Vacuolar iron transporters mediate resistance to triadimefon in plant pathogenic fungi

Abstract

Abstract Triadimefon resistance in Puccinia striiformis f. sp. tritici ( Pst ), the fungal pathogen of wheat stripe rust, is increasingly observed in China, but the mechanisms beyond cytochrome P450 14α-demethylase ( Cyp51 ) mutations remain unknown. Through bulked segregant analysis and RNA-seq of a sexual Pst population, we identify three vacuolar iron transporter genes ( CCC1 ), PstCCC1.1 , PstCCC1.2 and PstCCC1.3 , as key triadimefon sensitivity determinants. Functional analyses show that RNAi-mediated silencing of these genes reduce fungicide resistance, while heterologous expression of natural mutants ( PstCCC1.2 R82C,S86P,E109K and PstCCC1.3 R306C,S310P,E333K ) in yeast and Fusarium graminearum reduce cytosolic iron content and thereby increase resistance to triadimefon. Combined microscale thermophoresis (MST) and isothermal titration calorimetry (ITC) show that these two mutants exhibit increased iron affinities, linking specific amino acid changes to the detoxification ability. This iron-mediated resistance mechanism is conserved across fungal species and represents a triazole resistance pathway independent of Cyp51, with implications for managing fungal diseases, especially stripe rust.