Dataset Fig6A_BOR1_3D-SCLIM
Project
411-Yoshinari-BorateTransporter
Title
3D-time-lapse images of vacuolar sorting of BOR1
Description
Plants maintain nutrient homeostasis by controlling the activities and abundance of nutrient transporters. In Arabidopsis thaliana, the borate (B) transporter BOR1 plays a role in the efficient translocation of B under low-B conditions. BOR1 undergoes polyubiquitination in the presence of sufficient B and is then transported to the vacuole via multivesicular bodies (MVBs) to prevent B accumulation in tissues at a toxic level. A previous study indicated that BOR1 physically interacts with mu subunits of adaptor protein complexes AP-3 and AP-4, both involved in vacuolar sorting pathways. In this study, we investigated the roles of AP-3 and AP-4 subunits in BOR1 trafficking in Arabidopsis. The lack of AP-3 subunits did not affect either vacuolar sorting or polar localization of BOR1-GFP, whereas the absence of AP-4 subunits resulted in a delay in high-B-induced vacuolar sorting without affecting polar localization. Super-resolution microscopy revealed a rapid sorting of BOR1-GFP into AP-4-positive spots in the trans-Golgi network (TGN) upon high-B supply. These results indicate that AP-4 is involved in sequestration of ubiquitinated BOR1 into a TGN-specific subdomain 'vacuolar-trafficking zone', and is required for efficient sorting of MVB and vacuole. Their findings have thus helped elucidate the rapid vacuolar sorting process facilitated by AP-4 in plant nutrient transporters.
Funding
JSPS Grant-in-Aid for Scientific Research on Innovative Areas (19H05763 to J.T.), JSPS Grant-in-Aid for Scientific Research
(A) (19H00934 to J.T.), JSPS Grant-in-Aid for Young Scientists
(A) (26712007 to J.T.), Grant-in-Aid for Scientific Research(B)
(22H02643 to T.U.), Grant-in-Aid for Challenging Research
(Exploratory) (22K19327 to T.U.), JST CREST (JPMJCR20E5 to T.U.), JST PRESTO (JPMJPR22D9 to A.Y.) and Asahi Glass Foun-
dation (to T.U.).
Title
Z-series and time-lapse SCLIM images of BOR1-GFP and AP4M-TagRFP in the root epidermal cells after treatment with high concentration of boric acid
Description
Z-series and time-lapse SCLIM (super-resolution confocal live-imaging microscopy) images of BOR1(borate transporter)-GFP and AP4M-TagRFP in the root epidermal cells after treatment with high (100 micromolar) concentration of boric acid. The images are also included that were processed using deconvolution algorithms with theoretical point-spread functions
in Volocity software.
Description
The roots of Arabidopsis thaliana grown on MGRL medium contain-
ing 0.5 micromolar boric acid, 1.5% gellan gum and 1% sucrose for 4 or 5 days.
Extrinsic variables
BOR1-GFP and AP4M-TagRFP were exogenously expressed.
A. thaliana roots were treated with 100 micromolar boric acid at the start of imaging.
Gene ontology: Biological processes
Gene ontology: Cellular components
Gene ontology: Molecular functions
active borate transmembrane transporter activity (
GO:0046715)
Protein names
Boron transporter 1 (
Q8VYR7)
AP-4 complex subunit mu (
Q9SB50)
Genetic methods
Agrobacterium-mediated floral-dip transformation
Dimensions
160x190x9x2x10,
160x190x9x2x10
Visualization methods
Green fluorescent protein from Aequorea
TagRFP
Contrast enhancing methods
Resolution enhancing methods
super resolution confocal live imaging
microscopy (SCLIM)
Sample preparation methods
living tissue
Body
Olympus-Evident IX73
Objective
Olympus-Evident UPLSAPO100XS