The effects of altered membrane fatty acid composition on the toxic interactions of heavy
metals with Saccharomyces cerevisiae were examined. Saccharomyces cerevisiae was
enriched with the polyunsaturated fatty acids (PUFAs) linoleate (18:2) and linolenate
(18:3) by growth in 18:2- or 18:3-supplemented medium. Incorporation of the exogenous
PUF As resulted in them comprising greater than 65% and 40% of the total fatty acids in
whole-cell and plasma membrane lipids, and nuclear membrane lipids, respectively.
Incorporation of the exogenous PUF As had no discernible adverse effects on cell
division. However, inhibition of cell division in the presence of Cd(N03)2 was
accentuated by growth in the presence of the di-unsaturated fatty acid linoleate.
Furthermore, susceptibility to both Cd2+ - and Cu2+ -induced plasma membrane
permeabilisation and whole cell toxicity was markedly accentuated in PUF A-enriched
cells, and increased with the degree of fatty acid unsaturation. The increased sensitivity
ofPUFA-enriched cells to membrane permeabilisation and whole-cell toxicity was
correlated with increased levels of lipid peroxidation in these cells. Cu2+ - and Cd2+_
induced lipid peroxidation was rapid and associated with a decline in plasma membrane
lipid order, detected by fluorescence depolarization measurements. Levels of the lipid
peroxidation products thiobarbituric acid-reactive substances (TBARS) and conjugated
dienes were markedly higher in PUF A-enriched cells, compared with unsupplemented
cells, following exposure to cadmium or copper. Thus, lipid peroxidation was
demonstrated as a major means of heavy metal toxicity in a microorganism for the first
time. In addition, the effects ofPUFA-enrichment on the interactions of heavy metals
with cellular nucleic acids were examined. Exposure ofPUFA-enriched cells to the
redox-active metals chromium and copper resulted in the uncoupling of DNA synthesis
from cell division, leading to sequential S phases. For example, DNA levels of up to 8C
were evident in 18:3-enriched cells after only 4.5 h exposure to 100 JJ.M Cu(N03h.
Using flow cytometry, the heterogeneity in susceptibility to copper toxicity of
exponential phase S. cerevisiae was also examined. Susceptibility towards copper
toxicity was demonstrated to be cell cycle stage-dependent, whereby G2/M phase cells
were found to be the most susceptible towards copper toxicity. Staining with the oxidantsensitive
probe 2',7' -dichlorodihydrofluorescein diacetate (H2DCFDA) revealed that the
greater copper sensitivity of G2/M phase cells correlated with elevated endogenous levels
of reactive oxygen species in these cells.
Howlett, Niall G.
Faculty of Health and Life SciencesDepartment of Biological and Medical Sciences
Howlett, Niall G.
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