The Effect of UDP-glucuronosyltransferase 1A1 Expression on the Mutagenicity and Metabolism of the Cooked-Food Carcinogen 2-Amino-1-methyl-6-phenylimidazo[4-5,b]pyridine in CHO cells Metadata

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Title

  • Main Title The Effect of UDP-glucuronosyltransferase 1A1 Expression on the Mutagenicity and Metabolism of the Cooked-Food Carcinogen 2-Amino-1-methyl-6-phenylimidazo[4-5,b]pyridine in CHO cells

Creator

  • Author: Malfatti, M A
    Creator Type: Personal
  • Author: Wu, R W
    Creator Type: Personal
  • Author: Felton, J S
    Creator Type: Personal

Contributor

  • Sponsor: United States. Department of Energy.
    Contributor Type: Organization
    Contributor Info: US Department of Energy (United States)

Publisher

  • Name: Lawrence Livermore National Laboratory
    Place of Publication: Livermore, California
    Additional Info: Lawrence Livermore National Lab., Livermore, CA (United States)

Date

  • Creation: 2004-08-13

Language

  • English

Description

  • Content Description: UDP-glucuronosyltransferase proteins (UGT) catalyze the glucuronidation of both endogenous and xenobiotic compounds. In previous studies UGT1A1 has been implicated in the detoxification of certain food-borne-carcinogenic-heterocyclic amines. To determine the importance of UDP-glucuronosyltransferase 1A1 (UGT1A1) in the biotransformation of the cooked-food carcinogen 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), genetically modified CHO cells that are nucleotide excision repair-deficient, and express cytochrome P4501A2 (UV5P3 cell line) were transfected with a cDNA plasmid of human UGT1A1 to establish the UDPglucuronosyltransferase 1A1 expressing 5P3hUGT1A1 cell line. Expression of the UGT1A1 gene was verified by screening neogene expressing clonal isolates (G-418 resistant) for their sensitivity to cell killing from PhIP exposure. Five of eleven clones were chosen for further analysis due to their resistance to cell killing. Western blot analysis was used to confirm the presence of the UGT1A1 and CYP1A2 proteins. All five clones displayed a 52 kDa protein band, which corresponded to a UGT1A1 control protein. Only four of the clones had a protein band that corresponded to the CYP1A2 control protein. Correct fragment size of the cDNAs in the remaining 4 clones was confirmed by RT-PCR and quantification of the mRNA product was accomplished by real-time RT-PCR. Expression of UGT1A1 in the transfected cells was 10{sup 4}-10{sup 5} fold higher relative to the UV5P3 parental cells. One clone (No.14) had a 10 fold higher increase in expression at 1.47 x 10{sup 5} over the other three clones. This clone was also the most active in converting N-hydroxy-PhIP to N-hydroxy-PhIP glucuronide conjugates in microsomal metabolism assays. Based on the D{sub 50} values, the cytotoxic effect of PhIP was decreased {approx}350 fold in the 5P3hUGT1A1 cells compared to the UV5P3 control cells. In addition no significant increase in mutation frequency was observed in the transfected cells. These results clearly indicate that UGT1A1 plays a critical role in PhIP biotransformation, providing protection against PhIP mediated cytotoxicity and mutagenicity.
  • Physical Description: page(s) pp. 205-214

Subject

  • Keyword: Cytochromes
  • Keyword: Cell Killing
  • Keyword: Plasmids
  • Keyword: Amines
  • Keyword: Carcinogens
  • Keyword: Genes
  • Keyword: Cho Cells
  • STI Subject Categories: 59 Basic Biological Sciences
  • Keyword: Glucuronide Conjugates
  • Keyword: Nucleotides
  • Keyword: Proteins
  • Keyword: Sensitivity
  • Keyword: Mutation Frequency
  • Keyword: Xenobiotics
  • Keyword: Metabolism
  • Keyword: Detoxification

Source

  • Journal Name: Mutation Research; Journal Volume: 570; Other Information: Publication date January 18, 2005; PDF-FILE: 33 ; SIZE: 0.3 MBYTES

Collection

  • Name: Office of Scientific & Technical Information Technical Reports
    Code: OSTI

Institution

  • Name: UNT Libraries Government Documents Department
    Code: UNTGD

Resource Type

  • Article

Format

  • Text

Identifier

  • Report No.: UCRL-JRNL-207729
  • Grant Number: W-7405-ENG-48
  • Office of Scientific & Technical Information Report Number: 15015880
  • Archival Resource Key: ark:/67531/metadc1418172
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