The NR4A orphan nuclear receptors are target genes of the novel drug c1 in cancer cells and potential mediators of drug induced apoptosis

THE NR4A ORPHAN NUCLEAR RECEPTORS ARE TARGET GENES OF THE NOVEL DRUG C1 IN CANCER CELLS AND POTENTIAL MEDIATORS OF DRUG INDUCED APOPTOSIS KALA RAMASESHAN (M.Sc) A THESIS SUBMITTED FOR THE DEGREE OF DOCTOR OF PHILOSOPHY DEPARTMENT OF PHYSIOLOGY NATIONAL UNIVERSITY OF SINGAPORE 2008 ACKNOWLEDGEMENTS I wish to acknowledge my deepest gratitude and appreciation to my supervisor, Dr Patrick Tan, Group Leader at the Genome Institute of Singapore and Principal Investigator at the National Cancer Centre of Singapore, and my co-supervisor Dr Shazib Pervaiz, Professor, Department of Physiology, Yong Loo Lin School of Medicine, NUS Their encouragement and guidance and their critique has been of paramount importance in directing the course of the work leading to this dissertation and all the knowledge and skills I have gained in the process I am very grateful to all my colleagues, who have helped me in one way or another during my stay in both the labs My appreciation also goes to my parents-in-law for their immense support during this period without which this work could not have been done Special thanks belong to my husband and our two boys for all the joy they bring to my life and my parents who have always encouraged me to my best i TABLE OF CONTENTS Acknowledgements i Table of Contents ii Summary viii List of Figures ix List of tables xi Abbreviations xii List of publications xiv PART 1 INTRODUCTION Oncogenesis and the development of cancer therapies 2 Nuclear Receptors 2.1 Classification of nuclear receptors 2.2 Nuclear receptors as modular proteins 2.2.1 Nuclear receptor co-factors 2.2.2 Specificity of target genes 2.3 Orphan nuclear receptors 2.3.1 Orphan nuclear receptors as lipid sensors 2.4 NR4A nuclear receptors 2.4.1 NR4A hormone response elements 2.4.2 NR4A mediated regulation of transcription factors 10 2.5 NR4A1 10 ii 2.5.1 Regulation of NR4A1 11 2.5.2 NR4A1 in apoptosis 12 2.5.3 NR4A1 binding elements 13 2.6 NR4A2 14 2.7 NR4A3 16 2.7.1 Regulation of NR4A3 17 2.7.2 NR4A3 in apoptosis 18 Apoptosis 19 3.1 Intrinsic and extrinsic pathways of apoptosis 20 3.2 Caspases 21 3.2.1 Caspase independent cell death 22 3.3 Mitochondrial outer membrane permeabilization 23 3.4 Apoptogenic and inhibitory proteins involved in apoptosis 26 3.5 Non apoptotic mechanisms of cell death 31 Microarray technologies in interpreting drug induced signaling 34 Novel photochemotherapeutic agent C1 36 5.1 Photodynamic therapy and preactivation 36 5.2 The genesis of C1 37 PART II AIMS OF THE STUDY 40 PART III MATERIALS AND METHODS 41 iii Cell culture 41 Drugs used 41 Viability assay 41 Cell proliferation assay 42 Colony formation assay 42 Cell cycle analysis 43 Cell morphology 43 Caspase activity measurement 43 Measurement of transmembrane potential 44 10 Western blot analysis 44 10.1 Antibody list for western blotting 45 10.2 Buffers used 46 11 Transfections 48 12 RNA isolation and reverse transcription 48 13 Real time PCR 48 14 Microarray experiments and data analysis 49 PART IV RESULTS 52 Drug C1 causes non classical apoptosis in MCF-7 cells 52 1.1 Drug C1 is selective to tumor cells 52 1.2 Drug C1 causes phenotypic changes in MCF-7 cells 55 1.3 MCF-7 cells show a dose response upon C1 treatment 55 in a colony formation assay iv 1.4 Drug C1 induces caspase activation in MCF-7 cells 56 1.5 Drug C1 causes a drop in the transmembrane potential 62 of MCF-7 cells 1.6 Analysis of cell cycle profile upon C1 drug treatment 65 1.7 C1 treatment causes release of apoptogenic factors from the 68 mitochondria in MCF-7 cells 1.8 Bax translocation from cytosol to mitochondria 70 1.9 PARP1 clevage is observed with C1 treatment of MCF-7 cells 72 2.0 Microarray analysis of gene expression changes induced by C1 73 treatment on HL60 cells 2.1 Analysis of chip quality 73 2.2 Drug treated samples show greater variance than control 74 samples 2.3 Gene expression variations at each timepoint between drug 78 treated and control samples 3.0 NR4A family of transcription factors are upregulated in C1 99 mediated apoptosis 4.0 Silencing the transcript of NR4A1 and NR4A3 impact 103 apoptosis 4.1 NR4A1 and NR4A3 transcripts are effectively silenced by 103 siRNA 4.2 Silencing NR4A3 transcript affects cell viability upon low 106 dose drug treatment v 4.3 Silencing NR4A3 transcript affects BrdU assimilation upon 107 low dose drug treatment 4.4 Silencing NR4A1 transcript affects cell viability upon C1 108 drug treatment 4.5 Silencing the NR4A3 transcript affects VDAC1 translocation 109 4.5.1 The coexpression neighbourhood of NR4A3 includes 109 VDAC1 4.5.2 VDAC1 protein levels and transcript levels upon C1 113 treatment 4.5.3 Silencing the NR4A3 transcript decreases VDAC1 113 translocation 4.6 Silencing NR4A3 does not affect PARP1 clevage, Bax 118 translocation or AIF translocation 4.7 Silencing NR4A3 does not affect jun levels 118 PART V DISCUSSION 122 Microarray technology and the uncovering of drug induced 122 signal transduction pathways Drug C1 is a potent inducer of apoptosis 125 NR4A family members are important in drug response 127 3.1 NR4A transcripts are short lived 127 3.2 NR4A transcript levels modulate response to drugs 128 vi 3.3 Interaction with mitochondrial proteins may channel 131 apoptotic response of NR4A family members PART VI CONCLUSIONS 134 PART VII REFERENCES 135 vii SUMMARY Previous work has shown that photoactivation of lipophilic agent merocyanine 540 generates a mixture of photoproducts (pMC540) that selectively