Enteroendocrine cells are an attractive model for study of differentiation due to expression of distinct markers for each cell type. Understanding enteroendocrine cell differentiation will require detailed knowledge of both the transcriptional controls for each of the hormone genes as well as the factors that control cell proliferation as cells terminally differentiate. A transcriptional enhancer that directs expression of the secretin gene to secretin-producing cells has been characterized and consists of an E box, 2 Sp 1 sites, and a rough upstream site which interacts with an as yet unidentified factor. A recently described, tissue specific helix loop helix protein, BETA2, binds to the E box and activates transcription. Gene targeting experiments reveal that secretin cells fail to develop in mice lacking a functional BETA2 gene, indicating a critical role for this factor in secretin gene expression. BETA2 interacts with the 300 kD nuclear protein, p300. p300 is known to function as a transcription coactivator and as an inhibitor of cell proliferation. Involvement of p300 in transcriptional control of the secretin gene may explain the exclusive localization of secretin cells in the nondividing villus compartment of the intestine. The four aims of this proposal will further examine the transcriptional controls of secretin cell differentiation.
Aim 1 will examine the ontogeny of intestinal BETA2 expression and determine whether BETA2 functions as a factor determining cell fate or cellular differentiation in enteroendocrine cells, using a BETA2 knockout mouse model.
Aim 2 will characterize the interactions between BETA2 and p300 both in vitro and in vivo. The specific domains of p300 and BETA2 required for their association will be defined by structure-function studies using a series of truncated BETA2 and p300 proteins.
Aim 3 will examine the role of p300 in controlling differentiation and proliferation of secretin cells by studying the effects of wild type and mutant SV40 large T antigens expressed in transgenic mice. The objective of aim 4 is to clone the factor that binds to the 5' end of the secretin enhancer and to characterize its function in secretin gene transcription. The proposed work is likely to provide important information toward understanding transcriptional control of cell fate determination and cellular proliferation of endocrine cells in the GI tract. it is likely that insights gained from studying endocrine cells will have broad applicability to our knowledge of how the gastrointestinal tract develops.
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