The antiapoptotic factor GADD45B has now been shown to regulate CAR-mediated transcription of the CYP2B6 gene in HepG2 cells and the Cyp2b10 in mouse primary hepatocytes. Together with our previous finding of direct CAR interaction with GADD45B (manuscript submitted for publication), present cell-based transfection assays have demonstrated that GADD45B is a new transcriptional coactivator of CAR. Because GADD45B is a factor that may transfer signals into various cellular pathways, an indirect effect of GADD45B to coactivate CAR cannot totally be eliminated as the possible regulatory mechanism at this present time. Nevertheless, the fact that expression levels of CYP2B10 mRNA by TCPOBOP in GADD45B-KO primary hepatocytes was reduced to 20% to the corresponding levels observed in wild-type primary hepatocytes indicates that coactivation by GADD45B appears to be critical for maximal CAR activation of transcription of the Cyp2b10 gene.
Responding to various endogenous stimuli, the Gadd45b
gene is activated, and GADD45B protein is involved in transducing various cell signals (Abdollahi et al., 1991
). It is now found that CAR enables the Gadd45b
gene to respond to xenobiotics. The biological consequences of xenobiotic activation of the Gadd45b
gene remain an attractive subject for future research. The CAR activators PB and TCPOBOP are nongenotoxic carcinogens, and chronic CAR activation by these activators promotes development of hepatocellular carcinoma (HCC) in rodents (Yamamoto et al., 2004
; Huang et al., 2005
). We have demonstrated that treatment with TCPOBOP attenuates tumor necrosis factor α
-induced cell death of mouse primary hepatocytes only in the presence of both CAR and GADD45B, suggesting that CAR may promote HCC by down-regulating the apoptotic JNK pathway (manuscript submitted for publication). PB induction of xenobiotic metabolizing enzymes such as CYP2B has also been suggested to be a factor responsible for PB promotion of HCC (Diwan et al., 2001
), although this suggestion has not yet been experimentally demonstrated. Our present finding that GADD45B is a CAR coactivator raises an intriguing question as to whether the role of CYP enzymes in HCC development should be revisited. When rodents are treated with CAR activators, liver GADD45B increases and perhaps puts the animals into a vicious cycle of continuous induction of the CYP2B enzyme (). Thus, in response to CAR activation, GADD45B coordinates attenuation of apoptosis and induction of the CYP2B enzyme and/or other CYP enzyme that may synthesize chemicals assisting HCC development.
Cross talk regulation of GADD45B to coactivate CAR. Whether CAR directly activates the Gadd45b gene remains a subject for future investigations.
The liver is endowed with the metabolic capability to detoxify xenobiotics, including therapeutic drugs to counter toxicity and carcinogenicity caused by xenobiotics. Microsomal CYP enzymes provide this metabolic capability with the flexibility and adaptability for the liver to deal with an unlimited number of xenobiotics and their structural diversity. This flexibility and adaptability comes from various unique characteristics of the CYP enzymes and CYP
genes, one of which is the capability of CYP
genes to exhibit altered transcriptional activity in response to xenobiotic exposure, thereby regulating the levels of CYP enzymes. Human CYP2B6 is one such CYP enzyme. CYP2B6 exhibits extreme individual variations in its basal and induced levels in human livers, which may affect how patients respond to drug therapy and human susceptibility to xenobiotic exposure (Zanger et al., 2007
). We previously found that insulin represses the mouse Cyp2b
gene by down-regulating CAR-mediated transcription indirectly through the forkhead transcription factor, FoxO1, but not by directly regulating CAR expression (Kodama et al., 2004
). Here, we have now identified GADD45B as being an example of indirect up-regulation of CAR-mediated transcription of the CYP2B
genes. Thus, factors such as GADD45B and FoxO1 provide CAR with a molecular mechanism by which endogenous hormones and stimuli become the critical determinants of hepatic metabolic capability of xenobiotics.
Inflammatory stimulation by lipopolysaccharide (LPS) treatment increases GADD45B through a nuclear factor-κ
B pathway in mouse liver (Zhang et al., 2005
). LPS treatment is known to strongly repress expression of the Cyp2b
gene and its induction by PB in mouse liver and primary hepatocytes (Morgan, 1997
; Li-Masters and Morgan, 2001
). These responses to LPS treatment appear to contradict our conclusion that GADD45B coactivates CAR-mediated transcription. However, a more recent report demonstrated that LPS treatment greatly reduces the levels of CAR protein in mouse liver, resulting in the liver's inability to activate the Cyp2b10
gene (Beigneux et al., 2002
). Although its molecular mechanism is not known, this LPS-dependent reduction seems to be a nonspecific event because LPS reduces not only the levels of CAR but also those of PXR and RXRs as well. Once the nonspecific reduction of CAR is eliminated, increases of GADD45B can be expected to be involved in coregulation of CAR-mediated induction of the CYP2B enzyme. The extent to which the inflammation signal via GADD45B modulates the hepatic levels of CYP2B
genes and also the other CAR-target genes remains an intriguing question at the present time.