induce cell death in human leukemia, lymphoma, and a variety of other tumor cell types in vitro and in vivo (Gulliya et al., 1994; Pervaiz et al., 1998) Earlier work has also shown that the photoproduct C1 causes activation of caspases, drop in transmembrane potential and release of cytochrome c from the mitochondria (Pervaiz et al., 1999) However, the signal transduction pathway that leads to cell death has not been elucidated As drugs may cause multiple effectors to come into play, it is essential to characterize the different transcription factors and pathways they induce for a comprehensive account of the mechanism of drug action The present study was designed to decipher the mechanism of C1 mediated cell death A high throughput method was used and a microarray analysis was performed to study the effect of drug C1 at various time points upon HL60 cells (a human promyelocytic leukemia cell line) The analysis showed that a large number of transcripts are upregulated in the early time points (table1) including the orphan nuclear receptor NR4A3 This study has validated here by real time PCR the upregulation of the orphan nuclear receptor NR4A3, and also NR4A1 which is a member of the same family of receptors This study characterizes the role of nuclear receptors NR4A1 and NR4A3 in drug C1 mediated apoptosis and has identified the functional relevance of the increase in the transcript level of NR4A1 and NR4A3 This thesis shows here that in MCF-7 breast cancer cells, silencing NR4A3 has an impact on its reponse to low dose drug treatment – silencing NR4A3 leads to attenuated response to drug C1 It also finds that silencing NR4A1 leads to an even greater effect of attenuation of cell death The results thus point to the importance of NR4A1 and NR4A3 in drug mediated apoptosis Results from this work also propose a mechanism of NR4A3 action – our observation that silencing NR4A3 leads to a decrease in the levels of VDAC1 (a major outer mitochondrial membrane protein) in the mitochondrial enriched fraction indicates that NR4A3’s effect on drug mediated apoptosis may involve signal transduction through VDAC1 - which has been implicated with a role in apoptosis (Elinder et al., 2005; Liu et al., 2006; Zaid et al., 2005) We find that the decrease in VDAC1 protein levels in the mitochondria upon NR4A3 silencing corresponds to abrogation of apoptosis, but only when the drug dosage is low At higher doses of the drug the silencing of NR4A3 and the subsequent lowering of VDAC1 levels in mitochondria not protect from cell death possibly because the overwhelming response from other signal transduction pathways render VDAC1 levels inconsequential Our findings suggest that in MCF-7 cells triggered by drug C1 there may be interaction of NR4A3 with the VDAC1 protein of the mitochondria viii LIST OF FIGURES Introduction Figure Caspase dependant and Caspase-independent routes to cell death 29 Figure Structure and components of the Permeability Transition Pore Complex 30 Results Figure Effect of drug C1 on cell survival of various cell types 54 Figure Morphological and long term effects of drug C1 on MCF-7 cells 59 Figure Drug C1 increases Caspase activity in MCF-7 cells upon high dose drug treatment 60 Figure Drug C1 does not cause change in activity of Caspases 2, 6, 8, 61 Figure Transmembrane potential changes following C1 treatment in MCF-7 cells 64 Figure Cell cycle analysis of MCF-7 cells treated with drug C1 shows DNA degradation at 72hrs but not earlier 67 Figure C1 drug treatment causes apoptotic response including PARP1 cleavage and translocation of apoptogenic factors to or from the mitochondria 71 Figure Analysis of variance test indicates that there is more changes in transcript levels in the drug treated samples than in control samples 75 Figure t-test results show that there are more changes in transcript levels at later time points in C1 drug treated samples than in controls 76 Figure 10 Drug C1 transiently increases transcription of NR4A3 in MCF-7 cells 100 ix Degterev, A., Boyce, M., and Yuan, J (2003) A decade of caspases Oncogene 22, 85438567 Degterev, A., Huang, Z., Boyce, M., Li, Y., Jagtap, P., Mizushima, N., Cuny, G D., Mitchison, T J., Moskowitz, M A., and Yuan, J (2005) Chemical inhibitor of nonapoptotic cell death with therapeutic potential for ischemic brain injury Nat Chem Biol 1, 112-119 Degterev, A., and Yuan, J (2008) Expansion and evolution of cell death programmes Nat Rev Mol Cell Biol 9, 378-390 DeRisi, J., Penland, L., Brown, P O., Bittner, M L., Meltzer, P S., Ray, M., Chen, Y., Su, Y A., and Trent, J M (1996) Use of a cDNA microarray to analyse gene expression patterns in human cancer Nat Genet 14, 457-460 Dermietzel, R., Hwang, T K., Buettner, R., Hofer, A., Dotzler, E., Kremer, M., Deutzmann, R., Thinnes, F P., Fishman, G I., Spray, D C., and et al (1994) Cloning and in situ localization of a brain-derived porin that constitutes a large-conductance anion channel in astrocytic plasma membranes Proc Natl Acad Sci U S A 91, 499-503 Ditsworth, D., Zong, W X., and Thompson, C B (2007) Activation of poly(ADP)ribose polymerase (PARP-1) induces release of the pro-inflammatory mediator HMGB1 from the nucleus J Biol Chem 282, 17845-17854 Elinder, F., Akanda, N., Tofighi, R., Shimizu, S., Tsujimoto, Y., Orrenius, S., and Ceccatelli, S (2005) Opening of plasma membrane voltage-dependent anion channels (VDAC) precedes caspase activation in neuronal apoptosis induced by toxic stimuli Cell Death Differ 12, 1134-1140 Essmann, F., Engels, I H., Totzke, G., Schulze-Osthoff, K., and Janicke, R U (2004) Apoptosis