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Pdx-1 is required for activation in vivo from a duodenum-specific enhancer
M R Dusing1, E A Florence, D A Wiginton
1Department of Pediatrics, Division of Developmental Biology, University of Cincinnati College of Medicine and Children's Hospital Research Foundation, Cincinnati, Ohio 45229, USA.
Insights
The transcription factor PDX-1 is essential for activating adenosine deaminase (ADA) gene expression in the developing duodenum. While PDX-1 binding is crucial in vivo, other proteins are also needed for full ADA enhancer function.
Area of Science:
- Developmental Biology
- Molecular Genetics
- Gastroenterology
Background:
- Adenosine deaminase (ADA) exhibits specific spatiotemporal expression in the developing mammalian small intestine.
- High-level ADA expression localizes to the duodenal villous epithelium during intestinal maturation.
Purpose of the Study:
- To identify regulatory elements controlling the duodenum-specific expression of the ADA gene.
- To investigate the role of duodenal proteins interacting with the ADA enhancer region.
Main Methods:
- Identification of proteins binding to a regulatory module in the second intron of the human ADA gene.
- Utilizing transgenic mouse models with mutated PDX-1 binding sites in the ADA enhancer.
- Performing co-transfection experiments to assess enhancer activity.
Main Results:
- The homeobox protein PDX-1 was identified as a key duodenal protein interacting with the ADA enhancer.
- Loss of PDX-1 binding in vivo led to a complete absence of high-level ADA activation in the duodenum.
- PDX-1 alone was insufficient for significant transactivation, indicating the involvement of other factors.
Conclusions:
- PDX-1 is an essential transcription factor for in vivo activation of the ADA gene in the duodenum.
- The ADA enhancer requires PDX-1 and additional interacting proteins for full functional activity.
- This study elucidates a critical regulatory mechanism for ADA expression during intestinal development.
Abstract:
The purine metabolic gene adenosine deaminase (ADA) is expressed along a defined spatiotemporal pattern in the developing mammalian small intestine, where high-level expression is limited to the villous epithelium of the duodenum. This activation is observed in rodents as the intestine completes the final maturation resulting in adult crypt-villus structures at 2-3 weeks postpartum. A regulatory module responsible for this pattern of expression has been identified in the second intron of the human ADA gene. Of the multiple duodenal proteins that can interact with this small duodenal enhancer region, the studies contained in this work describe the identification of five of these proteins as the dispersed homeobox protein PDX-1. This transcription factor exhibits a profile of expression in the small intestine similar to that observed for ADA, making it an ideal candidate factor for the duodenum-specific ADA enhancer. Loss of PDX-1 binding, via a PDX-1 mutated enhancer transgenic construction, resulted in complete loss of high-level activation in the duodenum, demonstrating the absolute requirement for this factor in vivo. However, co-transfection experiments suggest that other proteins that bind the enhancer are also required for enhancer function because PDX-1 alone was incapable of significant transactivation.
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