Polydatin, a natural analogue of resveratrol, has many biological activities. The better bioavailability of polydatin than resveratrol makes it an ideal candidate for therapy. Polydatin has protective effects against various diseases (cardiovascular, neurological, inflammatory, etc.) including cancer. However, its mechanism of action has not been fully established. Therefore, the present study was initiated to explore the mechanism/s associated with chemotherapeutic effects of polydatin in in vitro using lung cancer A549 cells. The effects of polydatin on cell proliferation and metastasis were assessed using various parameters like MTT, colony formation, DNA damage, apoptosis, and wound healing. Polydatin treatment reduced the proliferation of A549 cells by inducing DNA damage and cell cycle arrest in a concentration-dependent manner. The inhibition of cell proliferation was induced by dual mechanism of senescence and apoptosis. Proteins involved in various pathways were studied using western blotting and immunocytochemistry. Interestingly, senescent and apoptotic cells induced a differential bystander response (proliferative/toxic) in naïve A549 cells. Our results show that polydatin can induce both senescence and apoptosis in A549 cells in a concentration-dependent manner and the differential bystander effects induced by polydatin are regulated by mTOR pathway.