resistance of MCF-7 breast carcinoma cells to ionizing radiation is independent of p53 and cell cycle control but caused by the lack of caspase-3 and a caffeine-inhibitable event Cancer Res 64, 7065-7072 Evans, R M (2005) The nuclear receptor superfamily: a rosetta stone for physiology Mol Endocrinol 19, 1429-1438 Fischer, U., Janicke, R U., and Schulze-Osthoff, K (2003) Many cuts to ruin: a comprehensive update of caspase substrates Cell Death Differ 10, 76-100 Forman, B M., Umesono, K., Chen, J., and Evans, R M (1995) Unique response pathways are established by allosteric interactions among nuclear hormone receptors Cell 81, 541-550 139 Fu, Y., Luo, L., Luo, N., Zhu, X., and Garvey, W T (2007) NR4A orphan nuclear receptors modulate insulin action and the glucose transport system: potential role in insulin resistance J Biol Chem 282, 31525-31533 Galluzzi, L., and Kroemer, G (2007) Mitochondrial apoptosis without VDAC Nat Cell Biol 9, 487-489 Garneau, N L., Wilusz, J., and Wilusz, C J (2007) The highways and byways of mRNA decay Nat Rev Mol Cell Biol 8, 113-126 Giguere, V (1999) Orphan nuclear receptors: from gene to function Endocr Rev 20, 689-725 Glass, C K., and Ogawa, S (2006) Combinatorial roles of nuclear receptors in inflammation and immunity Nat Rev Immunol 6, 44-55 Green, D R., and Kroemer, G (2004) The pathophysiology of mitochondrial cell death Science 305, 626-629 Gronemeyer, H., Gustafsson, J A., and Laudet, V (2004) Principles for modulation of the nuclear receptor superfamily Nat Rev Drug Discov 3, 950-964 Guan, X., Madabushi, A., Chang, D Y., Fitzgerald, M E., Shi, G., Drohat, A C., and Lu, A L (2007) The human checkpoint sensor Rad9-Rad1-Hus1 interacts with and stimulates DNA repair enzyme TDG glycosylase Nucleic Acids Res 35, 6207-6218 Gulliya, K S., and Pervaiz, S (1989) Elimination of clonogenic tumor cells from HL-60, Daudi, and U-937 cell lines by laser photoradiation therapy: implications for autologous bone marrow purging Blood 73, 1059-1065 Gulliya, K S., Pervaiz, S., Dowben, R M., and Matthews, J L (1990) Tumor cell specific dark cytotoxicity of light-exposed merocyanine 540: implications for systemic therapy without light Photochem Photobiol 52, 831-838 Gulliya, K S., Sharma, R K., Matthews, J L., Benniston, A C., Harriman, A., and Nemunaitis, J J (1994) In vitro and in vivo growth suppression of MCF-7 human breast cancer by novel photoproducts and tamoxifen Cancer 74, 1725-1732 Hanahan, D., and Weinberg, R A (2000) The hallmarks of cancer Cell 100, 57-70 Hardy, K., Mansfield, L., Mackay, A., Benvenuti, S., Ismail, S., Arora, P., O'Hare, M J., and Jat, P S (2005) Transcriptional networks and cellular senescence in human mammary fibroblasts Mol Biol Cell 16, 943-953 140 Hazel, T G., Misra, R., Davis, I J., Greenberg, M E., and Lau, L F (1991) Nur77 is differentially modified in PC12 cells upon membrane depolarization and growth factor treatment Mol Cell Biol 11, 3239-3246 Hazel, T G., Nathans, D., and Lau, L F (1988) A gene inducible by serum growth factors encodes a member of the steroid and thyroid hormone receptor superfamily Proc Natl Acad Sci U S A 85, 8444-8448 Heinzel, T., Lavinsky, R M., Mullen, T M., Soderstrom, M., Laherty, C D., Torchia, J., Yang, W M., Brard, G., Ngo, S D., Davie, J R., et al (1997) A complex containing NCoR, mSin3 and histone deacetylase mediates transcriptional repression Nature 387, 4348 Hirpara, J L., Clement, M V., and Pervaiz, S (2001) Intracellular acidification triggered by mitochondrial-derived hydrogen peroxide is an effector mechanism for drug-induced apoptosis in tumor cells J Biol Chem 276, 514-521 Holler, N., Zaru, R., Micheau, O., Thome, M., Attinger, A., Valitutti, S., Bodmer, J L., Schneider, P., Seed, B., and Tschopp, J (2000) Fas triggers an alternative, caspase-8independent cell death pathway using the kinase RIP as effector molecule Nat Immunol 1, 489-495 Hong, C Y., Park, J H., Ahn, R S., Im, S Y., Choi, H S., Soh, J., Mellon, S H., and Lee, K (2004) Molecular mechanism of suppression of testicular steroidogenesis by proinflammatory cytokine tumor necrosis factor alpha Mol Cell Biol 24, 2593-2604 Inaoka, Y., Yazawa, T., Uesaka, M., Mizutani, T., Yamada, K., and Miyamoto, K (2008) Regulation of NGFI-B/Nur77 gene expression in the rat ovary and in leydig tumor cells MA-10 Mol Reprod Dev 75, 931-939 Inuzuka, H., Tokumitsu, H., Ohkura, N., and Kobayashi, R (2002) Transcriptional regulation of nuclear orphan receptor, NOR-1, by Ca(2+)/calmodulin-dependent protein kinase cascade FEBS Lett 522, 88-92 Janicke, R U., Sprengart, M L., Wati, M R., and Porter, A G (1998) Caspase-3 is required for DNA fragmentation and morphological changes associated with apoptosis J Biol Chem 273, 9357-9360 Jiang, C., Wan, X., He, Y., Pan, T., Jankovic, J., and Le, W (2005) Age-dependent dopaminergic dysfunction in Nurr1 knockout mice Exp Neurol 191, 154-162 Kagawa, S., Gu, J., Honda, T., McDonnell, T J., Swisher, S G., Roth, J A., and Fang, B (2001) Deficiency of caspase-3 in MCF7 cells blocks Bax-mediated nuclear fragmentation but not cell death Clin Cancer Res 7, 1474-1480 141 Kaufmann, S H., Desnoyers, S., Ottaviano, Y., Davidson, N E., and Poirier, G G (1993) Specific proteolytic cleavage of poly(ADP-ribose) polymerase: an early marker of chemotherapy-induced apoptosis Cancer Res 53, 3976-3985 Ke, N., Claassen, G., Yu, D H., Albers, A., Fan, W., Tan, P., Grifman, M., Hu, X., Defife, K., Nguy, V., et al (2004) Nuclear hormone receptor NR4A2 is involved in cell transformation and apoptosis Cancer Res 64, 8208-8212 Khwaja, A., and Tatton, L (1999) Resistance to