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Figure 1: Molecular characterization of a novel KIAA1549:BRAF fusion alteration in a sporadic pediatric pilocytic astrocytoma. (a) Axial T1-weighted 1.5-Tesla gadolinium-enhanced MRI scan reveals a cystic lesion in the cerebellum with a peripheral enhancing nodule (arrow). Hematoxylin and eosin staining demonstrates a classic pilocytic astrocytoma with compact and loose areas (b), including Rosenthal fibers (arrow) and eosinophilic granular bodies (arrowhead) (c). The tumor is composed of cells with strong GFAP expression (d) and rare Ki-67 immunoreactivity (arrowhead; (e)). Direct amplification of RNA from this tumor demonstrates a 599 bp fragment, which creates a novel fusion KIAA1549:BRAF transcript in which exon 16 of the KIAA1549 gene is joined to BRAF sequences in the middle of exon 10. The bars below the predicted amino acid sequence correspond to BRAF exon 10 (red), BRAF exon 11 (green), and KIAA1549 exon 16 (blue) (f). Immunostaining with phospho-ERK-Thr202/Tyr204 antibodies demonstrates increased ERK activation in the PA tumor (bottom panel). Normal adult human frontal lobe (NB) from an autopsy specimen was included as reference tissue in the top panel (g). Western blot demonstrates 282-fold increase in ERK activation (phospho-ERK-Thr202/Tyr204; p-ERK; Cell Signaling Technologies, catalog no. 4370S) in the tumor (PA) relative to normal human brain (NB). Total ERK is included as internal control for protein loading (h).