Asymmetrical dose responses shape the evolutionary trade-off between antifungal resistance and nutrient use

Antimicrobial resistance is an emerging threat for public health. The success of resistance mutations depends on the trade-off between the benefits and costs they incur. This trade-off is largely unknown and uncharacterized for antifungals. Here, we systematically measure the effect of all amino acid substitutions in the yeast cytosine deaminase Fcy1, the target of the antifungal 5-fluorocytosine (5-FC, flucytosine). We identify over 900 missense mutations granting resistance to 5-FC, a large fraction of which appear to act through destabilization of the protein. The relationship between 5-FC resistance and growth sustained by cytosine deamination is characterized by a sharp trade-off, such that small gains in resistance universally lead to large losses in canonical enzyme function. We show that this steep relationship can be explained by differences in the dose–response functions of 5-FC and cytosine. Finally, we observe the same trade-off shape for the orthologue of FCY1 in Cryptoccocus neoformans , a human pathogen. Our results provide a powerful resource and platform for interpreting drug target variants in fungal pathogens as well as unprecedented insights into resistance–function trade-offs. Studying all amino acid substitutions in the yeast cytosine deaminase Fcy1, the target of the antifungal 5-FC, the authors show a sharp trade-off between 5-FC resistance and growth sustained by cytosine deamination.