the cytotoxic effects of tumor necrosis factor alpha can be overcome by inhibition of a FADD/caspase-dependent signaling pathway J Biol Chem 274, 36817-36823 Kirkbride, K C., Townsend, T A., Bruinsma, M W., Barnett, J V., and Blobe, G C (2008) Bone morphogenetic proteins signal through the transforming growth factor-beta type III receptor J Biol Chem 283, 7628-7637 Kitagawa, H., Ray, W J., Glantschnig, H., Nantermet, P V., Yu, Y., Leu, C T., Harada, S., Kato, S., and Freedman, L P (2007) A regulatory circuit mediating convergence between Nurr1 transcriptional regulation and Wnt signaling Mol Cell Biol 27, 74867496 Kokoszka, J E., Waymire, K G., Levy, S E., Sligh, J E., Cai, J., Jones, D P., MacGregor, G R., and Wallace, D C (2004) The ADP/ATP translocator is not essential for the mitochondrial permeability transition pore Nature 427, 461-465 Koltsova, E K., Wiest, D L., and Vavilova, T P (2007) Transcription factors NFAT2 and Egr1 cooperatively regulate the maturation of T-lymphoma in vitro Biochemistry (Mosc) 72, 954-961 Kroemer, G., Galluzzi, L., and Brenner, C (2007) Mitochondrial membrane permeabilization in cell death Physiol Rev 87, 99-163 Labelle, Y., Zucman, J., Stenman, G., Kindblom, L G., Knight, J., Turc-Carel, C., Dockhorn-Dworniczak, B., Mandahl, N., Desmaze, C., Peter, M., and et al (1995) Oncogenic conversion of a novel orphan nuclear receptor by chromosome translocation Hum Mol Genet 4, 2219-2226 Lammi, J., Rajalin, A M., Huppunen, J., and Aarnisalo, P (2007) Cross-talk between the NR3B and NR4A families of orphan nuclear receptors Biochem Biophys Res Commun 359, 391-397 Laudet, V., Hanni, C., Coll, J., Catzeflis, F., and Stehelin, D (1992) Evolution of the nuclear receptor gene superfamily Embo J 11, 1003-1013 Lavinsky, R M., Jepsen, K., Heinzel, T., Torchia, J., Mullen, T M., Schiff, R., Del-Rio, A L., Ricote, M., Ngo, S., Gemsch, J., et al (1998) Diverse signaling pathways 142 modulate nuclear receptor recruitment of N-CoR and SMRT complexes Proc Natl Acad Sci U S A 95, 2920-2925 Lavoie, P L., Budry, L., Balsalobre, A., and Drouin, J (2008) Developmental Dependence on NurRE and EboxNeuro for Expression of Pituitary POMC Mol Endocrinol Le, W D., Xu, P., Jankovic, J., Jiang, H., Appel, S H., Smith, R G., and Vassilatis, D K (2003) Mutations in NR4A2 associated with familial Parkinson disease Nat Genet 33, 85-89 Lee, H K., Hsu, A K., Sajdak, J., Qin, J., and Pavlidis, P (2004) Coexpression analysis of human genes across many microarray data sets Genome Res 14, 1085-1094 Letai, A., Bassik, M C., Walensky, L D., Sorcinelli, M D., Weiler, S., and Korsmeyer, S J (2002) Distinct BH3 domains either sensitize or activate mitochondrial apoptosis, serving as prototype cancer therapeutics Cancer Cell 2, 183-192 Levine, M., and Tjian, R (2003) Transcription regulation and animal diversity Nature 424, 147-151 Li, H., Kolluri, S K., Gu, J., Dawson, M I., Cao, X., Hobbs, P D., Lin, B., Chen, G., Lu, J., Lin, F., et al (2000) Cytochrome c release and apoptosis induced by mitochondrial targeting of nuclear orphan receptor TR3 Science 289, 1159-1164 Li, L Y., Luo, X., and Wang, X (2001) Endonuclease G is an apoptotic DNase when released from mitochondria Nature 412, 95-99 Lightman, S L (2008) The neuroendocrinology of stress: a never ending story J Neuroendocrinol 20, 880-884 Lin, B., Kolluri, S K., Lin, F., Liu, W., Han, Y H., Cao, X., Dawson, M I., Reed, J C., and Zhang, X K (2004) Conversion of Bcl-2 from protector to killer by interaction with nuclear orphan receptor Nur77/TR3 Cell 116, 527-540 Liu, Z G., Smith, S W., McLaughlin, K A., Schwartz, L M., and Osborne, B A (1994) Apoptotic signals delivered through the T-cell receptor of a T-cell hybrid require the immediate-early gene nur77 Nature 367, 281-284 Luo, J., and Elledge, S J (2008) Cancer: Deconstructing oncogenesis Nature 453, 995996 Luthi, A U., and Martin, S J (2007) The CASBAH: a searchable database of caspase substrates Cell Death Differ 14, 641-650 143 Maiuri, M C., Zalckvar, E., Kimchi, A., and Kroemer, G (2007) Self-eating and selfkilling: crosstalk between autophagy and apoptosis Nat Rev Mol Cell Biol 8, 741-752 Mangelsdorf, D J., Thummel, C., Beato, M., Herrlich, P., Schutz, G., Umesono, K., Blumberg, B., Kastner, P., Mark, M., Chambon, P., and Evans, R M (1995) The nuclear receptor superfamily: the second decade Cell 83, 835-839 Margolis, R N., Evans, R M., and O'Malley, B W (2005) The Nuclear Receptor Signaling Atlas: development of a functional atlas of nuclear receptors Mol Endocrinol 19, 2433-2436 Martinez-Gonzalez, J., Rius, J., Castello, A., Cases-Langhoff, C., and Badimon, L (2003) Neuron-derived orphan receptor-1 (NOR-1) modulates vascular smooth muscle cell proliferation Circ Res 92, 96-103 Maruyama, K., Tsukada, T., Bandoh, S., Sasaki, K., Ohkura, N., and Yamaguchi, K (1995) Expression of NOR-1 and its closely related members of the steroid/thyroid hormone receptor superfamily in human neuroblastoma cell lines Cancer Lett 96, 117122 Maruyama, K., Tsukada, T., Bandoh, S., Sasaki, K., Ohkura, N., and Yamaguchi, K (1997) Retinoic acids differentially regulate NOR-1 and its closely related orphan nuclear receptor genes in breast cancer cell line MCF-7 Biochem Biophys Res Commun 231, 417-420 Matsumura, H., Shimizu, Y., Ohsawa, Y., Kawahara, A., Uchiyama, Y., and Nagata, S (2000) Necrotic death pathway in Fas receptor signaling J Cell Biol 151, 1247-1256 McCarthy, N J., Whyte, M K., Gilbert, C S., and Evan, G I (1997) Inhibition of Ced3/ICE-related proteases does not prevent cell death induced by oncogenes, DNA damage, or the Bcl-2 homologue