The Adenosine Analog Tubercidin Inhibits Glycolysis in Trypanosoma brucei as Revealed by an RNA Interference Library

Mark E. Drew, James C. Morris, Zefeng Wang, Lance Wells, Marco Sanchez, Scott M. Landfear, Paul T. Englund

Research output: Contribution to journalArticle

50 Scopus citations

Abstract

We used an RNA interference (RNAi) library in a forward genetic selection to study the mechanism of toxicity of tubercidin (7-deazaadenosine) to procyclic Trypanosoma brucei. Following transfection of cells with an RNAi-based genomic library, we used 5 μM tubercidin to select a drug-resistant cell line. Surprisingly, we found in these resistant cells that the hexose transporters had been silenced. We subsequently found that silencing of hexokinase, a glycolytic enzyme, also yielded tubercidin-resistant parasites. These observations suggested that glycolysis could be a target of tubercidin action and that RNAi silencing of glycolytic enzymes was gradual enough to allow the parasites to adapt to alternative sources of energy. Indeed, adaptation of procyclic trypanosomes to a glucose-independent metabolism by reduction of glucose in the culture medium caused tubercidin resistance. High pressure liquid chromatography analysis of glycolytic intermediates from parasites treated with tubercidin showed a dose-dependent increase in concentration of 1,3-bisphosphoglycerate, a substrate of phosphoglycerate kinase. Furthermore, tubercidin triphosphate inhibited recombinant T. brucei phosphoglycerate kinase activity in vitro with an IC50 of 7.5 μM. We conclude that 5 μM tubercidin kills trypanosomes by targeting glycolysis, especially by inhibition of phosphoglycerate kinase.

Original languageEnglish (US)
Pages (from-to)46596-46600
Number of pages5
JournalJournal of Biological Chemistry
Volume278
Issue number47
DOIs
StatePublished - Nov 21 2003

    Fingerprint

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this