The overall objective of this ongoing project is to elucidate the molecular mechanisms that regulate transcription of intestine- specific genes. In the first funding period of this proposal the intestine-specific promoter of the sucrase-isomaltase (SI) gene has been well characterized. We have shown that an evolutionarily conserved SI gene promoter is capable of directing transcription to enterocytes in complex patterns that recapitulate the expression of the endogenous SI gene. The results of these transgenic mouse experiments provide a firm foundation for focusing on this promoter to dissect critical mechanisms of transcriptional initiation in enterocytes. Three primary DNA regulatory elements, and their associated DNA binding proteins, are important for transcriptional activation of the SI promoter. The three types of transcription factors include caudal-related homeodomain proteins (Cdx1 and Cdx2), GATA transcription factors, and hepatocyte nuclear factor 1 (HNF1). We present new data that another intestinal gene, intestinal phospholipase A/lysophospholipase (IPAL), is expressed in a pattern very similar to SI and has the binding sites in its promoter for the same three transcription factors. This finding could provide a breakthrough in our attempt to develop general paradigms for transcription of a particular group of intestinal genes. In this competing renewal, we propose to explore a model of intestinal gene transcription that incorporates multiple transcription factors acting in a combinatorial network. This will be accomplished through four Specific Aims:
Aim 1. To define the functional domains of Cdx proteins and identify other members of the transcriptional network of proteins that interact with those domains. These experiments will result in characterization of the protein contacts made by Cdxl and Cdx2 and how those contacts function in transcriptional initiation.
Aim 2. To examine the function of intestinal GATA transcription factors (GATA4, 5, and 6) on the GATA element in the SI promoter.
Aim 3. To analyze the function of the 'PAL promoter, clone the mouse 'PAL gene, and examine regulation in transgenic mice.
Aim 4, To explore the functional interactions between DNA regulatory elements and their cognate DNA binding proteins, particularly Cdx and GATA proteins. These data will allow us to propose a multidimensional model for transcriptional activation in the intestine, By understanding the molecular mechanisms of these transcriptional processes, generalized paradigms for intestinal gene transcription will emerge. As we assemble the complex puzzle of interacting proteins that regulate transcription of the SI and 'PAL promoters, new mechanisms will be discovered that may have important implications for the processes of intestinal development and differentiation.

National Institute of Health (NIH)
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Research Project (R01)
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General Medicine A Subcommittee 2 (GMA)
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University of Pennsylvania
Internal Medicine/Medicine
Schools of Medicine
United States
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Xiao, Fang; Crissey, Mary Ann S; Lynch, John P et al. (2005) Intestinal metaplasia with a high salt diet induces epithelial proliferation and alters cell composition in the gastric mucosa of mice. Cancer Biol Ther 4:669-75
Rankin, Erinn B; Xu, Wei; Silberg, Debra G et al. (2004) Putative intestine-specific enhancers located in 5' sequence of the CDX1 gene regulate CDX1 expression in the intestine. Am J Physiol Gastrointest Liver Physiol 286:G872-80
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Boudreau, Francois; Rings, Edmond H H M; van Wering, Herbert M et al. (2002) Hepatocyte nuclear factor-1 alpha, GATA-4, and caudal related homeodomain protein Cdx2 interact functionally to modulate intestinal gene transcription. Implication for the developmental regulation of the sucrase-isomaltase gene. J Biol Chem 277:31909-17
Suh, Eun Ran; Ha, Chong S; Rankin, Erinn B et al. (2002) DNA methylation down-regulates CDX1 gene expression in colorectal cancer cell lines. J Biol Chem 277:35795-800
Boudreau, F; Zhu, Y; Traber, P G (2001) Sucrase-isomaltase gene transcription requires the hepatocyte nuclear factor-1 (HNF-1) regulatory element and is regulated by the ratio of HNF-1 alpha to HNF-1 beta. J Biol Chem 276:32122-8
Rings, E H; Boudreau, F; Taylor, J K et al. (2001) Phosphorylation of the serine 60 residue within the Cdx2 activation domain mediates its transactivation capacity. Gastroenterology 121:1437-50
Suh, E; Wang, Z; Swain, G P et al. (2001) Clusterin gene transcription is activated by caudal-related homeobox genes in intestinal epithelium. Am J Physiol Gastrointest Liver Physiol 280:G149-56
Lorentz, O; Suh, E R; Taylor, J K et al. (1999) CREB-binding [corrected] protein interacts with the homeodomain protein Cdx2 and enhances transcriptional activity. J Biol Chem 274:7196-9
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