DRP1-Dependent Endocytosis is Essential for Polar Localization and Boron-Induced Degradation of the Borate Transporter BOR1 inArabidopsis thaliana

Abstract

Boron (B) is essential for plants but toxic in excess. The borate efflux transporter BOR1 is expressed in various root cells and localized to the inner/stele-side domain of the plasma membrane (PM) under low-B conditions. BOR1 is rapidly degraded through endocytosis upon sufficient B supply. The polar localization and degradation of BOR1 are considered important for efficient B translocation and avoidance of B toxicity, respectively. In this study, we first analyzed the subcellular localization of BOR1 in roots, cotyledons and hypocotyls, and revealed a polar localization in various cell types. We also found that the inner polarity of BOR1 is established after completion of cytokinesis in the root meristem. Moreover, variable-angle epifluorescence microscopy visualized BOR1-green fluorescent protein (GFP) as particles in the PM with significant lateral movements but in restricted areas. Importantly, a portion of BOR1-GFP particles co-localized with DYNAMIN-RELATED PROTEIN 1A (DRP1A), which is involved in scission of the clathrin-coated vesicles, and they disappeared together from the PM. To examine the contribution of DRP1A-mediated endocytosis to BOR1 localization and degradation, we developed an inducible expression system of the DRP1A K47A variant. The DRP1A variant prolonged the residence time of clathrin on the PM and inhibited endocytosis of membrane lipids. The dominant-negative DRP1A blocked endocytosis of BOR1 and disturbed its polar localization and B-induced degradation. Our results provided insight into the endocytic mechanisms that modulate the subcellular localization and abundance of a mineral transporter for nutrient homeostasis in plant cells.

Inroads into Internalization: Five Years of Endocytic Exploration

“…Consistent with their function in endocytosis, DRP1s colocalize with clathrin in dynamic PM CCPs (Konopka et al, 2008;Fujimoto et al, 2010), and drp1 mutants display abnormal CME (Collings et al, 2008). BOR1-GFP, which is polarly localized on the inner/stele-side PM of root cells, colocalizes with DRP1A, and inducible expression of the dominant negative DRP1A K47A mutant inhibits BOR1-GFP endocytosis, boron-induced degradation, and polar localization (Yoshinari et al, 2016).…”

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

“…Arabidopsis thaliana Col-0 ecotype harboring the pBOR1:BOR1-GFP construct (Takano et al 2010; Yoshinari et al 2016) and organelle marker constructs mCherry-Got1p (Wave18R), mCherry-RabE1D (Wave27R), mCherry-RabA5D (Wave24R), mCherry-RHA1 (Wave7R, Geldner et al 2009), VHA-a1-mRFP (Viotti et al 2010), pTOL5:TOL5-mCherry (Korbei et al 2013) were generated by crossing. Plants in F1 or F2 generation were used for confocal imaging.…”

“…indicating that BOR1 undergoes constitutive endocytosis. In Arabidopsis, DYNAMIN-RELATED PROTEIN 1A (DRP1A), which is involved in clathrin-mediated endocytosis, is required for constitutive endocytosis and polar localization of BOR1 (Yoshinari et al 2016).…”

Altered Expression of a Malate-Permeable Anion Channel, OsALMT4, Disrupts Mineral Nutrition

“…OsBOR1 is required for B uptake and accumulation, especially when B is limited, because reducing OsBOR1 expression reduces B uptake and B loading into the xylem (Nakagawa et al, 2007). In B-deficient Arabidopsis plants, AtBOR1 transporters are expressed in the pericycle and endodermal cells of roots, where they also transport B to the xylem (Yoshinari et al, 2016). Interestingly, members of this family of transporters also contribute to B tolerance in plants, as illustrated by HvBOT1 in barley.…”