Despite therapeutic advancements, there has been little improvement in the survival status of patients with oral squamous cell carcinoma (OSCC). HOX antisense intergenic RNA (HOTAIR) has been shown to be dysregulated in several cancer types. However, the roles of HOTAIR in OSCC remain largely unknown. In this study, we investigated the association of HOTAIR expression with clinicopathological features in OSCC patients and the crucial roles of HOTAIR in the modulation of tumor progression. Our results showed that HOTAIR was highly expressed both in OSCC tissue samples and cell lines compared with corresponding normal oral mucosa tissues and human oral keratinocytes. Its overexpression was positively correlated with TNM (tumor-node-metastases) stage, histological grade, and regional lymph node metastasis. The knockdown of HOTAIR by short hairpin RNA significantly decreased the migration, invasion, and epithelial-mesenchymal transition of OSCC cells in vitro. Moreover, there was a negative correlation between HOTAIR and microRNA-326 expression in OSCC tissue samples and cell lines. Luciferase reporter and loss-of-function assays revealed that HOTAIR acted as a competitive endogenous RNA effectively sponging miR-326, thereby regulating the derepression of metastasis-associated gene 2 (MTA2). Finally, the expression and clinical significance of MTA2 were analyzed in another cohort of OSCC tissue samples. High MTA2 expression was significantly correlated with clinicopathological features of advanced OSCC and poor prognosis for patients with OSCC. Collectively, HOTAIR overexpression promoted the progression of OSCC. The HOTAIR-miR-326-MTA2 axis may contribute to a better understanding of OSCC pathogenesis and be a potential therapeutic target for OSCC.© 2020 Wiley Periodicals, Inc.