Data from: Too much of a good thing: The unique and repeated paths toward copper adaptation

Copper is a micronutrient essential for growth due to its role as a
co-factor in enzymes involved in respiration, defense against oxidative
damage, and iron uptake. Yet too much of a good thing can be lethal, and
yeast cells typically do not have tolerance to copper levels much beyond
the concentration in their ancestral environment. Here, we report a
short-term evolutionary study of Saccharomyces cerevisiae exposed to
levels of copper sulfate that are inhibitory to the initial strain. We
isolated and identified adaptive mutations soon after they arose, reducing
the number of neutral mutations, to determine the first genetic steps that
yeast take when adapting to copper. We analyzed 34 such strains through
whole-genome sequencing and by assaying fitness within different
environments; we also isolated a subset of mutations through tetrad
analysis of four lines. We identified a multi-layered evolutionary
response. In total, 57 single base-pair mutations were identified across
the 34 lines. In addition, gene amplification of the copper
metallothionein protein, CUP1, was rampant, as was chromosomal aneuploidy.
Four other genes received multiple, independent mutations in different
lines (the vacuolar transporter genes VTC1 and VTC4; the plasma membrane
H+-ATPase PMA1; and MAM3, a protein required for normal mitochondrial
morphology). Analyses indicated that mutations in all four genes, as well
as CUP1 copy number, contributed significantly to explaining variation in
copper tolerance. Our study thus finds that evolution takes both common
and less trodden pathways toward evolving tolerance to an essential, but
highly toxic, micronutrient.