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Abstract Background Long noncoding RNA POU class 3 homeobox 3 (POU3F3) is upregulated in esophageal squamous-cell carcinomas. The present study aimed to investigate the role of POU3F3 in non-small cell lung cancer (NSCLC). Methods A total of 80 patients with NSCLC (adenocarcinoma) admitted by Guangdong Provincial Hospital of Integrated Traditional Chinese and Western Medicine between May 2016 and May 2018 were enrolled in this study. All patients were diagnosed by histopathological approaches. Expression levels of POU3F3 and microRNA-30d-5p (miR-30d-5p) in cancer and non-tumor tissues from these NSCLC patients were determined by qRT-PCR. Cell transfections were performed to assess interactions between miR-30d-5p and POU3F3. Cell proliferation, Transwell migration and invasion assays were performed to investigate the role of miR-30d-5p and POU3F3 in the regulation of cell proliferation, migration and invasion. Results POU3F3 was upregulated, while miR-30d-5p was downregulated in cancer tissues than in adjacent healthy tissues of NSCLC patients. Correlation analysis showed that expression levels of POU3F3 and miR-30d-5p were inversely correlated in tumor tissues. Overexpression of miR-30d-5p did not affect the expression of POU3F3, while overexpression of POU3F3 resulted in the suppression of miR-30d-5p in NSCLC cell lines. Overexpression of POU3F3 mediated enhanced proliferation, migration and invasion of NSCLC cells. In addition, overexpression of miR-30d-5p played an opposite role and attenuated the effects of overexpressing POU3F3 on cancer cell proliferation, migration and invasion. Conclusions POU3F3 might positively regulate NSCLC cell proliferation, migration and invasion through downregulation of miR-30d-5p.