Background Dysregulation of microRNAs (miRNAs) was found to play crucial roles in varieties of cancers, which affect tumor proliferation and migration. MiR-27a-3p has been identified as a tumor-related miRNA in liver cancer, lung cancer, and colorectal cancer. However, the function of miR-27a-3p in triple-negative breast cancer (TNBC) and its possible molecular mechanisms have still not been elucidated. Methods QRT-PCR technique was used to detect the expression of miR-27a-3p in TNBC and normal breast cell lines or the effects of miR-27a-3p knockdown and overexpression in TNBC cell lines. Proliferation and migration were measured by CCK-8 method, colony formation, wound healing, and Transwell assays, respectively. Furthermore, we used a dual-luciferase reporter gene assay and Western blot analysis to identify GSK3β as a target of miR-27a-3p. Results In this study, we found that miR-27a-3p expression was significantly elevated in TNBC cell lines. Database analysis suggested that TNBC patients with a high expression of miR-27a-3p have poorer overall survival possibilities. Overexpression of miR-27a-3p promotes TNBC cells proliferation, colony formation, and cell migration in vitro. Nevertheless, dual-luciferase reporter result showed that miR-27a-3p directly targeted the 3ʹ-UTR regions of GSK3β mRNA and negatively regulated its expression. Lastly, we demonstrated that miR-27a-3p inactivates Wnt/β-catenin signaling pathway via targeting GSK3β. Conclusion These results indicate that expression of miR-27a-3p was highly expressed in TNBC and promoted tumor progression through attenuating GSK3β and may have a potential molecular-targeted strategy for TNBC therapy.