| Summary: | The therapeutic potential of Musashi (MSI) RNA-binding proteins, important stemness-associated gene expression regulators, remains insufficiently understood in breast cancer. This study identifies the interplay between MSI protein expression, stem cell characteristics, radioresistance, cell invasiveness and migration. MSI-1, MSI-2 and Notch pathway elements were investigated via quantitative polymerase chain reaction (qPCR) in 19 triple-negative breast cancer samples. Measurements were repeated in MDA-MB-231 cells after MSI-1 and -2 siRNA-mediated double knockdown, with further experiments performed after <i>MSI</i> silencing. Flow cytometry helped quantify expression of CD44 and leukemia inhibitory factor receptor (LIFR), changes in apoptosis and cell cycle progression. Proliferation and irradiation-induced effects were assessed using colony formation assays. Radiation-related proteins were investigated via Western blots. Finally, cell invasion assays and digital holographic microscopy for cell migration were performed. MSI proteins showed strong correlations with Notch pathway elements. <i>MSI</i> knockdown resulted in reduction of stem cell marker expression, cell cycle progression and proliferation, while increasing apoptosis. Cells were radiosensitized as radioresistance-conferring proteins were downregulated. However, <i>MSI</i>-silencing-mediated LIFR downregulation resulted in enhanced cell invasion and migration. We conclude that, while <i>MSI</i> knockdown results in several therapeutically desirable consequences, enhanced invasion and migration need to be counteracted before knockdown advantages can be fully exploited.
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