Effects of lactate dehydrogenase suppression and glycerol-3-phosphate dehydrogenase overexpression on cellular metabolism

Dae won Jeong, Il Taeg Cho, Tae Soo Kim, Gun Won Bae, Ik Hwan Kim, Ick Young Kim

Research output: Contribution to journalArticle

30 Citations (Scopus)

Abstract

In order to conduct a physiological functional study of lactate dehydrogenase (LDH) and glycerol-3-phosphate dehydrogenase (GPDH), we engineered a CHO dhfr- cell, by overexpressing either the anti-sense LDH-A RNA (anti-LDH cells) or GPDH (GP3 cells), or both (GP3/ anti-LDH cells). LDH activity in the cell cytosol, and lactate content and pHe change in the growth media were found to decrease according to the order: cell lines GP3/anti-LDH > anti-LDH > GP3 > CHO. Intracellular ATP contents, representing the extent of respiration rate, also decreased, according to a rank order as follows: GP3 > CHO > GP3/anti-LDH > anti-LDH. We also attempted to identify and characterize any physiological changes occurring in the cells which harbored diverse metabolic pathways. First, anti-LDH cells with heightened respiration rates were found to display a higher degree of sensitivity to the prooxidant tert-butyl hydroperoxide (tBOOH), and the mitochondrial complex III inhibitor, antimycin A, than the GPDH-expressing cells (GP3 and GP3/anti-LDH), which have a lower respiration rate. Second, the anti-sense LDH-A RNA-expressing cells (anti-LDH and GP3/anti-LDH) evidenced a higher degree of resistance to apoptosis by cell-cell contact inhibition, and a faster doubling time (∼19 h compared with 2∼6 h) than the CHO and GP3 cells. Additionally, cell growth in an extended culture under HCO-3- free conditions to induce a steep acidification could be maintained with the anti-sense LDH-A RNA-expressing cells, but could not be maintained with the CHO and GP3 cells. Third, we observed that the most appropriate cell line for the optical production of a certain therapeutic protein (Tissue-Plasminogen Activator) was the GP3/anti-LDH cells. Collectively, our data indicate a variety of physiological roles for LDH and GPDH, including cellular acidosis, oxidoresistance, apoptosis by both acidosis and cell-cell contact inhibition, cell growth, and the generation of recombinant proteins.

Original languageEnglish
Pages (from-to)1-8
Number of pages8
JournalMolecular and Cellular Biochemistry
Volume284
Issue number1-2
DOIs
Publication statusPublished - 2006 Mar

Keywords

  • Acidosis
  • Apoptosis
  • Cellgrowth
  • Glycolysis
  • Oxidoresistance
  • Recombinant protein production
  • Respiration rate

ASJC Scopus subject areas

  • Molecular Biology
  • Clinical Biochemistry
  • Cell Biology

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