Expression of bovine cytosolic 5'-nucleotidase (cN-II) in yeast: nucleotide pools disturbance and its consequences on growth and homologous recombination

PLoS One. 2013 May 17;8(5):e63914. doi: 10.1371/journal.pone.0063914. Print 2013.

Abstract

Cytosolic 5'-nucleotidase II is a widespread IMP hydrolyzing enzyme, essential for cell vitality, whose role in nucleotide metabolism and cell function is still to be exactly determined. Cytosolic 5'-nucleotidase overexpression and silencing have both been demonstrated to be toxic for mammalian cultured cells. In order to ascertain the effect of enzyme expression on a well-known eukaryote simple model, we expressed cytosolic 5'-nucleotidase II in Saccharomyces cerevisiae, which normally hydrolyzes IMP through the action of a nucleotidase with distinct functional and structural features. Heterologous expression was successful. The yeast cells harbouring cytosolic 5'-nucleotidase II displayed a shorter duplication time and a significant modification of purine and pyrimidine derivatives concentration as compared with the control strain. Furthermore the capacity of homologous recombination in the presence of mutagenic compounds of yeast expressing cytosolic 5'-nucleotidase II was markedly impaired.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 5'-Nucleotidase / metabolism*
  • Analysis of Variance
  • Animals
  • Blotting, Western
  • Cattle / metabolism*
  • Cell Division / physiology
  • Cytosol / enzymology*
  • DNA Primers / genetics
  • Electrophoresis, Polyacrylamide Gel
  • Homologous Recombination / physiology*
  • Nucleotides / analysis*
  • Saccharomyces cerevisiae / growth & development*
  • Saccharomyces cerevisiae / metabolism
  • Temperature

Substances

  • DNA Primers
  • Nucleotides
  • 5'-Nucleotidase

Grants and funding

This work was financially supported by a grant from the REGIONE AUTONOMA DELLA SARDEGNA, L.R. 07/08/2007, CRP 3360 (TITLE: Modulation of expression by inducible silencing; heterologous expression in S. cerevisiae; two-hybrid system. Three models to make clear the physiological role of cytosolic 5′nucleotidase II). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.