Rhus verniciflua Stokes has been widely used as a traditional medicinal plant with a variety of pharmacological activities. We investigated the mechanisms involved in mediating the effects of Rhus verniciflua Strokes (R. verniciflua) extract in human chronic myelogenous leukemia K562 cells, including caspase-dependent apoptotic pathways related to cell-cycle arrest, as well as the inhibition of nuclear factor NF-κB activation and upregulation of the mitogen-activated protein kinase (MAPK) signaling pathway. R. verniciflua extract suppressed the abnormal cellular proliferation of K562 cells in a dose- and time‑dependent manner and increased the quantitative proportions of cells involved in the early and late process of apoptosis. Furthermore, R. verniciflua extract significantly mediated the mRNA levels of pro-apoptotic and anti-apoptotic regulators, such as Bcl-2, Bax, Mcl-1 and survivin in apoptotic cells. Particularly, the treatment of K562 cells with R. verniciflua extract augmented the caspase‑3 activity and increased the expression of caspase‑3 protein, while co-treatment with R. verniciflua extract and the permeant pan‑caspase inhibitor Z-VAD-FMK and caspase‑3 inhibitor Z-DEVD-FMK inversely enhanced the proliferation of K562 cells. The extract of R. verniciflua inhibited the activation of NF-κB and the phosphorylation of ERK. Collectively, these results indicated that the extract of R. verniciflua inhibited the proliferation of human chronic myelogenous leukemia K562 cells by activating the apoptotic process via caspase‑3 overexpression and the regulation of the NF-κB and MAPK signaling.