Laboratory evolution of copper tolerant yeast strains.
BACKGROUND
Yeast strains endowed with robustness towards copper and/or enriched in intracellular Cu might find application in biotechnology processes, among others in the production of functional foods. Moreover, they can contribute to the study of human diseases related to impairments of copper metabolism. In this study, we investigated the molecular and physiological factors that confer copper tolerance to strains of baker's yeasts.


RESULTS
We characterized the effects elicited in natural strains of Candida humilis and Saccharomyces cerevisiae by the exposure to copper in the culture broth. We observed that, whereas the growth of Saccharomyces cells was inhibited already at low Cu concentration, C. humilis was naturally robust and tolerated up to 1 g · L-1 CuSO4 in the medium. This resistant strain accumulated over 7 mg of Cu per gram of biomass and escaped severe oxidative stress thanks to high constitutive levels of superoxide dismutase and catalase. Both yeasts were then "evolved" to obtain hyper-resistant cells able to proliferate in high copper medium. While in S. cerevisiae the evolution of robustness towards Cu was paralleled by the increase of antioxidative enzymes, these same activities decreased in evolved hyper-resistant Candida cells. We also characterized in some detail changes in the profile of copper binding proteins, that appeared to be modified by evolution but, again, in a different way in the two yeasts.


CONCLUSIONS
Following evolution, both Candida and Saccharomyces cells were able to proliferate up to 2.5 g · L-1 CuSO4 and to accumulate high amounts of intracellular copper. The comparison of yeasts differing in their robustness, allowed highlighting physiological and molecular determinants of natural and acquired copper tolerance. We observed that different mechanisms contribute to confer metal tolerance: the control of copper uptake, changes in the levels of enzymes involved in oxidative stress response and changes in the copper-binding proteome. However, copper elicits different physiological and molecular reactions in yeasts with different backgrounds.
Dipartimento di Biotecnologie e Bioscienze, Università degli Studi di Milano-Bicocca, Piazza della Scienza 2, 20126 Milano, Italy.
Giusy Manuela
GM
Adamo
Stefania
S
Brocca
Simone
S
Passolunghi
Benedetto
B
Salvato
Marina
M
Lotti
D005075:Biological Evolution; D018533:Biomass; D002175:Candida / Q000201:enzymology; D002374:Catalase / Q000235:genetics / Q000378:metabolism; D003300:Copper / Q000494:pharmacology*; D018384:Oxidative Stress; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000187:drug effects* / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D013482:Superoxide Dismutase / Q000235:genetics / Q000378:metabolism
D011994:Recombinant Proteins; D029701:Saccharomyces cerevisiae Proteins; D003300:Copper; D002374:Catalase; D013482:Superoxide Dismutase