Abstract
Pancreatic β-cell-restricted expression of insulin is established through several critical cis-regulatory elements located in the insulin gene promoter region. The principal cis elements are A-boxes, E1, and C1/RIPE3b. The β-cell-enriched transcription factors Pdx1 and Beta2 bind to the A-boxes and E1 element, respectively. A β-cell-specific trans-acting factor binding to C1/RIPE3b (termed RIPE3b1 activator) was detected by electrophoretic mobility shift assay and has been identified as MafA, a member of the Maf family of basic leucine zipper (bZip) proteins. Here, ATF2, a member of the ATF/CREB family of basic leucine zipper proteins, was identified as a component of the RIPE3b1 activator. ATF2 alone was unable to bind to the C1/RIPE3b element but acquired binding capacity upon complex formation with MafA. ATF2 also interacted with Pdx1 and Beta2, and co-expression of ATF2, MafA, Pdx1, and Beta2 resulted in a synergistic activation of the insulin promoter. Immunohistochemical analysis of mouse pancreas tissue sections showed that ATF2 is enriched in islet endocrine cells, including β-cells. RNAi-mediated knockdown of MafA or ATF2 in the MIN6 β-cell line resulted in a significant decrease in endogenous levels of insulin mRNA. These data indicate that ATF2 is an essential component of the positive regulators of the insulin gene expression.
Highlights
The principal cis-elements involved in regulating -cell-restricted expression are A-boxes and the E1 and C1/RIPE3b elements
We demonstrated that ATF2, a member of the ATF/CREB family of basic leucine zipper (bZip) proteins, is a component of the RIPE3b1 activator
Previous studies have shown that ATF2 is involved in the regulation of insulin expression through binding to cAMP-responsive element (CRE)-like elements in the promoter region [19, 20]
Summary
The principal cis-elements involved in regulating -cell-restricted expression are A-boxes ( known as A1, A3, and GG2 elements in humans) and the E1 and C1/RIPE3b elements. HeLa cells were transfected with a luciferase reporter gene driven by the human insulin promoter together with various combinations of expression plasmids for ATF2 and MafA, and Pdx1 and Beta2, two other important insulin promoter activators.
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