Peptidylarginine deiminases (PADIs) convert peptidylarginine into citrulline via posttranslational modification. One member of the family, PADI4, plays an important role in immune cell differentiation and cell death. To elucidate the participation of PADI4 in haematopoietic cell death, we examine whether inducible overexpression of PADI4 enhances the apoptotic cell death. PADI4 reduced the viability in a dose- and time-dependent manner of human leukemia HL-60 cells and human acute T leukemia Jurkat cells. The apoptosis-inducing activities were determined by nuclear condensation, DNA fragmentation, sub-G1 appearance, loss of mitochondrial membrane potential (delta psi(m)), release of mitochondrial cytochrome c into cytoplasm and proteolytic activation of caspase 9 and 3. Following PADI4 overexpression, cells arrest in G1 phase significantly before their entrance into apoptotic cell death. PADI4 increases tumor suppressor p53 and its downstream p21 to control cell cycle. In the detections of protein expression and kinase activity, all protein levels of cyclin-dependent kinases (CDKs) and cyclins are not reduced except cyclin D, however, CDK2 (G1 entry S phase) and CDK1 (G2 entry M phase) enzyme activities are inhibited by conditionally inducible PADI4. p53 also expands its other downstream Bax to induce cytochrome c release from mitochondria. According to these data, we suggest that PADI4 induces apoptosis mainly through cell cycle arrest and mitochondria-mediated pathway. Furthermore, p53 features in PADI4-induced apoptosis by increasing intracellular p21 to control cell cycle and by Bax accumulation to decline Bcl-2 function, destroy delta psi(m), release cytochrome c to cytoplasm and activate the caspase cascade.