Nuclear receptor CAR represses TNFα-induced cell death by interacting with the anti-apoptotic GADD45B

Laboratory of Reproductive and Developmental Toxicology, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, North Carolina, United States of America.
PLoS ONE (Impact Factor: 3.23). 04/2010; 5(4):e10121. DOI: 10.1371/journal.pone.0010121
Source: PubMed


Phenobarbital (PB) is the most well-known among numerous non-genotoxic carcinogens that cause the development of hepatocellular carcinoma (HCC). PB activates nuclear xenobiotic receptor Constitutive Active/Androstane Receptor (CAR; NR1I3) and this activation is shown to determine PB promotion of HCC in mice. The molecular mechanism of CAR-mediated tumor promotion, however, remains elusive at the present time. Here we have identified Growth Arrest and DNA Damage-inducible 45beta (GADD45B) as a novel CAR target, through which CAR represses cell death.
PB activation of nuclear xenobiotic receptor CAR is found to induce the Gadd45b gene in mouse liver throughout the development of HCC as well as in liver tumors. Given the known function of GADD45B as a factor that represses Mitogen-activated protein Kinase Kinase 7 - c-Jun N-terminal Kinase (MKK7-JNK) pathway-mediated apoptosis, we have now demonstrated that CAR interacts with GADD45B to repress Tumor Necrosis Factor alpha ( TNFalpha)-induced JNK1 phosphorylation as well as cell death. Primary hepatocytes, prepared from Car(+/+), Car(-/-), Gadd45b(+/+) and Gadd45b(-/-) mice, were treated with TNFalpha and Actinomycin D to induce phosphorylation of JNK1 and cell death. Co-treatment with the CAR activating ligand TCPOBOP (1,4 bis[2-(3,5-dichloropyridyloxy)]benzene) has resulted in repression of both phosphorylation and cell death in the primary hepatocytes from Car(+/+) but not Car(-/-) mice. Repression by TCPOBOP was not observed in those prepared from Gadd45b(-/-) mice. In vitro protein-protein interaction and phosphorylation assays have revealed that CAR interacts with MKK7 and represses the MKK7-mediated phosphorylation of JNK1.
CAR can form a protein complex with GADD45B, through which CAR represses MKK7-mediated phosphorylation of JNK1. In addition to activating the Gadd45b gene, CAR may repress death of mouse primary hepatocytes by forming a GADD45B complex and repressing MKK7-mediated phosphorylation of JNK1. The present finding that CAR can repress cell death via its interaction with GADD45B provides an insight for further investigations into the CAR-regulated molecular mechanism by which PB promotes development of HCC.

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    • "To date, many groups have reported possible mechanisms for the CAR-mediated hepatocyte proliferation in mice. For example, CAR induced the transcription of the genes encoding modulators of p53 tumor suppressor protein, such as Gadd45 and Mdm2 [9], [10], [11]. Another report demonstrated that CAR-induced hepatocyte hyperplasia was mediated by the expression of the oncogene c-Myc and its target Foxm1 [12]. "
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    ABSTRACT: Xenobiotic-responsive nuclear receptors pregnane X receptor (PXR), constitutive active/androstane receptor (CAR) and peroxisome proliferator-activated receptor α (PPARα) play pivotal roles in the metabolic functions of the liver such as xenobiotics detoxification and energy metabolism. While CAR or PPARα activation induces hepatocyte proliferation and hepatocarcinogenesis in rodent models, it remains unclear whether PXR activation also shows such effects. In the present study, we have investigated the role of PXR in the xenobiotic-induced hepatocyte proliferation with or without CAR activation by 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP) and phenobarbital, or PPARα activation by Wy-14643 in mice. Treatment with TCPOBOP or phenobarbital increased the percentage of Ki-67-positive nuclei as well as mRNA levels of cell proliferation-related genes in livers as expected. On the other hand, treatment with the PXR activator pregnenolone 16α-carbonitrile (PCN) alone showed no such effects. Surprisingly, PCN co-treatment significantly augmented the hepatocyte proliferation induced by CAR activation with TCPOBOP or phenobarbital in wild-type mice but not in PXR-deficient mice. Intriguingly, PXR activation also augmented the hepatocyte proliferation induced by Wy-14643 treatment. Moreover, PCN treatment increased the RNA content of hepatocytes, suggesting the induction of G0/G1 transition, and reduced mRNA levels of Cdkn1b and Rbl2, encoding suppressors of cell cycle initiation. Our present findings indicate that xenobiotic-induced hepatocyte proliferation mediated by CAR or PPARα is enhanced by PXR co-activation despite that PXR activation alone does not cause the cell proliferation in mouse livers. Thus PXR may play a novel and unique role in the hepatocyte/liver hyperplasia upon exposure to xenobiotics.
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    • "The reasons for these apparent differences are not clear but may lie in the nature of the mouse models used to generate the results. Mechanistically, GADD45B (Yamamoto et al., 2010), an antiapoptotic factor, as well as Mdm2 (Huang et al., 2005), a negative regulator of the p53 tumor suppressor, have been implicated as pathways activated by CAR and contributory to the enhanced tumorigenic response in CAR wild-type animals. "
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