Pediatric low-grade gliomas (PLGGs) are a heterogeneous group of tumors that collectively represent the most common childhood brain cancer. Despite favorable outcomes with surgical and adjuvant therapies, majority of patients suffer from long-term treatment-related morbidities and recurrent/inoperable disease. This necessitates a deeper understanding of PLGG biology to aid development of molecular diagnostics and low-toxicity targeted therapeutics. Hitherto, PLGGs have been defined by activating mutations that dysregulate the MAPK signaling pathway, leading to clinical testing of RAF/MAPK inhibitors for PLGGs. Interestingly, recent large-scale sequencing efforts discovered novel gene fusions in PLGGs and we identified the unique recurrent association of tumor suppressor Quaking (QKI) with distinct proto-oncogenes, MYB and RAF1, in different PLGG sub-types. We hypothesized that MYB-QKI and QKI-RAF1 function via novel oncogenic mechanisms invoking a two-hit mechanism of gain-of-function in the MYB/RAF1 oncoproteins collaborating with QKI loss of putative tumor suppressor function, describing for the first time a unique gene fusion setting involving both fusion partners with implications for therapeutic targeting. Utilizing heterologous cell model systems and in vivo mouse models, we found MYB-QKI and QKI-RAF1 are capable of driving oncogenesis. Furthermore, MYB-QKI is a specific driver mutation defining angiocentric gliomas and mediates tumorigenesis via a tri-partite mechanism: (1)MYB oncogenic activation via truncation, (2)rearrangement led enhancer translocation that drives MYB-QKI expression and (3)LOH of QKI tumor suppressor. In contrast, QKI-RAF1 drives some pilocytic astrocytomas via aberrant activation of the MAPK pathway in a QKI-dimerization dependent manner. We also found differential response to RAF targeted therapy in QKI-RAF1, compared to BRAF fusions in PLGGs, due to QKI-mediated dimerization. Hence, our study highlights distinct roles for the same gene, QKI in supporting the oncogenic functions of MYB and RAF1 in different PLGG-gene fusions. Overall, our study has uncovered distinct molecular mechanisms associated with different QKI gene fusions in PLGGs. We show that MYB-QKI is specific to angiocentric gliomas and mediates a unique oncogenic program, and with QKI-RAF1 we demonstrate how mutational context guides differential response to targeted therapy. Therefore, our study has important clinical implications on molecular diagnostics and targeted therapy for these rather understudied class of childhood brain tumors.