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http://hdl.handle.net/11375/20628
Title: | Investigating the Role of the RNA-Binding Protein MUSASHI-2 (MSI2) in Normal Hematopoiesis and Leukemia |
Authors: | Holzapfel, Nicholas |
Advisor: | Hope, Kritstin |
Department: | Biochemistry |
Keywords: | RNA-Binding Protein, Musashi, Hematopoiesis |
Publication Date: | 2016 |
Abstract: | Musashi-2 (MSI2), a member of the Musashi family of RNA-binding proteins, is thought to play a critical role in the maintenance of stem cell populations and in the formation of aggressive tumours. Multiple studies indicate that MSI2 plays an important role in the maintenance of hematopoietic stem cell (HSC) populations and recent studies in humans identify MSI2 as an independent prognostic factor for overall survival in patients with Acute Myeloid Leukemia (AML). Importantly, though correlative studies implicate MSI2 as a contributor to aggressive disease in human AML, no study to date has attempted to analyze the functional role of MSI2 in primary human AML samples. Furthermore, though MSI2 is critical for the maintenance of HSCs, the mechanisms through which MSI2 functions are unknown. The work presented in this thesis elucidates the biochemical mechanisms through which MSI2 functions and examines the functional role of MSI2 in human AML. Using a lentiviral-mediated shRNA knockdown of MSI2, I demonstrate that MSI2 is critical for the maintenance of human AML. A loss of MSI2 greatly impairs the ability of AML samples to maintain disease in a xenotransplantation assay. MSI2 is an RNA binding protein that is thought to repress the translation of target mRNAs in the cytoplasm and prevent the maturation of microRNAs (miRNAs) in the nucleus. The targets of MSI2 are believed to be potent regulators of stem-ness and dysregulation of these targets could very well contribute to neoplastic transformation. Cross-linking immunoprecipitation followed by next generation sequencing (CLIP-Seq), revealed the RNA binding properties of MSI2 and the RNA targets bound by MSI2. To identify novel MSI2 protein interactors, the MSI2 locus was endogenously tagged with the promiscuous biotin ligase BirA* and subjected to BioID analysis. When compared to appropriate controls, we were able to robustly identify proteins that associate with MSI2. The analysis of one of these protein binding partners, Insulin-like growth factor 2 mRNA binding protein 2 (IGF2BP2) reveals a critical role in the normal function of HSCs. |
URI: | http://hdl.handle.net/11375/20628 |
Appears in Collections: | Open Access Dissertations and Theses |
Files in This Item:
File | Description | Size | Format | |
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Holzapfel_Nicholas_T_2016September_PhD.pdf | 18.61 MB | Adobe PDF | View/Open |
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