文档介绍：The KEEP ON GOING Protein of Arabidopsis Recruits
the ENHANCED DISEASE RESISTANCE1 Protein to
Trans-work/Early Endosome Vesicles1[W][OA]
Yangnan Gu and Roger W. Innes*
Department of Biology, Indiana University, Bloomington, Indiana 47405–7107
Loss-of-function mutations in the Arabidopsis (Arabidopsis thaliana) ENHANCED DISEASE RESISTANCE1 (EDR1) gene confer
enhanced resistance to powdery mildew infection, enhanced senescence, and enhanced programmed cell death under both
abiotic and biotic stress conditions. All edr1-mediated phenotypes can be suppressed by a specific missense mutation (keg-4)in
the KEEP ON GOING (KEG) gene, which encodes a multidomain protein that includes a RING E3 ligase domain, a kinase
domain, ankyrin repeats, and HERC2-like (for HECT and RCC1-like) repeats. The molecular and cellular mechanisms
underlying this suppression are poorly understood. Using confocal laser scanning microscopy and fluorescent protein fusions,
we determined that KEG localizes to trans-work/early endosome (TGN/EE) vesicles. Both the keg-4 mutation, which
is located in the carboxyl-terminal HERC2-like repeats, and deletion of the entire HERC2-like repeats reduced endosomal
localization of KEG and increased localization to the endoplasmic reticulum and cytosol, indicating that the HERC2-like
repeats facilitate the TGN/EE targeting of KEG. EDR1 colocalized with KEG to the TGN/EE when coexpressed but localized
primarily to the endoplasmic reticulum when expressed alone. Yeast two-hybrid and coimmunoprecipitation analyses
revealed that EDR1 and KEG physically interact. Deletion of the HERC2-like repeats abolished the interaction between KEG
and EDR1 as well as the KEG-induced TGN/EE localization of EDR1, indicating that the recruitment of EDR1 to the TGN/EE
is based on a direct interaction between EDR1 and KEG mediated by the HERC2-like repeats. Collectively, these data suggest
that EDR1 and KEG function together to regulate endocytic trafficking and/or the fo