Glioblastoma (GBM), or grade IV astrocytoma, is a malignant brain cancer that contains subpopulations of proliferative and invasive cells that coordinately drive primary tumor growth, progression, and recurrence after therapy. Here, we have analyzed functions for megalencephalic leukoencephalopathy with subcortical cysts 1 (Mlc1), an eight-transmembrane protein normally expressed in perivascular brain astrocyte end feet that is essential for neurovascular development and physiology, in the pathogenesis of GBM. We show that Mlc1 is expressed in human stem-like GBM cells (GSCs) and is linked to the development of primary and recurrent GBM. Genetically inhibiting MLC1 in GSCs using RNAi-mediated gene silencing results in diminished growth and invasion in vitro as well as impaired tumor initiation and progression in vivo. Biochemical assays identify the receptor tyrosine kinase Axl and its intracellular signaling effectors as important for MLC1 control of GSC invasive growth. Collectively, these data reveal key functions for MLC1 in promoting GSC growth and invasion, and suggest that targeting the Mlc1 protein or its associated signaling effectors may be a useful therapy for blocking tumor progression in patients with primary or recurrent GBM.