These included the genes encoding insulin [68,70,71,126], Glut2 [127,128], glucokinase [129,130], islet amyloid polypeptide (IAPP) [63,75,131,132], Pax4 [133], Nkx6.1 [69,134], MafA [72], liver receptor homolog (Lrh) 1 [135], and Pdx1 itself [37]. In recent years, the direct regulation of many of these genes by Pdx1 has been confirmed using loss-of-function studies in islets and intact animals, and occupancy by Pdx1 of the endogenous genes in islets and β cell lines has been confirmed using the chromatin immunoprecipitation (ChIP) assay [70].…”

Cooperative Transcriptional Regulation of the Essential Pancreatic Islet Gene NeuroD1 (Beta2) by Nkx2.2 and Neurogenin 3

“…Nkx2.2 null mice lack all ␤ cells, most ␣ cells, and a subset of PP cells, which are replaced by the ghrelinproducing population (10,26). In vitro and in vivo data suggest that Nkx2.2 mediates these early islet cell fate decisions by functioning both as a repressor and activator of transcription depending on the developmental context (27)(28)(29)(30)(31). To date, however, only a small number of transcriptional targets of Nkx2.2 have been identified, including Ins2 and MafA (27,30).…”

“…In vitro and in vivo data suggest that Nkx2.2 mediates these early islet cell fate decisions by functioning both as a repressor and activator of transcription depending on the developmental context (27)(28)(29)(30)(31). To date, however, only a small number of transcriptional targets of Nkx2.2 have been identified, including Ins2 and MafA (27,30). Nkx2.2 consensus sites are also present in several regions on the Nkx2.2 promoter, and it has been speculated that Nkx2.2 could control its own transcription through a feedback loop (29,32).…”