Mechanisms of Stem Cell Maintenance and Cell Differentiation in the Intestinal Epithelium

Constant regeneration of the intestinal epithelium, a dynamic tissue with vital digestive and barrier functions, depends on proliferation of resident stem cells and their differentiation into mature cell types. This epithelium thus provides an ideal model to study stem cells and mechanisms of cell differentiation in an adult tissue. The identification of a proliferative population of intestinal stem cells (ISCs) at the base of intestinal crypts presents the prospect of understanding their regulation by extrinsic and intrinsic factors. Although activation of Wnt signaling in ISCs is thought to be one crucial function of the ISC niche, the cellular source of Wnt ligands is uncertain. Chapter 2 addresses this question through genetic elimination of Wnt ligand secretion in candidate niche cell populations. The data reveal that Wnts originating in any of the sources considered in the literature – the epithelium (including Paneth cells) and sub-epithelial myofibroblasts – are not required for ISC function. These data support models of highly complex cell redundancy or alternative, non-Wnt ligands. Chapter 3 investigates the cell-intrinsic contributions of an intestine-restricted transcription factor (TF), CDX2, to important ISC behaviors. Cdx2 loss in vivo perturbs ISC proliferation and differentiation, distinct from its functions in mature enterocytes. Analysis of candidate direct CDX2 target genes in ISCs suggests that CDX2 modulates Fibroblast Growth Factor signaling, thus opening new avenues of investigation. Although cells that differentiate from ISCs rely on gene expression changes mediated by TFs, loss of several individual TFs in vivo has modest effects on intestinal function. Chapter 4 characterizes the functional interactions of CDX2, which has many properties of a master regulator, with two other intestinal TFs: GATA4 and Hepatocyte Nuclear Factor 4 alpha (HNF4A). Analysis of compound mutant mouse intestines elucidated combinatorial roles for CDX2 with GATA4 in crypt cell proliferation and with HNF4A in enterocyte differentiation. Building on this foundation, Chapter 5 describes preliminary investigations into another TF, HNF1A, in intestinal gene regulation and its relationship with CDX2 in controlling cell differentiation and tissue architecture. These studies highlight the complexities of TF interactions and the functions of diverse TF complexes in controlling tissue-specific genes during cell differentiation