Bak J Cell Biol 136, 215-227 McEvoy, A N., Murphy, E A., Ponnio, T., Conneely, O M., Bresnihan, B., FitzGerald, O., and Murphy, E P (2002) Activation of nuclear orphan receptor NURR1 transcription by NF-kappa B and cyclic adenosine 5'-monophosphate response elementbinding protein in rheumatoid arthritis synovial tissue J Immunol 168, 2979-2987 McMurray, H R., Sampson, E R., Compitello, G., Kinsey, C., Newman, L., Smith, B., Chen, S R., Klebanov, L., Salzman, P., Yakovlev, A., and Land, H (2008) Synergistic response to oncogenic mutations defines gene class critical to cancer phenotype Nature 453, 1112-1116 Metivier, R., Reid, G., and Gannon, F (2006) Transcription in four dimensions: nuclear receptor-directed initiation of gene expression EMBO Rep 7, 161-167 144 Micheau, O., and Tschopp, J (2003) Induction of TNF receptor I-mediated apoptosis via two sequential signaling complexes Cell 114, 181-190 Milbrandt, J (1988) Nerve growth factor induces a gene homologous to the glucocorticoid receptor gene Neuron 1, 183-188 Mills, P B., Struys, E., Jakobs, C., Plecko, B., Baxter, P., Baumgartner, M., Willemsen, M A., Omran, H., Tacke, U., Uhlenberg, B., et al (2006) Mutations in antiquitin in individuals with pyridoxine-dependent seizures Nat Med 12, 307-309 Miura, M., Zhu, H., Rotello, R., Hartwieg, E A., and Yuan, J (1993) Induction of apoptosis in fibroblasts by IL-1 beta-converting enzyme, a mammalian homolog of the C elegans cell death gene ced-3 Cell 75, 653-660 Mix, K S., Attur, M G., Al-Mussawir, H., Abramson, S B., Brinckerhoff, C E., and Murphy, E P (2007) Transcriptional repression of matrix metalloproteinase gene expression by the orphan nuclear receptor NURR1 in cartilage J Biol Chem 282, 94929504 Monden, T., Kishi, M., Hosoya, T., Satoh, T., Wondisford, F E., Hollenberg, A N., Yamada, M., and Mori, M (1999) p120 acts as a specific coactivator for 9-cis-retinoic acid receptor (RXR) on peroxisome proliferator-activated receptor-gamma/RXR heterodimers Mol Endocrinol 13, 1695-1703 Morita, T., Mayanagi, T., and Sobue, K (2007) Dual roles of myocardin-related transcription factors in epithelial mesenchymal transition via slug induction and actin remodeling J Cell Biol 179, 1027-1042 Muller, M., Strand, S., Hug, H., Heinemann, E M., Walczak, H., Hofmann, W J., Stremmel, W., Krammer, P H., and Galle, P R (1997) Drug-induced apoptosis in hepatoma cells is mediated by the CD95 (APO-1/Fas) receptor/ligand system and involves activation of wild-type p53 J Clin Invest 99, 403-413 Mullican, S E., Zhang, S., Konopleva, M., Ruvolo, V., Andreeff, M., Milbrandt, J., and Conneely, O M (2007) Abrogation of nuclear receptors Nr4a3 and Nr4a1 leads to development of acute myeloid leukemia Nat Med 13, 730-735 Nagel, S., Meyer, C., Quentmeier, H., Kaufmann, M., Drexler, H G., and MacLeod, R A (2008) MEF2C is activated by multiple mechanisms in a subset of T-acute lymphoblastic leukemia cell lines Leukemia 22, 600-607 Nagy, L., Kao, H Y., Chakravarti, D., Lin, R J., Hassig, C A., Ayer, D E., Schreiber, S L., and Evans, R M (1997) Nuclear receptor repression mediated by a complex containing SMRT, mSin3A, and histone deacetylase Cell 89, 373-380 145 Nilsson, J., Lund, E., and Lund, A (2001) The effects of UV-irradiation on the ESRdosimetry of tooth enamel Appl Radiat Isot 54, 131-139 Ofir, R., Seidman, R., Rabinski, T., Krup, M., Yavelsky, V., Weinstein, Y., and Wolfson, M (2002) Taxol-induced apoptosis in human SKOV3 ovarian and MCF7 breast carcinoma cells is caspase-3 and caspase-9 independent Cell Death Differ 9, 636-642 Ohkubo, T., Ohkura, N., Maruyama, K., Sasaki, K., Nagasaki, K., Hanzawa, H., Tsukada, T., and Yamaguchi, K (2000) Early induction of the orphan nuclear receptor NOR-1 during cell death of the human breast cancer cell line MCF-7 Mol Cell Endocrinol 162, 151-156 Ohkura, N., Hijikuro, M., Yamamoto, A., and Miki, K (1994) Molecular cloning of a novel thyroid/steroid receptor superfamily gene from cultured rat neuronal cells Biochem Biophys Res Commun 205, 1959-1965 Oppenheim, J J., and Yang, D (2005) Alarmins: chemotactic activators of immune responses Curr Opin Immunol 17, 359-365 Pearen, M A., Ryall, J G., Maxwell, M A., Ohkura, N., Lynch, G S., and Muscat, G E (2006) The orphan nuclear receptor, NOR-1, is a target of beta-adrenergic signaling in skeletal muscle Endocrinology 147, 5217-5227 Pena, A N., and Pereira-Smith, O M (2007) The role of the MORF/MRG family of genes in cell growth, differentiation, DNA repair, and thereby aging Ann N Y Acad Sci 1100, 299-305 Peng, H., Tan, L., Osaki, M., Zhan, Y., Ijiri, K., Tsuchimochi, K., Otero, M., Wang, H., Choy, B K., Grall, F T., et al (2008) ESE-1 is a potent repressor of type II collagen gene (COL2A1) transcription in human chondrocytes J Cell Physiol 215, 562-573 Perlmann, T., and Jansson, L (1995) A novel pathway for vitamin A signaling mediated by RXR heterodimerization with NGFI-B and NURR1 Genes Dev 9, 769-782 Pervaiz, S (2001) Reactive oxygen-dependent photochemotherapeutic agents Faseb J 15, 612-617 production of novel Pervaiz, S., Harriman, A., and Gulliya, K S (1992) Protein damage by photoproducts of merocyanine 540 Free Radic Biol Med 12, 389-396 Pervaiz, S., Hirpara, J L., and Clement, M V (1998) Caspase proteases mediate apoptosis induced by anticancer agent preactivated MC540 in human tumor cell lines Cancer Lett 128, 11-22 Pervaiz, S., and Olivo, M (2006) Art and science of photodynamic therapy Clin Exp Pharmacol Physiol 33, 551-556 146 Pervaiz, S., Seyed, M A., Hirpara, J L., Clement, M V., and Loh, K W (1999) Purified photoproducts of merocyanine 540 