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bioRxiv · 10.1101/342345

TOL proteins mediate vacuolar sorting of the borate transporter BOR1 in Arabidopsis thaliana.

Yoshinari, A.·Korbei, B.·Takano, J.
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Abstract

Boron (B) is an essential micronutrient for plants, however, it shows cytotoxicity at high concentrations. A borate transporter BOR1 is required for efficient transport of boron (B) toward the root stele in Arabidopsis thaliana. BOR1 shows polar localization in the plasma membrane of various root cells toward the stele-side under B limitation. To avoid over-accumulation of B, BOR1 in the plasma membrane is rapidly internalized and transported into the vacuole for proteolysis after high-B supply in an ubiquitination-dependent manner. Although BOR1 has been predicted to be transported into multi-vesicular bodies/late endosomes (MVB/LEs) via the endosomal sorting complex required for transport (ESCRT) machinery, experimental evidence was absent so far. In this study, we investigated the intracellular localization of BOR1 by visualizing endomembrane compartments, and tested the involvement of ESCRT-0-like proteins TOM1-LIKEs (TOLs) in the vacuolar sorting of BOR1. Under low-B conditions, a large portion of cytoplasmic BOR1 was localized in the trans-Golgi networks/early endosomes (TGN/EEs) labeled with VHA-a1 subunit. Pharmacological interference of endosomal recycling using brefeldin A induced colocalization of BOR1 with RabA5D, which labels recycling vesicles associated with the TGN. These data suggest that BOR1 cycles between plasma membrane and TGN/EE via RabA5D-positive endomembrane compartments under low-B conditions. On the other hand, under high-B conditions, BOR1 was localized in the inside of TOL5-positive MVB/LEs. To examine the roles of TOL proteins in intracellular trafficking of BOR1, we analyzed BOR1-GFP localization in the TOL quintuple mutant (tolQ; tol2-1tol3-1tol5-1tol6-1tol9-1) after high-B supply. In the tolQ mutant, vacuolar sorting of BOR1 was delayed, while the polar localization of BOR1 was not disturbed. Taken together, BOR1 is constantly transported to the TGN/EE by endocytosis and recycled to the plasma membrane likely via RabA5D-positive endomembrane compartments under low-B conditions. On the other hand, BOR1 is transported to the vacuole via TOL5-positive MVB/LEs under high-B conditions. TOL proteins are required for sorting of ubiquitinated BOR1 into MVB/LE for vacuolar degradation but not for the polar trafficking of BOR1.

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Yoshinari, A., Korbei, B., Takano, J.. 2018-06-08. TOL proteins mediate vacuolar sorting of the borate transporter BOR1 in Arabidopsis thaliana.. https://doi.org/10.1101/342345

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