Exosomes are deemed to be an important tool of intercellular communicators in cancer cells. Our study investigated the role of PRR34 long non-coding RNA antisense RNA 1 (PRR34-AS1) in regulating exosome secretion in hepatocellular carcinoma (HCC) cells.Quantitative real-time polymerase chain reaction (RT-qPCR) analyzed the expression of PRR34-AS1. We assessed the function of PRR34-AS1 on the biological changes of THLE-3 cells and HCC cells. The downstream interaction between RNAS was assessed by mechanistic experiments.PRR34-AS1 expression was upregulated in HCC cells in comparison to THLE-3 cells. PRR34-AS1 depletion repressed HCC cell proliferation, migration and invasion as well as EMT phenotype, while PRR34-AS1 up-regulation accelerated the malignant phenotypes of THLE-3 cells. PRR34-AS1 recruited DDX3X to stabilize the mRNA level of exosomal protein Rab27a. Moreover, PRR34-AS1 facilitated the malignant phenotypes of THLE-3 cells by elevating Rab27a expression to promote the exosome secretion of VEGF and TGF-β in HCC cells.The current study revealed a novel function of PRR34-AS1 in accelerating exosome secretion in HCC cells and offered an insight into lncRNA function in the regulation of tumor cell biology.© 2022. The Author(s).