Malignant glioma is a highly aggressive cancer, known as one of the most dangerous types of primary brain tumor occurring in the central nervous system (CNS). Septin 9 (SEPT9) has been involved in tumor growth. However, its exact roles in regulating glioma development have not been fully understood. In the present study, we found that SEPT9 expression levels were markedly up-regulated in glioma tissues and cell lines. High expression of SEPT9 predicted a poor overall survival in patients with glioma. SEPT9 knockdown significantly reduced the proliferation, migration and invasion of glioma cells. Moreover, epithelial-mesenchymal transition (EMT) markers, including N-cadherin, matrix metalloproteinase-9 (MMP9), Vimentin and Twist, were significantly reduced by SEPT9 knockdown; however, the expression of E-cadherin was elevated by SEPT9 silence. This EMT process in glioma cells was dependent on the expression transforming growth factor (TGF)-β1. In addition, the clinical analysis suggested that SEPT9 gene expression had a positive correlation with TGF-β1 in patients with primary glioma at different grades. Furthermore, knockdown of SEPT9 significantly reduced the glioma progression in vivo. The anti-metastasis regulated by the knockdown of SEPT9 was further confirmed in mouse model, as evidenced by the reduced number of lung metastatic nodules. Our results supported that reducing SEPT9 expression could inhibit glioma progression through the suppression of EMT induced by TGF-β1.