Intestinal stem cells (ISCs) reside at the base of glandular structures, termed crypts, in the small intestinal epithelium. ISCs are defined by functional characteristics of self-renewal and multipotency, are thought to exist as multiple distinct populations, and exhibit remarkable plasticity during homeostasis as well in post-damage regeneration. The genetic regulation of this plasticity, especially in regards to myriad chromatin regulator elements, remains incompletely understood. At this career stage, my central goal is to supplement my expertise in ISC biology with new training in genomics in order to establish an independent career focused on chromatin regulation in ISCs. My preliminary data demonstrate that Tet1, which is a critical regulator DNA demethylation, is specifically upregulated in cell populations consistent with active and reserve ISCs. Additionally, subunits of the PRC2 complex, which is recruited to target sites by TET1, are differentially expressed between these two ISC populations. The central hypothesis of this study is that Tet1 establishes distinct gene regulatory networks in active and reserve ISCs. To test this hypothesis and meet my training goals, I will take a two-pronged approach to: (1) characterize the Tet1/PRC2 gene regulatory network by RNA- and ChIP-seq (Aims 1 and 2) and (2) apply these genome-scale findings to understanding functional outcomes in Tet1-null ISCs. This strategy will allow me to gain expertise in the experimental, conceptual, and practical approaches necessary to succeed in my independent career goals, as well as provide ample data to build a foundation for subsequent independent funding.
The intestinal epithelium, which is responsible for digestive and barrier functions in the gastrointestinal tract, undergoes constant, rapid renewal by intestinal stem cells (ISCs). ISC function is shared between ?active? and ?reserve? stem cell pools, which have different context-dependent roles in intestinal physiology, disease, and repair. However, the genomic mechanisms that differentiate ISC populations from one another remain poorly characterized. Tet1, which regulates genome structure by active DNA demethylation as well as recruitment of other chromatin modifying protein complexes like PRC2, is strongly expressed in ISCs. This proposal aims to delineate the role of Tet1 in establishing distinct ISC cell states through integration of genomic and functional analyses.
| Gracz, Adam D; Samsa, Leigh Ann; Fordham, Matthew J et al. (2018) Sox4 Promotes Atoh1-Independent Intestinal Secretory Differentiation Toward Tuft and Enteroendocrine Fates. Gastroenterology 155:1508-1523.e10 |
