From the ‡ Molecular Biology of Selenium Section, Laboratory of Cancer Prevention, Center for Cancer Research, NCI, National Institutes of Health, Bethesda, Maryland 20892, §Neurobiology of Selenium, Neuroscience Research Center, Institute forExperimental Endocrinology, Charite Universita tsmedizin Berlin, 10117 Berlin, Germany, ¶ Science Applications International Corporation, Frederick Cancer Research and Development Center, Frederick, Maryland 21702, the School of Biological Sciences and Institute of Molecular Biology and Genetics, Seoul National University, Seoul 151-742, Korea, and the **Department of Biochemistry, University of Nebraska, Lincoln, Nebraska 68588

Whereas the p53 tumor suppressor protein plays a central role in cellular checkpoints that respond to damage or stress to prevent tumorigenesis, the transcriptional control of the p53 gene has remained unclear. We show that chemotherapeutic agents induce p53 transcription and that p73 or p53 transactivates endogenous p53 expression through direct binding to the p53 promoter. Silencing of p53 or p73 by RNA interference significantly suppresses p53 transcription under physiologic conditions or in response to cellular stress. Mutational analysis of the human p53 promoter localized a p53 DNA-binding site, which confers p53-or p73-dependent p53 promoter activation. Importantly, impaired p53-mediated autoregulation of p53 transcription by inducible-interfering RNA results in aberrant cell cycle regulation and suppression of p53-mediated apoptosis. Thus, a positive feedback loop regulates human p53 expression and involves p73 and p53. Disruption of p53 transcription contributes to defective checkpoint control. (Cancer Res 2006; 66 (14): 6982-9)

p53-dependent apoptosis is a major determinant of its tumor suppressor activity and can be triggered by hypoxia. No p53 target is known to be induced by p53 or to mediate p53-dependent apoptosis during hypoxia. We report that p53 can directly upregulate expression of Bnip3L, a cell death inducer. During hypoxia, Bnip3L is highly induced in wild-type p53-expressing cells, in part due to increased recruitment of p53 and CBP to Bnip3L. Apoptosis is reduced in hypoxia-exposed cells with functional p53 following Bnip3L knockdown. In vivo, Bnip3L knockdown promotes tumorigenicity of wild-type versus mutant p53-expressing tumors. Thus, Bnip3L, capable of attenuating tumorigenicity, mediates p53-dependent apoptosis under hypoxia, which provides a novel understanding of p53 in tumor suppression.

The candidate tumor suppressor KILLER/DR5 is a DNA damageinducible p53-regulated death receptor for the tumor necrosis factorrelated apoptosis-inducing ligand (TRAIL), a promising agent for cancer therapy. The majority of studies on KILLER/DR5 have been focused on its role in TRAIL-induced apoptosis. However, its contribution to the inhibition of tumor growth and its role as a determinant of chemosensitivity are poorly understood. In the present study, we have generated stable human colon cancer cell lines, in which the function of KILLER/DR5 was ablated using inducible RNA interference. Inducible silencing of KILLER/DR5 in vivo by exposure of mice to doxycycline led to accelerated growth of bioluminescent tumor xenografts and conferred resistance to the chemotherapeutic agent 5-fluorouracil. Our results suggest that KILLER/DR5 may be a critical determinant for tumorigenicity and chemosensitivity.