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Mitogen activated protein kinase kinase kinase 18 (MAPKKK18) mediated signaling cascade plays important roles in Arabidopsis drought stress tolerance. However, the post-translational modu-lation patterns of MAPKKK18 are not characterized. In this study, we found that the protein level of MAPKKK18 was tightly controlled by the 26S pro-teasome. Ubiquitin ligases RGLG1 and RGLG2 ubiquitinated MAPKKK18 at lysine residue K32 and K154, and promoted its degradation. Deletion of RGLG1 and RGLG2 stabilized MAPKKK18 and fur-ther enhanced the drought stress tolerance of MAPKKK18-overexpression plants. Our data dem-onstrate that RGLG1 and RGLG2 negatively regulate MAPKKK18-mediated drought stress tolerance in Arabidopsis.