Yellow stripe-like (YSL) transporters are required for the transportation of metal-phytosiderophores and are structurally related to metal-nicotianamine complexes. Some studies also reported the involvement of YSL transporters in pathogen-induced defense. However, the molecular mechanisms of YSL genes involved in biotic stress responses are still not clear, especially in cereal crops. This study aimed to functionally characterize TaYS1A during the interaction of wheat and Puccinia striiformis f. sp. tritici (Pst), the causal agent of stripe rust disease. TaYS1A was localized in the cell membrane of wheat protoplasts and Nicotiana benthamiana cells. TaYS1A was significantly up-regulated in wheat leaves after being infected with the avirulent Pst isolate CYR23 and after treatment with salicylic acid (SA). Silencing of TaYS1A by the virus-induced gene silencing method enhanced the susceptibility of wheat to Pst accompanied by reducing the accumulation of SA and H2O2 and down-regulating the transcriptions of TaPR1 and TaPR2. In addition, TaYS1A was found to interact with TaNH2, a homolog of OsNH2, by yeast-two-hybrid assay, and silencing of TaYS1A diminished the expression of TaNH2. Our findings suggested the existence of positive regulation of TaYS1A in providing resistance against Pst by modulating SA-induced signaling and offered new insight into the biological role of YSL in wheat against pathogens.