FAT1 modulates EMT and stemness genes expression in hypoxic glioblastoma.

Abstract

Glioblastoma (GBM) is characterized by the presence of hypoxia, stemness and local invasiveness. We have earlier demonstrated that FAT1 promotes invasiveness, inflammation and upregulates HIF-1α expression and its signaling in hypoxic GBM. Here, we have identified the role of FAT1 in regulating EMT (epithelial-mesenchymal transition) and stemness characteristics in GBM. The expression of FAT1, EMT (Snail/LOX/Vimentin/N-cad), stemness (SOX2/OCT4/Nestin/REST) and hypoxia markers (HIF-1α/VEGF/PGK1/CA9) was upregulated in ≥39% of GBM tumors (n = 31) with significant positive correlation (p ≤ 0.05) of the expression of FAT1 with LOX/Vimentin/SOX2/HIF-1α/PGK1/VEGF/CA9. Furthermore, positive correlation (p ≤ 0.01) of FAT1 with Vimentin/N-cad/SOX2/REST/HIF-1α has been observed in TCGA GBM-dataset (n = 430). Analysis of cells (U87MG/A172) exposed to severe hypoxia (0.2%O2 ) revealed elevated mRNA expression of FAT1, EMT (Snail/LOX/Vimentin/N-cad), stemness (SOX2/OCT4/Nestin/REST) and hypoxia markers (HIF-1α/PGK1/VEGF/CA9) as compared to their normoxic (20%O2 ) counterparts. FAT1 knockdown in U87MG/A172 maintained in severe hypoxia and in normoxic primary glioma cultures led to significant reduction of EMT/stemness markers as compared to controls. HIF-1α knockdown in U87MG cells markedly reduced the expression of all the EMT/stemness markers studied except for Nestin and SOX2 which were more under the influence of FAT1. This indicates FAT1 has a novel regulatory effect on EMT/stemness markers both via or independent of HIF-1α. The functional relevance of our study was corroborated by significant reduction in the number of soft-agar colonies formed in hypoxic-siFAT1 treated U87MG cells. Hence, our study for the first time reveals FAT1 as a novel regulator of EMT/stemness in hypoxic GBM and suggests FAT1 as a potential therapeutic candidate.