Medulloblastoma-associated DDX3X mutants are oncogenic having a defect in translation-promoting activity but functional in stress granule formation and interferon signaling

Abstract

DDX3X, a DEAD box-containing RNA helicase, is known to play diverse roles in RNA metabolism, stress response, innate immunity, and cancer. Medulloblastoma is the single most common malignant brain tumor in children. DDX3X is recurrently mutated in the WNT and SHH subgroups of medulloblastoma. CRISPR-Cas9 mediated DDX3X knockout was successful in the HEK293FT cells but generated only non-truncating indels in the medulloblastoma cells suggesting DDX3X is necessary for the viability of the cells. Downregulation of DDX3X expression using shRNA also brought about a considerable reduction in proliferation, clonogenic potential, and anchorage-independent growth of the medulloblastoma cells. Thus, DDX3X expression was found to be essential for the survival, growth, and malignant potential of the medulloblastoma cells consistent with the non-truncating nature of medulloblastoma-associated DDX3X mutations. The medulloblastoma-associated DDX3X mutants were found to be defective in their ability to drive the translation of mRNAs with complex 5-UTR that is dependent on the ATP-dependent helicase activity of DDX3X. These helicase defective DDX3X mutants could restore the expression of interferon signaling genes and malignant potential lost upon DDX3X knockdown in medulloblastoma cells. Their N-terminal domain is intact and was found to be functional in stress granule formation. DDX3X mutants upregulated expression of malignancy-related genes suggesting tumor suppressive role for the helicase activity in the medulloblastoma pathogenesis. Inhibitors of the N-terminal domain of DDX3X which is essential for the viability of medulloblastoma cells could have therapeutic potential in the treatment of WNT and SHH subgroup medulloblastomas.