Abstracts C1-C3

Category C.  Cancer Biology/ Molecular Biology /Developmental Biology


C1.  Investigating a Role for Musashi-1 in Intestinal Development and Organ Size Regulation

Thelma T. Chiremba1, Liang Xu1, Kristi L. Neufeld1, 2

1Department of Molecular Biosciences, University of Kansas, Lawrence, KS, USA;
2Department of Cancer Biology, University of Kansas Medical Center, Kansas City, KS,USA

Proper organ size depends on the coordinated balance between cell proliferation and cell death. The dysregulation of organ size control is associated with various human pathologies, including cancer and degenerative disorders. As such, identifying and characterizing the molecular regulators of organ size can illuminate potential therapeutic targets for diseases that are associated with aberrant tissue growth. Our lab is interested in Musashi-1 (Msi-1), an RNA-binding protein that is overexpressed in a considerable subset of colorectal cancers. Msi-1 regulates target mRNA translation by binding to the 3’-untranslated region (3’-UTR) and inhibiting initiation of translation. Known targets of Msi-1, including p21, Adenomatous polyposis coli (Apc), and Numb, are involved in the regulation of cell proliferation, cell differentiation, and cell death. These processes are essential in tissue development and in the maintenance of tissue homeostasis; thus, the translational control conferred by Msi-1 might be crucial in organ size regulation. In order to understand the in vivo regulatory role of Msi-1 during development, we generated a mouse model that allowed regulated and inducible Msi-1 overexpression. Here we provide evidence that the ubiquitous overexpression of Msi-1 in young transgenic mice decreases overall body size and results in premature death. These Msi-1-overexpressing mice failed to maintain normal organ to body proportions; in particular, their colon and small intestines were larger when compared to wild-type mice. Our findings thus far implicate Msi-1 in the regulation of intestinal organ size during early mammalian development. Current studies are aimed at elucidating mechanisms through which Msi-1 controls intestinal tissue development.



C2.  Increased Levels of APC in Goblet Cells Linked to Endoplasmic Reticulum Stress and Non-O-Glycosylated Muc2 in Ulcerative Colitis Patients

Christian Gomez, Maged Zeineldin, and Kristi L. Neufeld

Department of Molecular Biosciences, University of Kansas, Lawrence, KS

Mutation of the tumor suppressor gene Adenomatous Polyposis Coli (APC) is an initiating event in >80% of all colorectal cancers. While best known as an antagonist of the Wnt signaling pathway, other functions of APC have yet to be fully investigated. We have found that there are increased levels of APC in a subset of intestinal Goblet cells of both mice and humans. Goblet cells produce and secrete mucins, which serve to protect the intestinal epithelium from pathogens and mechanical damage.  The number of intestinal Goblet cells showing high levels of APC increased in mice treated with Dextran Sulfate Sodium (DSS) to induce colitis as well as in human inflammatory bowel disease patients with either Crohn’s disease or Ulcerative Colitis (UC). The goblet cells in these ulcerative colitis colon crypts also have non-O-glycosylated Muc2 present, indicating perturbation of normal goblet cell function. Our lab has previously generated a mouse model with compromised nuclear import of APC (ApcmNLS/mNLS mice). We found that ApcmNLS/mNLS mice treated with DSS showed significantly decreased colonic expression of the mucin Muc2 than treated wildtype mice, while the Muc2 that is produced shows reduced glycosylation. Muc2 is a highly glycosylated 5179 amino acid protein secreted exclusively by Goblet cells and is predominantly responsible for making up the two intestinal mucus layers Defects in the production of Muc2 are linked to spontaneous colitis and inflammation which can increase the risk of colon cancer. Muc2 production is tightly controlled as its overproduction or accumulation could lead to ER stress and result in the activation of the unfolded protein response (UPR). Accumulation of non-O-glycosylated Muc2 has also been linked to activation of the UPR in UC patients. Here we show that high levels of APC in patients with UC correlate with the accumulation of non-O-glycosylated Muc2 and ER stress.



C3.  Suppression of intestinal tumorigenesis in Apc mutant mice by Musashi-1 deletion

Andy R. Wolfe1, Amanda Ernlund1, William McGuinness1, Carl Lehmann1, Kaitlyn Carl1, Nicole Balmaceda1, Kristi L. Neufeld1,2

1Department of Molecular Biosciences, University of Kansas
2Department of Cancer Biology, University of Kansas Medical Center

The RNA-binding protein Musashi 1 (Msi-1) regulates target mRNA translation and is implicated in colon tumorigenesis.  We previously identified the tumor suppressor Adenomatous Polyposis Coli (Apc), a Wnt pathway antagonist, as a Msi-1 target.  Truncation of Apc initiates ~80% of all colorectal cancers and Msi-1 levels are high in colon tumors. In cultured colon cells, Msi-1 blocks Apc translation and Apc indirectly inhibits transcription of Msi-1, a Wnt target gene. Based on these findings, we previously proposed an Apc/Msi-1 double negative feedback loop and also that Msi-1 might make a good anti-cancer therapeutic target. Here, we tested whether the Msi-1 upregulation that results from Apc loss contributes to colorectal tumorigenesis by eliminating Msi-1 with a germline deletion in two Apc-mutant mouse models of intestinal cancer. Both ApcMin and Apc1322T mice showed a dramatic reduction in intestinal polyp number if they lacked Msi-1. This result provides genetic evidence that Msi-1 plays a key role in intestinal tumorigenesisMoreover, intestinal epithelial cells from Apc+/+ mice lacking Msi-1 displayed more Apc, and less b-catenin protein and decreased Wnt target gene mRNA than wildtype mice. This evidence of altered Apc/Msi-1 double negative feedback in mice lacking Msi-1 provides a possible underlying mechanism for the reduced intestinal polyp numbers seen in these mice.  Finally, mice lacking Msi-1 showed defects in intestinal cell proliferation and stem cell populations, implicating these features in the polyp resistant phenotype. Together, these results provide genetic evidence that Msi-1 contributes to intestinal tumorigenesis and validate the pursuit of Msi-1 inhibitors as chemo-prevention/therapeutic agents. 



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