Regulation of Human Cytochrome P4501A2 Gene (CYP1A2) Expression

Injae Chung 대한약학회 1997 大韓藥學會 總會 및 學術大會 Vol.1997 No.1

No Role of Protected Region B of Human Cytochrome P4501A2 Gene (CYP1A2) As an AP-1 Response Element

Chung, Injae,Jung, Kihwa 德成女子大學校 藥學硏究所 2002 藥學論文誌 Vol.13 No.1

Cytochrome P4501A2 (CYP1A2) is a member of the cytochrome P450 family of isozymes involved in the phase I drug metabolism of vertebrates. CYP1A2 is responsible for the activation of a number of aromatic amines to mutagenic and carcinogenic forms. Thus, the level of CYP1A2, which varies among different populations, may determine an individual's susceptibility to these chemicals. We have previously reported on the importance of a cis element named PRB (protected region B) in the regulation of human Cytochrome P4501A2 (CYP1A2) gene, sequence(-2218 to -2187 bp) revealed a putative AP-1 binding site, TGACTAA, at -2212 bp(Chung and Brenick, 1997). To elucidate the role of AP-1 in CYP1A2 regulation, we transiently overexpressed c-Jun and c-Fos transcription factors in human hepatoma HepG2 cells, and examined their influence on the CYP1A2 promoter activity by reporter gene assays. Cotransfection of the c-Jun and the c-Fos expression vectors increased the induced transactivation by five to six fold from the CYP1A2 promoter constructs. However, deletion of the PRB element did not affect the degree of activation by the c-Jun and the c-Fos. Therefore, it is unlikely that the c-Jun and the c-fos activate the CYP1A2 promoter through this AP-1 consensus-like sequence in the PRB region.

Regulation of the Constitutive Expression of the Human CYP1A2 Gene : Cis Elements and Their Interactions with Proteins

CHUNG, INJAE,BRESNICK, EDWARD 德成女子大學校 藥學硏究所 1997 藥學論文誌 Vol.8 No.1

Cytochorome P4501A2(CYP1A2) is a member of the cytochrome P450 family that is involved in phase I drug metabolism in vertebrates. To understand how the constitutive expression of the human CYP1A2 gene is regulated, its 5'flanking region was analyzed. The promother activity of a human CYP1A2 gene sequence[base pairs(bp)-3203 to + 58bp] was measured in transiently transfected HepG2 cells using fusion constructs containing the luciferase reporter gene. Using 5'-end deletion analysis, two functionally important cis elements, i.e., a proximal 42-bp DNA from bp -72 to bp -31 and a distal 259-bp DNA from bp -2352 to bp -2094, were identified. The proximal sequence (bp -72 to -31) contained CCAAT and GC boxes, with which well characterized transcription factors such as nuclear factor-1/CCAAT transcription factor and simian virus 40 promoter factor-1 could interact. With regard to the 259-bp fragment (bp -2352 to bp -2094), gel mobility shift analyses with HepG2 nuclear lysates indicated high affinity, specific interactions of several trans-acting factors. Three protein binding sites within the 259-bp fragment were identified by DNasel footprinting analysis; these sites contained activator protein-1, nuclear factor-E1.7, and one-half hepatic nuclear factor-1(HNF-1)binding consensus sequences. Only the region from bp -2124 to bp -2098, in which the HNF-1 binding site was located, was markedly protected by a HepG2 nuclear extract, compared with a MCF7 human breast cancer nuclear extract. These results suggested that the 259-bp DNA fragment contained positive regulator binding sites and HNF-1 could contribute to the liver-specific expression of human CYP1A2.

