Utilizing proteomic approach to identify nuclear translocation related serine kinase phosphorylation site of GNMT as downstream effector for benzo[a]pyrene

J Food Drug Anal. 2019 Apr;27(2):603-609. doi: 10.1016/j.jfda.2018.12.007. Epub 2019 Jan 8.

Abstract

Glycine N-methyltransferase (GNMT) protein is highly expressed in certain tissues, such as liver, pancreas, and prostate. GNMT serves multiple roles which include a methyl group transfer enzyme and a liver tumor suppressor. Benzo(a)pyrene (BaP), a family member of polycyclic aromatic hydrocarbon (PAH), is a known environmental carcinogen found in coal tar, tobacco smoke, barbecued food and incomplete combustion of auto fuel. BaP recruits cytochrome P450 to transform itself into benzo(a)pyrene-7,8-diol-9,10-epoxide (B(a)PDE), which covalently interacts with DNA causing tumorigenesis. BaP can be detoxified through GNMT and induces GNMT translocation into the cellular nucleus. GNMT translocation is accompanied by phosphorylation, but the role of phosphorylation in GNMT remains to be explored. Using liquid chromatography coupled with tandem mass spectrometry, this study identified serine 9 of GNMT as the phosphorylation site upon BaP treatment. When serine 9 was mutated and lost the capability to be phosphorylated, the occurrence of BaP-induced GNMT nuclear translocation was dramatically decreased. Also, this mutant from of GNMT lost the ability of phosphorylation and increased cytochrome P450 1A1 (Cyp1a) expression upon BaP treatment. In addition, protein kinase C (PKC) and c-Jun NH2-terminal kinase (JNK) may be required for such phosphorylation. Further characterization of phosphorylated GNMT for its link to BaP may bring new insights into chemical detoxification.

Keywords: Benzo(a)pyrene; GNMT; Phosphorylation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Benzo(a)pyrene / pharmacology*
  • Cells, Cultured
  • Chromatography, Liquid
  • Glycine N-Methyltransferase / analysis*
  • Glycine N-Methyltransferase / antagonists & inhibitors*
  • Glycine N-Methyltransferase / metabolism
  • HEK293 Cells
  • Hep G2 Cells
  • Humans
  • Phosphorylation / drug effects
  • Protein Serine-Threonine Kinases / analysis*
  • Protein Serine-Threonine Kinases / antagonists & inhibitors*
  • Protein Serine-Threonine Kinases / metabolism
  • Proteomics*
  • Tandem Mass Spectrometry

Substances

  • Benzo(a)pyrene
  • Glycine N-Methyltransferase
  • Protein Serine-Threonine Kinases

Grants and funding

This work was supported in part by the following grants: Program to Upgrade the R&D Capabilities of Private Universities grant (No. MOST105-2632-B-037-001), Ministry of Science and Technology of Taiwan (No. MOST104-2320-B-037-031-MY3 and MOST107-2320-B-037-003), Aim for the Top Universities Grant from Kaohsiung Medical University (No. KMU-TP104E11, KMU-TP104E14, KMU-TP104E37 and KMUTP105E08), Aim for the Top 500 Universities grant from Kaohsiung Medical University (No. KMU-DT105004, KMUDT106002), KMU Global Networking Talent Plan (No. 105KMUOR02 and 105KMUOR05) and Taiwan Protein Project (No. AS-KPQ-105-TPP). This work was also financially supported by the Research Center for Environmental Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan from The Featured Areas Research Center Program within the framework of the Higher Education Sprout Project by the Ministry of Education (MOE) in Taiwan.