trigger cytochrome C release and caspase 8dependent apoptosis in human leukemia and melanoma cells Blood 93, 4096-4108 Peter, M E., and Krammer, P H (1998) Mechanisms of CD95 (APO-1/Fas)-mediated apoptosis Curr Opin Immunol 10, 545-551 Philips, A., Lesage, S., Gingras, R., Maira, M H., Gauthier, Y., Hugo, P., and Drouin, J (1997) Novel dimeric Nur77 signaling mechanism in endocrine and lymphoid cells Mol Cell Biol 17, 5946-5951 Pirih, F Q., Nervina, J M., Pham, L., Aghaloo, T., and Tetradis, S (2003) Parathyroid hormone induces the nuclear orphan receptor NOR-1 in osteoblasts Biochem Biophys Res Commun 306, 144-150 Podar, K., Tonon, G., Sattler, M., Tai, Y T., Legouill, S., Yasui, H., Ishitsuka, K., Kumar, S., Kumar, R., Pandite, L N., et al (2006) The small-molecule VEGF receptor inhibitor pazopanib (GW786034B) targets both tumor and endothelial cells in multiple myeloma Proc Natl Acad Sci U S A 103, 19478-19483 Pols, T W., Ottenhoff, R., Vos, M., Levels, J H., Quax, P H., Meijers, J C., Pannekoek, H., Groen, A K., and de Vries, C J (2008) Nur77 modulates hepatic lipid metabolism through suppression of SREBP1c activity Biochem Biophys Res Commun 366, 910-916 Rajpal, A., Cho, Y A., Yelent, B., Koza-Taylor, P H., Li, D., Chen, E., Whang, M., Kang, C., Turi, T G., and Winoto, A (2003) Transcriptional activation of known and novel apoptotic pathways by Nur77 orphan steroid receptor Embo J 22, 6526-6536 Rao, R V., Hermel, E., Castro-Obregon, S., del Rio, G., Ellerby, L M., Ellerby, H M., and Bredesen, D E (2001) Coupling endoplasmic reticulum stress to the cell death program Mechanism of caspase activation J Biol Chem 276, 33869-33874 Rashmi, R., Pillai, S G., Vijayalingam, S., Ryerse, J., and Chinnadurai, G (2008) BH3only protein BIK induces caspase-independent cell death with autophagic features in Bcl2 null cells Oncogene 27, 1366-1375 Ravagnan, L., Gurbuxani, S., Susin, S A., Maisse, C., Daugas, E., Zamzami, N., Mak, T., Jaattela, M., Penninger, J M., Garrido, C., and Kroemer, G (2001) Heat-shock protein 70 antagonizes apoptosis-inducing factor Nat Cell Biol 3, 839-843 Repa, J J., Turley, S D., Lobaccaro, J A., Medina, J., Li, L., Lustig, K., Shan, B., Heyman, R A., Dietschy, J M., and Mangelsdorf, D J (2000) Regulation of absorption and ABC1-mediated efflux of cholesterol by RXR heterodimers Science 289, 15241529 147 Rius, J., Martinez-Gonzalez, J., Crespo, J., and Badimon, L (2004) Involvement of neuron-derived orphan receptor-1 (NOR-1) in LDL-induced mitogenic stimulus in vascular smooth muscle cells: role of CREB Arterioscler Thromb Vasc Biol 24, 697702 Rius, J., Martinez-Gonzalez, J., Crespo, J., and Badimon, L (2006) NOR-1 is involved in VEGF-induced endothelial cell growth Atherosclerosis 184, 276-282 Rostovtseva, T K., Antonsson, B., Suzuki, M., Youle, R J., Colombini, M., and Bezrukov, S M (2004) Bid, but not Bax, regulates VDAC channels J Biol Chem 279, 13575-13583 Rovere-Querini, P., Capobianco, A., Scaffidi, P., Valentinis, B., Catalanotti, F., Giazzon, M., Dumitriu, I E., Muller, S., Iannacone, M., Traversari, C., et al (2004) HMGB1 is an endogenous immune adjuvant released by necrotic cells EMBO Rep 5, 825-830 Roy, S., Shor, A C., Bagui, T K., Seto, E., and Pledger, W J (2008) Histone deacetylase represses the transcription of cyclin D3 J Cell Biochem Saparbaev, M., Prakash, L., and Prakash, S (1996) Requirement of mismatch repair genes MSH2 and MSH3 in the RAD1-RAD10 pathway of mitotic recombination in Saccharomyces cerevisiae Genetics 142, 727-736 Sebbagh, M., Renvoize, C., Hamelin, J., Riche, N., Bertoglio, J., and Breard, J (2001) Caspase-3-mediated cleavage of ROCK I induces MLC phosphorylation and apoptotic membrane blebbing Nat Cell Biol 3, 346-352 Shi, Y., Evans, J E., and Rock, K L (2003) Molecular identification of a danger signal that alerts the immune system to dying cells Nature 425, 516-521 Shimizu, S., Kanaseki, T., Mizushima, N., Mizuta, T., Arakawa-Kobayashi, S., Thompson, C B., and Tsujimoto, Y (2004) Role of Bcl-2 family proteins in a nonapoptotic programmed cell death dependent on autophagy genes Nat Cell Biol 6, 12211228 Shimizu, S., Konishi, A., Kodama, T., and Tsujimoto, Y (2000) BH4 domain of antiapoptotic Bcl-2 family members closes voltage-dependent anion channel and inhibits apoptotic mitochondrial changes and cell death Proc Natl Acad Sci U S A 97, 31003105 Shimizu, S., Narita, M., and Tsujimoto, Y (1999) Bcl-2 family proteins regulate the release of apoptogenic cytochrome c by the mitochondrial channel VDAC Nature 399, 483-487 148 Shin, H J., Park, K K., Lee, B H., Moon, C K., and Lee, M O (2003) Identification of genes that are induced after cadmium exposure by suppression subtractive hybridization Toxicology 191, 121-131 Shintani, T., and Klionsky, D J (2004) Autophagy in health and disease: a double-edged sword Science 306, 990-995 Shipp, M A., Ross, K N., Tamayo, P., Weng, A P., Kutok, J L., Aguiar, R C., Gaasenbeek, M., Angelo, M., Reich, M., Pinkus, G S., et al (2002) Diffuse large B-cell lymphoma outcome prediction by gene-expression profiling and supervised machine learning Nat Med 8, 68-74 Sibayama-Imazu, T., Fujisawa, Y., Masuda, Y., Aiuchi, T., Nakajo, S., Itabe, H., and Nakaya, K (2008) Induction of apoptosis in PA-1 ovarian cancer cells by vitamin K(2) is associated with an increase in the level of TR3/Nur77 and its accumulation in mitochondria and nuclei J Cancer Res Clin Oncol 134, 803-812 Slee, E A., Adrain, C., and Martin, S J (2001) Executioner caspase-3, -6, and -7 perform distinct, non-redundant roles during the demolition phase of apoptosis J Biol