Electrophile response element-mediated induction of the cystine/glutamate exchange transporter gene expression

J Biol Chem. 2002 Nov 22;277(47):44765-71. doi: 10.1074/jbc.M208704200. Epub 2002 Sep 13.

Abstract

In mammalian cultured cells, the cystine/glutamate exchange transport mediated by system x(c)- is important to maintain intracellular GSH levels. System x(c)- consists of two protein components, xCT and the heavy chain of 4F2 antigen. The activity of system x(c)- is induced by various stimuli, including electrophilic agents like diethyl maleate. In the present study, we have investigated the mechanism of the transcriptional regulation of xCT mRNA by diethyl maleate. The xCT gene consisted of twelve exons and sequence analysis identified four electrophile response element (EpRE)-like sequences between -230 and -1 in the 5'-flanking region, designated EpRE-1 to EpRE-4. To identify sequences mediating the constitutive and induced expression of xCT, a series of 5'-deletion mutants created from the 5'-flanking region were cloned into a luciferase reproter vector and transfected into BHK21 cells. The 5'-deletion analysis revealed that the sequence between -116 and -82 is essential for the basal expression and the sequence between -226 and -116 containing EpRE-1 is essential in response to diethyl maleate. Mutational analysis demonstrated that EpRE-1 is critically involved in the response to diethyl maleate. Other stress agents like arsenite, cadmium, and hydroquinone seemed to induce system x(c)- activity via the same sequence. Furthermore, the experiments using the mouse embryonic fibroblasts derived from the Nrf2-deficient mice revealed that the induction of xCT gene by electrophilic agents is mediated by Nrf2. EpRE occurs in a broad spectrum of genes for the proteins that are involved in the defense against xenobiotics and regulates their expression. The present results have demonstrated that xCT is a novel member of this protein family.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 5' Flanking Region
  • Amino Acid Transport System y+*
  • Animals
  • Base Sequence
  • Buthionine Sulfoximine / metabolism
  • Carrier Proteins / genetics*
  • Carrier Proteins / metabolism*
  • Cell Line
  • Cricetinae
  • Cysteine / metabolism
  • Cystine / metabolism
  • DNA-Binding Proteins / metabolism
  • Enzyme Inhibitors / metabolism
  • Fibroblasts / cytology
  • Fibroblasts / metabolism
  • Fusion Regulatory Protein 1, Heavy Chain / genetics
  • Fusion Regulatory Protein 1, Heavy Chain / metabolism
  • Gene Expression Regulation*
  • Genes, Reporter
  • Glutathione / metabolism
  • Humans
  • Maleates / metabolism*
  • Mice
  • Mutation
  • NF-E2-Related Factor 2
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Response Elements / genetics*
  • Sequence Analysis, DNA
  • Trans-Activators / metabolism

Substances

  • Amino Acid Transport System y+
  • Carrier Proteins
  • DNA-Binding Proteins
  • Enzyme Inhibitors
  • Fusion Regulatory Protein 1, Heavy Chain
  • Maleates
  • NF-E2-Related Factor 2
  • NFE2L2 protein, human
  • Nfe2l2 protein, mouse
  • Recombinant Fusion Proteins
  • SLC7A11 protein, human
  • Slc7a11 protein, mouse
  • Trans-Activators
  • Cystine
  • Buthionine Sulfoximine
  • diethyl maleate
  • Glutathione
  • Cysteine