PROJECT SUMMARY The intestinal epithelium is in a dynamic equilibrium of proliferation, differentiation and apoptosis along the crypt-villus gradient. Normal intestinal homeostasis is disturbed during states of infection, inflammation and malignant transformation (adenomatous polyps, colorectal cancer). The pathogenesis of sporadic colorectal cancer involves distinct pathways with characteristic genomic and genetic alterations. Despite significant advances in leveraging our understanding of these pathways for diagnostic, prognostic, and therapeutic strategies, colorectal cancer (CRC) remains a leading cause of cancer-related mortality. This underscores a specific need to identify and understand novel, therapeutically tractable pathways in intestinal homeostasis that may drive CRC. Our work has introduced and elucidated the novel role of mRNA binding proteins in intestinal/colonic epithelial homeostasis, as well as aberrations including hyperproliferation, altered metabolism and transformation. LIN28B, an mRNA binding protein, also critical in embryonic stem cells, post- transcriptionally regulates the let-7 microRNA family and results in suppression of differentiation. In turn, Let-7 microRNAs have diverse mRNA targets, including IMP1 (Igf2 mRNA binding protein-1), another mRNA binding protein. We have demonstrated that LIN28B and IMP1 separately drive tumor-initiating cell phenotypes associated with their roles in regulating proliferation and differentiation during normal homeostasis; however, it remains unclear if LIN28B-mediated hyperproliferation, altered differentiation, and associated tumorigenesis requires IMP1. In addition, it remains unknown if the tumor promoting or metastatic roles of IMP1 depend entirely on its regulation by Let-7 downstream of LIN28B. Our overarching hypothesis of this proposal is that LIN28B and IMP1 comprise cooperative roles in controlling post-transcriptionally the pathways (some known) associated with epithelial cell fate, which may favor malignant transformation. We will pursue this hypothesis through the following interrelated Specific Aims: Aim 1: To evaluate the interdependence of LIN28B and IMP1 in proliferation, differentiation, and malignant transformation in vivo. Aim 2: To determine and validate common and divergent pathways mediated by LIN28B and IMP1 via RNA-Sequencing (“transcriptome”) and ribosome profiling (“translatome”) in vivo. Aim 3: To evaluate Let-7 dependent regulation of IMP1 to promote hyperproliferation and metastasis.