Prostate apoptosis response-4 (Par-4) is a leucine zipper protein linked to apoptotic cell death in prostate cancer and neuronal tissues. The leucine zipper domain of Par-4 (Leu.zip) mediates protein-protein interactions that are essential for sensitization of cells to apoptosis, and overexpression of Leu.zip blocks Par-4 activity in a dominant negative fashion. Ischemia-reperfusion-induced renal injury (IRI) is clinically important because it typically damages renal tubular epithelial cells and glomerular cells, and it is the most common cause of acute renal failure (ARF). We now report that Par-4 is expressed in renal tubule cells and that aberrant expression of Par-4 activity plays a crucial role in activating apoptotic pathways in well-characterized models of renal IRI. Increased levels of Par-4 were observed following chemical ischemia-reperfusion in HK-2 cells in vitro and in mouse renal tubular cells following bilateral clamping of renal pedicles in vivo. Inhibition of Par-4 expression by specific par-4 antisense oligonucleotides largely prevented HK-2 cell apoptosis induced by IRI. Overexpression of Par-4 in these cells exacerbated mitochondrial dysfunction and caspase activation and conferred increased sensitivity to IRI-induced apoptosis. Expression of Leu.zip, a dominant negative regulator of Par-4, largely prevented mitochondrial dysfunction and caspase activation and significantly inhibited IRI-induced apoptosis in HK-2 cells. In addition, transfection of Par-4 increased while transfection of Leu.zip decreased necrosis in HK-2 cells following prolonged IRI. These results identify Par-4 as a novel and early mediator of renal tubule cell injury following IRI and provide a potential target for developing new therapeutic strategies for renal IRI and ARF.