Mechanisms and biological consequences of the nuclear receptor CAR activation

National Institute Of Environmental Health Sciences

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Abstract

CAR activation mechanism: What is unique about this system is the fact that xenobiotics do not directly bind to CAR to activate it. We previously determined that threonine 48 of endogenous CAR is phosphorylated in mouse primary hepatocytes and that phenobarbital treatment de-phosphorylates this threonine, activating CAR and translocating it into the nucleus. We have now identified protein phosphatase 2A as the enzyme that de-phosphorylates threonine 48 of CAR. Moreover, receptor for activated C-kinase1 (RACK1) was characterized as the essential regulatory subunit that activates the PP2A core enzyme to de-phosphorylate threonine 48 of CAR. We have also defined the growth factor-MEK1/2-ERK1/2 pathway as a cell signaling mechanism that represses de-phosphorylation of threonine 48, thus repressing CAR activation. Activated P-ERK1/2 directly interacts with phosphorylated CAR and prevents PP2A from interacting with CAR and de-phosphorylating threonine 48. Phenobarbital treatment results in inactivation of P-ERK1/2 and enables PP2A to de-phosphorylate threonine 48 in mouse primary hepatocytes. Our investigations have defined this de-phosphorylation as the principle regulator of CAR activation and functions. Xenobiotic-signal crosstalk mechanism: Upon activation by xenobiotics, CAR regulates genes differently from one another, conferring specificity to CAR-regulated gene expression. CAR acquires this specificity via crosstalk with cell signaling. We have identified various endogenous cell signals as the essential regulator of CAR activation and function: pituitary factor, p38 signaling, SGK2 signaling and the early response factor GADD45 (growth arrest and DNA-damage inducible 45. Given these findings, we are investigating the molecular mechanisms by which these signaling regulate CAR activation and functions. CAR-mediated diseases: Chronic treatment with drugs, such as phenobarbital, is known to activate CAR and cause hepatocellular carcinoma (HCC) in rodents. We have now characterized GADD45 as a CAR target for phenobarbital promotion of HCC development: CAR interacts with GADD45 protein and this interaction inhibits phosphorylation of JNK1, thus repressing apoptosis and possibly promoting tumor genesis. 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) treatment is known to cause severe liver injury and proliferate postnatal hepatic progenitor oval cells. Utilizing CAR KO mice, we have determined that DDC activates CAR and this activation is essential for the developments of liver injury and oval cell proliferation.