Effects of mGST A4 transfection on 4-hydroxynonenal-mediated apoptosis and differentiation of K562 human erythroleukemia cells

Ji Zhong Cheng, Sharad S. Singhal, Manjit Saini, Jyotsana Singhal, John T. Piper, F. J.G.M. Van Kuijk, Piotr Zimniak, Yogesh C. Awasthi, Sanjay Awasthi

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Cellular levels of downstream products of membrane lipid oxidation appear to regulate differentiation in K562 human erythroleukemia cells. 4- Hydroxynonenal (4-HNE) is a diffusible and relatively stable product of peroxidation of arachidonic and linoleic acids, cellular levels of which are regulated through metabolism to glutathione (GSH) conjugate by glutathione S- transferases (GSTs). A group of immunologically related α-class mammalian GSTs expressed in mice (mGST A4-4), rat (rGST A4-4), human (hGST A5.8), and other species, as well as the more distantly related human hGST A4-4, preferentially utilize 4-HNE as a substrate and are suggested to be major determinants of intracellular levels of 4-HNE. Present studies were designed to examine the effects of 4-HNE on K562 cells and to study the effect of transfection of mGSTA4-4 in these cells. Exposure of K562 cells to 20 μM 4- HNE for 2 h resulted in a rapid erythroid differentiation of K562 cells, as well as apoptosis evidenced by characteristic DNA laddering. Stable transfection of cells with mGST A4-4 resulted in a fivefold increase in GST- specific activity toward 4-HNE compared with wild-type or vector-only transfected cells. The mGST A4-4-transfected cells were resistant to the cytotoxic, apoptotic, and differentiating effects of 4-HNE. The mGST A4 transfection also conferred resistance to direct oxidative stress (IC50 of H2O2 22, 23, and 35 μM for wild-type, vector-transfected, and mGST A4- transfected cells, respectively), mGST A4-4-transfected cells also showed a higher rate of proliferation compared with wild-type or vector-transfected K562 cells (doubling time 22.1 ± 0.7, 31 ± 1.2, and 29 ± 0.6 h, respectively). Cellular 4-HNE levels determined by mass spectrometry were lower in mGST A4-4-transfected cells compared to cells transfected with vector alone (5.9 pmol/5 x 107 cells and 62.9 pmol/5 x 107 cells, respectively). Our studies show that 4-HNE can induce erythroid differentiation in K562 cells and that overexpression of mGST A4 suppresses 4-HNE levels and inhibits erythroid differentiation and apoptosis.

Original languageEnglish (US)
Pages (from-to)29-36
Number of pages8
JournalArchives of Biochemistry and Biophysics
Issue number1
StatePublished - Dec 1 1999

Bibliographical note

Funding Information:
These studies were supported in part by NIH Grants CA77495 (SA), CA 27967 (YCA), ES 07804 (PZ), and Research to Prevent Blindness (FJGM).


  • 4-hydroxynonenal
  • Apoptosis
  • Differentiation
  • Erythroleukemia
  • GST
  • K562


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