Chem 276, 7320-7326 Smith, A G., Luk, N., Newton, R A., Roberts, D W., Sturm, R A., and Muscat, G E (2008) Melanocortin-1 receptor signalling markedly induces the expression of the NR4A nuclear receptor subgroup in melanocytic cells J Biol Chem Song, K H., Park, J I., Lee, M O., Soh, J., Lee, K., and Choi, H S (2001) LH induces orphan nuclear receptor Nur77 gene expression in testicular Leydig cells Endocrinology 142, 5116-5123 Srivastava, R K., Sasaki, C Y., Hardwick, J M., and Longo, D L (1999) Bcl-2mediated drug resistance: inhibition of apoptosis by blocking nuclear factor of activated T lymphocytes (NFAT)-induced Fas ligand transcription J Exp Med 190, 253-265 Stasik, I., Rapak, A., Kalas, W., Ziolo, E., and Strzadala, L (2007) Ionomycin-induced apoptosis of thymocytes is independent of Nur77 NBRE or NurRE binding, but is accompanied by Nur77 mitochondrial targeting Biochim Biophys Acta 1773, 1483-1490 Stocco, C O., Zhong, L., Sugimoto, Y., Ichikawa, A., Lau, L F., and Gibori, G (2000) Prostaglandin F2alpha-induced expression of 20alpha-hydroxysteroid dehydrogenase involves the transcription factor NUR77 J Biol Chem 275, 37202-37211 Suzuki, T., Nishiyama, K., Yamamoto, A., Inazawa, J., Iwaki, T., Yamada, T., Kanazawa, I., and Sakaki, Y (2000) Molecular cloning of a novel apoptosis-related gene, human Nap1 (NCKAP1), and its possible relation to Alzheimer disease Genomics 63, 246-254 149 Suzuki, Y., Imai, Y., Nakayama, H., Takahashi, K., Takio, K., and Takahashi, R (2001) A serine protease, HtrA2, is released from the mitochondria and interacts with XIAP, inducing cell death Mol Cell 8, 613-621 Swetloff, A., and Ferretti, P (2005) Changes in E2F5 intracellular localization in mouse and human choroid plexus epithelium with development Int J Dev Biol 49, 859-865 Takeda, I., Maruya, S., Shirasaki, T., Mizukami, H., Takahata, T., Myers, J N., Kakehata, S., Yagihashi, S., and Shinkawa, H (2007) Simvastatin inactivates beta1integrin and extracellular signal-related kinase signaling and inhibits cell proliferation in head and neck squamous cell carcinoma cells Cancer Sci 98, 890-899 Tata, J R (2002) Signalling through nuclear receptors Nat Rev Mol Cell Biol 3, 702710 Tay, S T., Leong, S H., Yu, K., Aggarwal, A., Tan, S Y., Lee, C H., Wong, K., Visvanathan, J., Lim, D., Wong, W K., et al (2003) A combined comparative genomic hybridization and expression microarray analysis of gastric cancer reveals novel molecular subtypes Cancer Res 63, 3309-3316 Taylor, R C., Cullen, S P., and Martin, S J (2008) Apoptosis: controlled demolition at the cellular level Nat Rev Mol Cell Biol 9, 231-241 Temkin, V., Huang, Q., Liu, H., Osada, H., and Pope, R M (2006) Inhibition of ADP/ATP exchange in receptor-interacting protein-mediated necrosis Mol Cell Biol 26, 2215-2225 Terstappen, G C., Schlupen, C., Raggiaschi, R., and Gaviraghi, G (2007) Target deconvolution strategies in drug discovery Nat Rev Drug Discov 6, 891-903 Thompson, J., and Winoto, A (2008) During negative selection, Nur77 family proteins translocate to mitochondria where they associate with Bcl-2 and expose its proapoptotic BH3 domain J Exp Med 205, 1029-1036 Tracy, K., Dibling, B C., Spike, B T., Knabb, J R., Schumacker, P., and Macleod, K F (2007) BNIP3 is an RB/E2F target gene required for hypoxia-induced autophagy Mol Cell Biol 27, 6229-6242 Upton, Z., Cuttle, L., Noble, A., Kempf, M., Topping, G., Malda, J., Xie, Y., Mill, J., Harkin, D G., Kravchuk, O., et al (2008) Vitronectin: growth factor complexes hold potential as a wound therapy approach J Invest Dermatol 128, 1535-1544 Vaux, D L., and Silke, J (2005) IAPs, RINGs and ubiquitylation Nat Rev Mol Cell Biol 6, 287-297 150 Virag, L., Salzman, A L., and Szabo, C (1998) Poly(ADP-ribose) synthetase activation mediates mitochondrial injury during oxidant-induced cell death J Immunol 161, 37533759 Wang, X., Yang, C., Chai, J., Shi, Y., and Xue, D (2002) Mechanisms of AIF-mediated apoptotic DNA degradation in Caenorhabditis elegans Science 298, 1587-1592 Wang, Y C., Kulp, S K., Wang, D., Yang, C C., Sargeant, A M., Hung, J H., Kashida, Y., Yamaguchi, M., Chang, G D., and Chen, C S (2008) Targeting endoplasmic reticulum stress and Akt with OSU-03012 and gefitinib or erlotinib to overcome resistance to epidermal growth factor receptor inhibitors Cancer Res 68, 2820-2830 Wansa, K D., Harris, J M., Yan, G., Ordentlich, P., and Muscat, G E (2003) The AF-1 domain of the orphan nuclear receptor NOR-1 mediates trans-activation, coactivator recruitment, and activation by the purine anti-metabolite 6-mercaptopurine J Biol Chem 278, 24776-24790 Wanschers, B F., van de Vorstenbosch, R., Schlager, M A., Splinter, D., Akhmanova, A., Hoogenraad, C C., Wieringa, B., and Fransen, J A (2007) A role for the Rab6B Bicaudal-D1 interaction in retrograde transport in neuronal cells Exp Cell Res 313, 3408-3420 Waterhouse, N J., Goldstein, J C., von Ahsen, O., Schuler, M., Newmeyer, D D., and Green, D R (2001) Cytochrome c maintains mitochondrial transmembrane potential and ATP generation after outer mitochondrial membrane permeabilization during the apoptotic process J Cell Biol 153, 319-328 Wei, M C., Lindsten, T., Mootha, V K., Weiler, S., Gross, A., Ashiya, M., Thompson, C B., and Korsmeyer, S J (2000) tBID, a membrane-targeted death ligand, oligomerizes BAK to release cytochrome c Genes Dev 14, 2060-2071 Wei, M C., Zong, W X., Cheng, E H., Lindsten, T., Panoutsakopoulou, V., Ross, A J., Roth, K A., MacGregor, G R., Thompson, C B., and Korsmeyer, S J (2001) Proapoptotic BAX and BAK: a requisite gateway to mitochondrial dysfunction and death