Human CYP1A2 Promoter Fused-Luciferase Gene Constructs Hardly Respond to Polycyclic Hydrocarbons in Transient Transfection Study in HepG2 Cells

Chung, Injae 德成女子大學校 藥學硏究所 2000 藥學論文誌 Vol.11 No.1

In previous study. both constitutive expression and 3-methylcholanthrene (3MC)-mediated elevation of CYP1A2 mRNA were demonstrated in human hepatoma HepG2 cells by reverse transcription-polymerase chain reaction (RT-PCR), suggesting that HepG2 cells would be appropriate for the study of human CYP1A2 regulation(Chung and Bresnick. 1994). Further studies were conducted to detemine the basis of this induction phenomenon that is observed in HepG2 cells. Since CYP1A1 gene. another polycyclic hydrocarbon(PH)-inducible gene. is regulated by PHs through their interactions via receptors with cis-elements, the 5-flanking region of human CYP1A2 gene was analyzed to search such responsive elements. The promoter activity of various lengths of CYP1A2 gene sequence (-3203/+58 bp) was measured in transiently-transfected HepG2 cells by fusion constructs containing the CAT,hGH or luciferase genes as a reporter. This region of the CYP1A2 gene,although containing a XRE, was only weakly responsive (less than 2 fold induction) to 10 nM of TCDD or 1 uM 3 MC treatment. This small enhancement of promoter activity is inconsistent with the previous observation, i.e.. 12 to 14 fold-enhanced CYP1A2 mRNA from 1 uM 3 MC treated HepG2 cells, suggesting that additional mechanisms would exist for PH-mediated induction of CYP1A2 in these cells.

Cancer Gene Therapy : Chemosensitization by an Enzyme-Prodrug Activation Strategy

(Injae Chung) 한국응용약물학회 1997 학술대회 Vol.1997 No.2

Prediction of Transonic Buffet Onset for a Supercritical Airfoil with Shock-Boundary Layer Interactions Using Navier-Stokes Solver

Injae Chung 한국항공우주학회 2017 International Journal of Aeronautical and Space Sc Vol.18 No.1

To predict the transonic buffet onset for a supercritical airfoil with shock-boundary layer interactions, a practical steady approach has been proposed. In this study, it is assumed that the airfoil flow is steady even when buffet onset occurs. Steady Navier-Stokes computations are performed on the supercritical airfoil. Using the aerodynamic parameters calculated from Navier-Stokes solver, various steady approaches for predicting buffet onset are discussed. Among the various steady approaches considered in this study, Thomas’ criterion based on Navier-Stokes computation has shown to be the most appropriate indicator of identifying the buffet onset for a supercritical airfoil with shock-boundary layer interactions. Good agreements have been obtained compared with the results of unsteady transonic wind tunnel tests. The present method is shown to be reliable and useful for transonic buffet onset for a supercritical airfoil with shock-boundary layer interactions in terms of practical engineering viewpoint.

Identification of Positive and Negative Regulatory Elements of the Human Cytochrome P4501A2(CYP1A2) Gene

Chung, Injae,Bresnick, Edward 德成女子大學校 藥學硏究所 1998 藥學論文誌 Vol.9 No.1

We previously demonstrated an enhancer-like positive regulatory element within a 259-bp sequence (-2352 to -2094 bp) of the human CYP1A2 gene in HepG2 cells. Three protein binding sites were identified by DNase I footprinting analyses within the 259-bp sequence: protected region A PRA; -2283 to -2243 bp), PRB(-2218 to -2187 bp), and PRC(-2124 to -2098 bp) (I. Chung and E. Bresnick, Mol. Pharmacol. 47, 677-685, 1995). In the present study, the functional significance of those protected regions was examined. Transfection experiments with delection and substitution mutants defined the PRB and PRC as containing positive and negative regulatory elements, respectively. Human breast carcinoma MCF-7 cells were cotransfected with a hepatocyte nuclear factor-1 (HNF-1) expression vector and CYP1A2 promoter- or thymidine kinase promoter-luciferase reporter gene constructs. HNF-1, which contributes to the liver specificity of genes, enhanced reporter gene activity in a PRC sequence-dependent manner. These results suggested that PRC could exist bound to a repressor which was displaceable by other transcription factors such as HNF-1. Results obtained by transfection of HepG2 hepatoma cells with various PRB substitution mutant-luciferase gene fusion constructs indicated that the entire sequence of PRB was necessary for promoter activity. Consequently, the regulation of CYP1A2 expression is very complex, requiring a number of both positive and negative regulatory factors.

