Some SLY1 alleles require multiple substitutions to suppress the lethal uso1∆ phenotype. (A–D) Locations of amino acid substitutions in four representative SLY1 alleles recovered in our screen. (E) Growth phenotypes show that most single substitutions are unable to suppress the loss of Uso1. Many of the same single mutants suppress the loss of Ypt1 (see Table S1). The multisite allele SLY1-5c, although retrieved in our primary screen, was unable to suppress the uso1∆ allele in secondary screening.