Deletion of can1/cat1 genes and expression of a dominant any1 mutation establish an effective canavanine selection in fission yeast

Abstract

Positive and counter-selectable markers have been successfully integrated as a part of numerous genetic assays in many model organisms. In this study, we investigate the mechanism of resistance to arginine analog canavanine and its applicability for genetic selection in Schizosaccharomyces pombe. Deletion of both arginine permease genes cat1 and can1 provides strong drug resistance, while the single can1 deletion does not have impact on canavanine resistance. Surprisingly, the widely used can1-1 allele does not match to the can1 gene but rather corresponds to the any1-523C>T allele. The strong canavanine-resistance conferred by this allele arises from an inability to deposit basic amino acid transporters on the cellular membrane. any1-523C>T leads to reduced post-translational modifications of Any1 regulated by the Tor2 kinase. We also demonstrate that any1-523C>T is a dominate allele. Our results uncover the mechanisms of canavanine-resistance in fission yeast and open the opportunity of using cat1, can1 and any1 mutant alleles in genetic assays.