The aim of the present study was to investigate the effect of microRNA (miR)‑539 on the proliferation and migration of gastric cancer cells, and explore the underlying mechanism. Gastric cancer cell lines with high or low miR‑539 and SRY‑box 5 (SOX5) expression levels were constructed by transfection. The proliferation of gastric cancer cells was then detected by Cell Counting Kit‑8 assay and cell migration was tested by transwell assay. The results revealed low expression of miR‑539 and high expression of SOX5 in gastric cancer tissues and cells as compared with the levels in normal tissues and cells, suggesting that there was a negative correlation between miR‑539 and SOX5. Dual‑luciferase reporter experiments demonstrated that miR‑539 directly targeted SOX5. The proliferation and migration of gastric cancer cells were negatively regulated by the overexpression of miR‑539, while positively regulated by the overexpression of SOX5. Notably, SOX5 overexpression attenuated the inhibitory effect of miR‑539 on gastric cancer cells. The results suggested that SOX5 is a target gene of miR‑539, and that miR‑539 inhibits the proliferation and migration of gastric cancer cells by targeting SOX5.