KRUPPEL-LIKE FACTOR 9 INHIBITS GLIOBLASTOMA STEMNESS THROUGH GLOBAL TRANSCRIPTION REPRESSION AND INHIBITION OF INTEGRIN ALPHA 6 AND CD151
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Glioblastoma (GBM) stem cells (GSCs) represent tumor-propagating cells with stem-like characteristics (stemness) that contribute disproportionately to GBM drug resistance and tumor recurrence. Understanding the mechanisms supporting GSC stemness is important for developing novel strategies that target tumor propagation to inhibit cancer progression and improve patient survival. Krüppel-like factor 9 (KLF9) has emerged as a regulator of cell differentiation, neural development, and oncogenesis; however, the molecular basis for KLF9’s diverse contextual functions has been unclear. We establish for the first time a genome-wide map of KLF9-regulated targets in human glioblastoma stem-like cells, and show that KLF9 functions as a transcriptional repressor and thereby regulates multiple signaling pathways involved in oncogenesis and regulation of cancer stem-like phenotype. A detailed analysis of two novel KLF9 targets suggests that KLF9 inhibits glioma cell stemness by repressing expression of integrin α6 and CD151. The expression of one candidate KLF9 target gene ITGA6 coding for integrin α6 was verified to be downregulated by KLF9 in GSCs. ITGA6 transcription repression by KLF9 altered GBM neurosphere cell behavior as evidenced by reduced cell adhesion to and migration through membrane coated with the integrin α6 ligand laminin. Forced expression of integrin α6 partially rescued GBM neurosphere cells from the differentiating and adhesion/migration-inhibiting effects of KLF9. Using GBM derived neurospheres, we identified cell surface tetraspanin family member CD151 as a novel regulator of GSC stemness and tumorigenicity. CD151 was found to be iii overexpressed in GBM tumors and GBM neurospheres enriched in GSCs. Silencing CD151 inhibited neurosphere self-renewal and cell proliferation and attenuated expression of stem cell markers and drivers. Conversely, forced CD151 expression promoted neurosphere self-renewal, cell migration and expression of stemness-associated transcription factors. Additionally, targeting CD151 inhibited glioma angiogenesis and growth of GBM neurosphere derived xenografts. CD151 was found to form complexes with integrins α3, α6 and β1 in neurosphere cells and blocking the CD151-intgerin α3/α6 interaction impaired sphere formation, migration and phosphorylation of AKT, a marker of integrin signaling. These findings identify CD151 and its interactions with integrins α3 and α6 as potential therapeutic targets for depleting stem cell populations and stemness-driving mechanisms in GBM.