Date of Award

2018

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Graduate Group

Cell & Molecular Biology

First Advisor

Anil K. Rustgi

Abstract

The intestinal epithelium spans proliferating crypts at its base to differentiated villi at the luminal surface and renews itself every 3-5 days. It maintains a dynamic equilibrium between proliferation, differentiation and apoptosis. The regulation of homeostasis, response to injury, regeneration and transformation is a complex set of dynamic processes. mRNA binding proteins (RBPs) are newly recognized regulators of intestinal homeostasis. The RBP:mRNA complexes act as rheostats of key signaling processes by regulating expression of already transcribed RNAs. Their functional effects are tissue and context dependent. The manner in which RBPs operate and their interactions with other pivotal pathways in colorectal cancer provide a framework for new insights and potential therapeutic applications. This thesis focuses upon the interplay between LIN28B and IGF2 mRNA binding protein (IMP1) in the regulation of intestinal epithelial regeneration and malignant transformation, unraveling a new perspective on these processes.

LIN28B, an mRNA binding protein, plays a critical role in regulating growth and proliferation in the intestinal epithelium. Previous work in our lab revealed that LIN28B promotes growth and tumorigenesis of the intestinal epithelium via suppression of mature let-7 miRNAs. LIN28B suppression of let-7 promotes upregulation of let-7 targets, including IMP1 (Insulin-like growth factor II mRNA-binding protein 1). Our lab has shown that transgenic mice expressing LIN28B from the mouse Vil1 promoter (Vil-Lin28b mice) have increased proliferation and tumor formation in the small intestine. IMP1 protein levels are upregulated in these mice epithelia and tumors but specific role of IMP1 in Lin28b-mediated tumorigenesis remains unknown. The current study tested the hypothesis that IMP1 may be required for LIN28B-mediated tumorigenesis and that LIN28B and IMP1 may cooperatively promote a tumor-initiating phenotype.

Additionally, IMP1 hypomorphic mice exhibit severe intestinal growth defects, yet it’s role in adult epithelium is unclear. We investigated the mechanistic contribution of epithelial IMP1 to intestinal homeostasis and repair. We evaluated IMP1 expression in Crohn’s disease patients followed by unbiased ribosome profiling in IMP1 knockout cells. We used irradiation and dextran sodium sulphate (DSS) induced colitis as injury models to evaluate regeneration in intestinal epithelium lacking IMP1.

These studies show that in the context of LIN28B overexpression, IMP1 loss led to increased tumor initiation and progression. Ribosome profiling and RNA sequencing revealed a potential role for IMP1 in negatively regulating the Wnt pathway, stem cell signature and other pathways associated with proliferation. This pro-proliferative effect with the loss of IMP1 has been previously observed in breast cancer and intestinal stroma. Additionally, IMP1 acts as a post-transcriptional regulator of gut epithelial repair post-colitis and irradiation, in part through modulation of autophagy. This study provides a new perspective on post-transcriptional regulation of autophagy as a contributing factor to the pathogenesis of inflammatory bowel disease.

In total, these studies provide new insights into the role of IMP1 in regulating homeostasis, response to injury, and tumorigenesis in the intestine. It provides evidence that IMP1 regulates the expression of its targets at both the transcriptional and translational levels.

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