OBJECTIVESeveral transcription factors are essential to pancreatic islet β-cell development, proliferation, and activity, including MafA and MafB. However, MafA and MafB are distinct from others in regard to temporal and islet cell expression pattern, with β-cells affected by MafB only during development and exclusively by MafA in the adult. Our aim was to define the functional relationship between these closely related activators to the β-cell.RESEARCH DESIGN AND METHODSThe distribution of MafA and MafB in the β-cell population was determined immunohistochemically at various developmental and perinatal stages in mice. To identify genes regulated by MafB, microarray profiling was performed on wild-type and MafB−/− pancreata at embryonic day 18.5, with candidates evaluated by quantitative RT-PCR and in situ hybridization. The potential role of MafA in the expression of verified targets was next analyzed in adult islets of a pancreas-wide MafA mutant (termed MafAΔPanc).RESULTSMafB was produced in a larger fraction of β-cells than MafA during development and found to regulate potential effectors of glucose sensing, hormone processing, vesicle formation, and insulin secretion. Notably, expression from many of these genes was compromised in MafAΔPanc islets, suggesting that MafA is required to sustain expression in adults.CONCLUSIONSOur results provide insight into the sequential manner by which MafA and MafB regulate islet β-cell formation and maturation.

The C1/RIPE3b1 (؊118/؊107 bp) binding factor regulates pancreatic-␤-cell-specific and glucose-regulated transcription of the insulin gene. In the present study, the C1/RIPE3b1 activator from mouse ␤TC-3 cell nuclear extracts was purified by DNA affinity chromatography and two-dimensional gel electrophoresis. C1/RIPE3b1 binding activity was found in the roughly 46-kDa fraction at pH 7.0 and pH 4.5, and each contained N-and C-terminal peptides to mouse MafA as determined by peptide mass mapping and tandem spectrometry. MafA was detected in the C1/RIPE3b1 binding complex by using MafA peptide-specific antisera. In addition, MafA was shown to bind within the enhancer region (؊340/؊91 bp) of the endogenous insulin gene in ␤TC-3 cells in the chromatin immunoprecipitation assay. These results strongly suggested that MafA was the ␤-cell-enriched component of the RIPE3b1 activator. However, reverse transcription-PCR analysis demonstrated that mouse islets express not only MafA but also other members of the large Maf family, specifically c-Maf and MafB. Furthermore, immunohistochemical studies revealed that at least MafA and MafB were present within the nuclei of islet ␤ cells and not within pancreas acinar cells. Because MafA, MafB, and c-Maf were each capable of specifically binding to and activating insulin C1 element-mediated expression, our results suggest that all of these factors play a role in islet ␤-cell function.Insulin is an essential regulator of metabolism. This hormone, which is synthesized by the ␤ cells of the islets of Langerhans, increases the storage of glucose, fatty acids, and amino acids through its actions in liver, adipose tissue, and muscle. Experiments performed in vivo with transgenic animals have established that the cis-acting elements controlling ␤-cell-selective expression are located within the insulin enhancer region, which is found between nucleotides Ϫ340 and Ϫ91 relative to the transcription start site. Several key control elements within the enhancer have been identified, including C2 (Ϫ317/ Ϫ311 bp), A3 (Ϫ201/Ϫ196 bp), C1 (Ϫ118/Ϫ107 bp), and E1 (Ϫ100/Ϫ91 bp) (37,60,67). Mutations that decrease the binding affinity of the A3, C1, and E1 activators also reduce glucose-regulated transcription (37,60,67).The activator of insulin C2-element stimulated transcription is Pax6 (61). Proteins in the Pax family all contain a paired box bipartite DNA-binding domain, although Pax6 also has a homeodomain. The Pdx-1 homeodomain protein (formerly known as IPF-1, STF-1, and IDX-1) is the regulator of A3 elementactivated expression (46,48,49,50), whereas the E1 activator is a heterodimer composed of proteins in the basic helix-loophelix family that are enriched in islets (i.e., BETA2 [42]) and generally distributed (i.e., HEB [51] and E2A [2,10,17,65]). In the adult pancreas, Pax6 (61) and BETA2 (42) are found in all islet cell types, whereas Pdx-1 appears to be found only in ␤ cells, a subset of islet ␦ cells (48,49), and exocrine acinar cells (71,80). These transcription factors are necessary for ma...

Pancreatic endocrine cell differentiation depends on transcription factors that also contribute in adult insulin and glucagon gene expression. Islet cell development was examined in mice lacking MafB, a transcription factor expressed in immature ␣ (glucagon ؉ ) and ␤ (insulin ؉ ) cells and capable of activating insulin and glucagon expression in vitro. We observed that MafB ؊/؊ embryos had reduced numbers of insulin ؉ and glucagon ؉ cells throughout development, whereas the total number of endocrine cells was unchanged. Moreover, production of insulin ؉ cells was delayed until embryonic day (E) 13.5 in mutant mice and coincided with the onset of MafA expression, a MafB-related activator of insulin transcription. MafA expression was only detected in the insulin ؉ cell population in MafB mutants, whereas many important regulatory proteins continued to be expressed in insulin ؊ ␤ cells. However, Pdx1, Nkx6.1, and GLUT2 were selectively lost in these insulin-deficient cells between E15.5 and E18.5. MafB appears to directly regulate transcription of these genes, because binding was observed within endogenous control region sequences. These results demonstrate that MafB plays a previously uncharacterized role by regulating transcription of key factors during development that are required for the production of mature ␣ and ␤ cells.insulin ͉ MafA ͉ pancreas development

Insulin gene expression is regulated by several islet-enriched transcription factors. However, MafA is the only ␤ cell-specific activator. Here, we show that MafA selectively induces endogenous insulin transcription in non-␤ cells. MafA was also first detected in the insulin-producing cells formed during the second and predominant phase of ␤ cell differentiation, and absent in the few insulin-positive cells found in Nkx6.1 ؊/؊ pancreata, which lack the majority of second-phase ␤ cells. These results demonstrate that MafA is a potent insulin activator that is likely to function downstream of Nkx6.1 during islet insulin-producing cell development.