Science 292, 727-730 Weih, F., Ryseck, R P., Chen, L., and Bravo, R (1996) Apoptosis of nur77/N10transgenic thymocytes involves the Fas/Fas ligand pathway Proc Natl Acad Sci U S A 93, 5533-5538 Williams, G T., and Lau, L F (1993) Activation of the inducible orphan receptor gene nur77 by serum growth factors: dissociation of immediate-early and delayed-early responses Mol Cell Biol 13, 6124-6136 Willson, T M., Brown, P J., Sternbach, D D., and Henke, B R (2000) The PPARs: from orphan receptors to drug discovery J Med Chem 43, 527-550 151 Wilson, T E., Fahrner, T J., Johnston, M., and Milbrandt, J (1991) Identification of the DNA binding site for NGFI-B by genetic selection in yeast Science 252, 1296-1300 Wolter, K G., Hsu, Y T., Smith, C L., Nechushtan, A., Xi, X G., and Youle, R J (1997) Movement of Bax from the cytosol to mitochondria during apoptosis J Cell Biol 139, 1281-1292 Woo, M., Hakem, R., Soengas, M S., Duncan, G S., Shahinian, A., Kagi, D., Hakem, A., McCurrach, M., Khoo, W., Kaufman, S A., et al (1998) Essential contribution of caspase 3/CPP32 to apoptosis and its associated nuclear changes Genes Dev 12, 806819 Wood, L D., Parsons, D W., Jones, S., Lin, J., Sjoblom, T., Leary, R J., Shen, D., Boca, S M., Barber, T., Ptak, J., et al (2007) The genomic landscapes of human breast and colorectal cancers Science 318, 1108-1113 Woronicz, J D., Calnan, B., Ngo, V., and Winoto, A (1994) Requirement for the orphan steroid receptor Nur77 in apoptosis of T-cell hybridomas Nature 367, 277-281 Wrange, O., and Gustafsson, J A (1978) Separation of the hormone- and DNA-binding sites of the hepatic glucocorticoid receptor by means of proteolysis J Biol Chem 253, 856-865 Wu, H Y., Lin, C Y., Lin, T Y., Chen, T C., and Yuan, C J (2008) Mammalian Ste20-like protein kinase mediates trophoblast apoptosis in spontaneous delivery Apoptosis 13, 283-294 Wulfkuhle, J., Espina, V., Liotta, L., and Petricoin, E (2004) Genomic and proteomic technologies for individualisation and improvement of cancer treatment Eur J Cancer 40, 2623-2632 Xiang, J., Chao, D T., and Korsmeyer, S J (1996) BAX-induced cell death may not require interleukin beta-converting enzyme-like proteases Proc Natl Acad Sci U S A 93, 14559-14563 Yamamoto, A., Suzuki, T., and Sakaki, Y (2001) Isolation of hNap1BP which interacts with human Nap1 (NCKAP1) whose expression is down-regulated in Alzheimer's disease Gene 271, 159-169 Yang, X., Downes, M., Yu, R T., Bookout, A L., He, W., Straume, M., Mangelsdorf, D J., and Evans, R M (2006) Nuclear receptor expression links the circadian clock to metabolism Cell 126, 801-810 Yin, C., Knudson, C M., Korsmeyer, S J., and Van Dyke, T (1997) Bax suppresses tumorigenesis and stimulates apoptosis in vivo Nature 385, 637-640 152 Yin, M J., Shao, L., Voehringer, D., Smeal, T., and Jallal, B (2003) The serine/threonine kinase Nek6 is required for cell cycle progression through mitosis J Biol Chem 278, 52454-52460 Yin, X M., Oltvai, Z N., and Korsmeyer, S J (1994) BH1 and BH2 domains of Bcl-2 are required for inhibition of apoptosis and heterodimerization with Bax Nature 369, 321-323 Youn, H D., Sun, L., Prywes, R., and Liu, J O (1999) Apoptosis of T cells mediated by Ca2+-induced release of the transcription factor MEF2 Science 286, 790-793 Yu, S W., Wang, H., Poitras, M F., Coombs, C., Bowers, W J., Federoff, H J., Poirier, G G., Dawson, T M., and Dawson, V L (2002) Mediation of poly(ADP-ribose) polymerase-1-dependent cell death by apoptosis-inducing factor Science 297, 259-263 Yuan, J., and Horvitz, H R (2004) A first insight into the molecular mechanisms of apoptosis Cell 116, S53-56, 51 p following S59 Yuan, J., Shaham, S., Ledoux, S., Ellis, H M., and Horvitz, H R (1993) The C elegans cell death gene ced-3 encodes a protein similar to mammalian interleukin-1 betaconverting enzyme Cell 75, 641-652 Zamzami, N., and Kroemer, G (2001) The mitochondrion in apoptosis: how Pandora's box opens Nat Rev Mol Cell Biol 2, 67-71 Zetterstrom, R H., Solomin, L., Mitsiadis, T., Olson, L., and Perlmann, T (1996) Retinoid X receptor heterodimerization and developmental expression distinguish the orphan nuclear receptors NGFI-B, Nurr1, and Nor1 Mol Endocrinol 10, 1656-1666 Zhang, T., Jia, N., Fei, E., Wang, P., Liao, Z., Ding, L., Yan, M., Nukina, N., Zhou, J., and Wang, G (2007) Nurr1 is phosphorylated by ERK2 in vitro and its phosphorylation upregulates tyrosine hydroxylase expression in SH-SY5Y cells Neurosci Lett 423, 118122 Zheng, T S., Hunot, S., Kuida, K., Momoi, T., Srinivasan, A., Nicholson, D W., Lazebnik, Y., and Flavell, R A (2000) Deficiency in caspase-9 or caspase-3 induces compensatory caspase activation Nat Med 6, 1241-1247 Zheng, Y., Shi, Y., Tian, C., Jiang, C., Jin, H., Chen, J., Almasan, A., Tang, H., and Chen, Q (2004) Essential role of the voltage-dependent anion channel (VDAC) in mitochondrial permeability transition pore opening and cytochrome c release induced by arsenic trioxide Oncogene 23, 1239-1247 153 ... Accordingly, the great and urgent need presently is to investigate the role of each of the plenitude of genes and proteins important in the progression of cancer NUCLEAR RECEPTORS The nuclear receptors are. .. Tan, Shazib Pervaiz, ? ?The NR4A orphan nuclear receptors are target genes of the novel drug C1 in cancer cells and potential mediators of drug induced apoptosis? ?? (manuscript under preparation) CONFERENCE... family of receptors This study characterizes the role of nuclear receptors NR4A1 and NR4A3 in drug C1 mediated apoptosis and has identified the functional relevance of the increase in the transcript