Human CYP1A2 Promoter Fused-Luciferase Gene Constructs Hardly Respond to Polycyclic Hydrocarbons in Transient Transfection Study in HepG2 Cells

Chung, Injae The Korean Society of Toxicology Korea Environment 2000 Toxicological Research Vol.16 No.2

In previous study, both constitutive expression and 3-methylcholanthrene (3MC)-mediated elevation of CYP1A2 mRNA were demonstrated in human hepatoma HepG2 cells by reverse transcription-polymerase chain reaction (RT-PCR), suggesting that HepG2 cells would be appropriate for the study of human CYP1A2 regulation(Chung and Bresnick, 1994). Further studies were conducted to determine the basis of this induction phenomenon that is observed in HepG2 cells. Since CYP1A1 gene, another polycyclic hydrocarbon(PH)-inducible gene, is regulated by PHs through their interactions via receptors with cis-elements, the 5'-flanking region of human CYP 1A2 gene was analyzed to search such responsive elements. The promoter activity of various lengths of CYP1A2 gene sequence (-3203/+58bp) was measured in transiently-transfected HepG2 cells by fusion constructs containing the CAT, hGH or luciferase genes as a reporter. This region of the CYP1A2 gene, although containing a XRE, was only weakly responsive (less than 2 fold induction) to 10 nM of TCDD or 1 $\mu$M 3 MC treatment. This small enhancement of promoter activity is inconsistent with the previous observation, i.e., 12 to 14 fold-enhanced CYP1A2 mRNA from 1 $\mu$M 3 MC treated HepG2 cells, suggesting that additional mechanisms would exist for PH-mediated induction of CYP1A2 in these cells.

Molecular Mechanisms of Regulation of Human Cytochrome P4501A2 Gene Expression

Injae Chung 한국생약학회 2004 Natural Product Sciences Vol.10 No.5

Cytochrome P4501A2 (CYP1A2) is responsible for the metabolic activation of a number of aromatic amines and amides to mutagenic and carcinogenic moieties. Considerable variations in the level of CYP1A2 expression in humans have been reported. Thus, the level of human CYP1A2 may determine an individuals susceptibility to these chemicals. Given its importance, the molecular mechanisms of CYP1A2 regulation have been studied by many groups. Direct interactions between transcription factors with the promoters of the gene represent one of the primary means by which the expression of CYP1A2 is controlled. In this review, several important cis elements, transcription factors and the effects of deacetylation/methylation of promoter regions that play an important role in the induction by PAHs as well as constitutive expression of human CYP1A2 are discussed.

Prediction of Transonic Buffet Onset for a Supercritical Airfoil with Shock-Boundary Layer Interactions Using Navier-Stokes Solver

Chung, Injae The Korean Society for Aeronautical and Space Scie 2017 International Journal of Aeronautical and Space Sc Vol.18 No.1

To predict the transonic buffet onset for a supercritical airfoil with shock-boundary layer interactions, a practical steady approach has been proposed. In this study, it is assumed that the airfoil flow is steady even when buffet onset occurs. Steady Navier-Stokes computations are performed on the supercritical airfoil. Using the aerodynamic parameters calculated from Navier-Stokes solver, various steady approaches for predicting buffet onset are discussed. Among the various steady approaches considered in this study, Thomas' criterion based on Navier-Stokes computation has shown to be the most appropriate indicator of identifying the buffet onset for a supercritical airfoil with shock-boundary layer interactions. Good agreements have been obtained compared with the results of unsteady transonic wind tunnel tests. The present method is shown to be reliable and useful for transonic buffet onset for a supercritical airfoil with shock-boundary layer interactions in terms of practical engineering viewpoint.