Prostate cancer (PCa) is a leading cause for death in men and the most commonly diagnosed malignancy globally. MicroRNA (miR)-583 expression levels have been discovered to be downregulated in recurrent PCa samples compared with non-recurrent cases. However, the precise functions and pathogenic mechanism of miR-583 in the development of PCa are vague, thus the aim of the present study was to investigate these. The expression levels of miR-583 and Janus kinase 1 (JAK1) in PCa tissues and cell lines were analyzed using reverse transcription-quantitative PCR and western blotting. The protein expression levels of phosphorylated (p)-STAT3 and STAT3 in PCa cell lines were also analyzed using western blotting. The effects of miR-583 and JAK1 on the proliferation and invasion of PCa cell lines cell lines were determined using MTT and Transwell assays, respectively. The binding interaction between miR-583 and the 3'-untranslated region of JAK1 were predicted by TargetScan, and further validated using dual luciferase reporter assays in PCa cell lines. The results revealed that the expression levels of miR-583 were downregulated, while those of JAK1 were upregulated in PCa tissues and cell lines (DU145 and PC3). The transfection with the miR-583 mimic inhibited the proliferation and invasion, as well as downregulating JAK1 and p-STAT3 protein expression levels in DU145 and PC3 cell lines. These effects were partially abolished following the overexpression of JAK1. Moreover, JAK1 was identified to be a target gene for miR-583 in DU145 and PC3 cell lines and the expression levels of miR-583 were revealed to be negatively correlated with JAK1 expression levels in PCa tissues. In conclusion, the findings of the present study suggested that miR-583 may inhibit the proliferation and invasion of PCa cells by targeting JAK1, thus providing a novel therapeutic target for patients with PCa. © 2021 Spandidos Publications. All rights reserved.