Plant Rho GTPase ROP6 Is Essential for Manganese Homeostasis in <i>Arabidopsis</i>

Manganese (Mn) is an indispensable mineral for plant growth and development. However, plants cultivated in acidic and poorly drained soils are vulnerable to Mn²⁺ toxicity due to its heightened increased bioavailability. Despite the crucial roles of the Rho of plant (ROP) GTPases in various cellular processes, their precise function in regulating Mn homeostasis remains elusive. In this study, we unveil a novel ROP6 GTPase signalling pathway that profoundly influences Mn phytotoxicity tolerance in Arabidopsis. Remarkably, the rop6 and dominant-negative ROP6 (rop6^DN) mutant plants displayed a dramatically sensitive phenotype to Mn toxicity, whereas ROP6-overexpression and constitutively activated ROP6 (rop6^CA) lines exhibited enhanced Mn stress tolerance. Immunoblot analysis corroborated that the ROP6 protein, especially the active form of ROP6, increased in abundance in the presence of high Mn levels. Further, we identified that ROP6 physically interacted and colocalized with Metal Tolerance Protein 8 (MTP8) in vivo. Mn transport complementation assays in yeast, combined with biochemical analyses, emphasized the essentiality of ROP6 for MTP8's transport activity. In addition, genetic analyses indicated that ROP6 acted upstream of MTP8 in the regulatory cascade. Collectively, our findings elucidate that ROP6 GTPase signalling positively modulates and enhances Mn stress tolerance in plants.