Sixteen carbazole alkaloids from Clausena vestita D. D. Tao were extracted, and their anti-tumor activities were evaluated. Among the extracts, Clauszoline-I exhibited an obvious growth inhibitory activity against several cancer cell lines through its ability to induce cell cycle arrest in the S and G2/M phases. A dramatic morphologic change with decreased F-actin staining and RhoA activity was found in Clauszoline-I treated HepG2 cells, in which the phosphorylation of PKCδ (Ser643) was inhibited. Our results indicated that induction cell cycle arrest by Clauszoline-I might be achieved by decreasing the RhoA activity via the inhibition of PKCδ phosphorylation. Display omitted ► Clauszoline-I is a carbazole alkaloid from Clausena vestita D. D. Tao. ► It induced cell cycle arrest and Chk1 redistribution from nucleus into cytoplasm. ► It caused morphological changes via repression of RhoA activation. ► It inhibited PKCδ (Ser643) phosphorylation.