Salinity stress progressively reduces plant growth and productivity, while plant has developed complex signaling pathways to confront salt stress. However, only a few genetic variants have been identified to mediate salt tolerance in the major crop rice, and the molecular mechanism remains poorly understood. Here, we identify ten candidate genes associated with salt-tolerance (ST) traits by performing a genome-wide association analysis in rice landraces. We characterize two ST-related genes, encoding transcriptional factor OsWRKY53 and Mitogen-activated protein Kinase Kinase OsMKK10.2, that mediate root Na+flux and Na+homeostasis. We further find that OsWRKY53 acts as a negative modulator regulating expression ofOsMKK10.2in promoting ion homeostasis. Furthermore, OsWRKY53 trans-repressesOsHKT1;5(high-affinity K+transporter 1;5), encoding a sodium transport protein in roots. We show that the OsWRKY53-OsMKK10.2 and OsWRKY53-OsHKT1;5 module coordinate defenses against ionic stress. The results shed light on the regulatory mechanisms underlying plant salt tolerance.