Biology subjects

Dragwidge, J. M.

Publications and source records attributed to Dragwidge, J. M..

4 recordsLinked to original sources

bioRxiv2025

Secretory carrier membrane proteins assist with aquaporin trafficking in Arabidopsis.

Aquaporins belonging to the plasma membrane intrinsic protein (PIP) subfamily are channel proteins that control water flow across cells, allowing plants to rapidly adjust hydraulic conductivity and thereby sustain growth, gas exchange, and recovery from drought. We show that secretory carrier membrane proteins (SCAMP) regulate the abundance of the aquaporins and thereby water transport in Arabidopsis root cells. SCAMPs are evolutionarily conserved multi-spanning transmembrane proteins. In animal cells, they function in secretion, endocytosis and autophagy. Knowledge on their role in plants is restricted to localization and trafficking experiments in heterologous systems and few genetic perturbation experiments. Here, we analysed all five members of the Arabidopsis SCAMP family. We identified conserved tyrosine motifs assisting in transport to the plasma membrane and N-terminally located NPF motifs that are required for internalization. SCAMPs dimerize both at the plasma membrane and endosomes, and dimerization is required for their internalization. Functionally, several PIPs were identified as common targets of multiple SCAMP isoforms. Triple and quintuple scamp mutants show mild developmental delay under standard growth conditions, but they are less sensitive to drought-induced leaf wilting. This observation cannot be explained by altered stomatal dynamics or densities, nor by differential soil water content. However, scamp mutant root protoplasts contain less PIPs and swell less under hypotonic conditions compared to wild type, indicating reduced PIP plasma membrane levels. In conclusion, our research identifies the SCAMP membrane trafficking proteins as regulators of PIP abundance at the plasma membrane in root cells. We propose that the reduced PIP levels in the scamp mutants act as a priming mechanism, allowing them to respond better to conditions of reduced water availability.

plant biology↗

bioRxiv2022

AtEH/Pan1 proteins drive phase separation of the TPLATE complex and clathrin polymerisation during plant endocytosis

Clathrin-mediated endocytosis (CME) is an essential cellular internalisation pathway involving the dynamic assembly of clathrin and accessory proteins to form membrane-bound vesicles. In plants, the evolutionarily ancient TSET/TPLATE complex (TPC) plays an essential, but not well-defined role in CME. Here, we show that two highly disordered TPC subunits, AtEH1 and AtEH2 function as scaffolds to drive biomolecular condensation of the complex. These condensates specifically nucleate on the plasma membrane through interactions with anionic phospholipids, and facilitate the dynamic recruitment and assembly of clathrin, early-, and late-stage endocytic accessory proteins. Importantly, clathrin forms ordered assemblies within the condensate environment. Biomolecular condensation therefore acts to promote dynamic protein assemblies throughout clathrin-mediated endocytosis. Furthermore, the disordered region sequence properties of AtEH1 regulate the material properties of the endocytic condensates in vivo and alteration of these material properties influences endocytosis dynamics, and consequently plant adaptive growth. HighlightsO_LIAtEH subunits are endocytic scaffolds which drive condensation of the TPC C_LIO_LIAtEH1 condensates nucleate on the plasma membrane via lipid interactions C_LIO_LICondensation of AtEH1/TPC facilitates clathrin re-arrangement and assembly C_LIO_LIAtEH1 IDR1 composition controls condensate properties to regulate endocytosis C_LI

cell biology↗

bioRxiv2022

An environmentally-responsive transcriptional state modulates cell identities during root development

Roots are fundamental organs for plant development and response to their environment: they anchor the plant to its growth substrate, uptake nutrients and water vital to plant growth, and can sense and respond to a variety of biotic and abiotic stresses. The architecture of root systems and their growth are known to be strongly affected by the environmental conditions found in the soil. However, the acquisition of cell identities at the root meristem is still mainly viewed as ontogenetically driven, where a small number of stem cells generate all the cell types through stereotyped divisions followed by differentiation, along a simple developmental trajectory. The extent to which environmental cues precisely shape and affect these developmental trajectories remains an open question. We used single-cell RNA-seq, combined with spatial mapping, to deeply explore the trajectories of cell states at the tip of Arabidopsis roots, known to contain multiple developing lineages. Surprisingly, we found that most lineage trajectories exhibit a stereotyped bifid topology with two developmental trajectories rather than one. The formation of one of the trajectories is driven by a strong and specific activation of genes involved in the responses to various environmental stimuli, that affects only of a subset of the cells in multiple cell types simultaneously, revealing another layer of patterning of cell identities in the root that is independent of cell ontogeny. We demonstrate the robustness of this environmentally responsive transcriptional state by showing that it is present in a mutant where cell type identities are greatly perturbed, as well as in different Arabidopsis ecotypes. We also show that the root can adapt the proportion of cells that acquire this particular state in response to environmental signals such as nutrient availability. The discovery of this cell state reveals new layers of cell identity that may underpin the adaptive potential of plant development.

plant biology↗

bioRxiv2018

NHX-type Na+(K+)/H+ antiporter activity is required for endomembrane trafficking and ion homeostasis in Arabidopsis thaliana

The regulation of ion and pH homeostasis of endomembrane organelles is critical for functional protein trafficking, sorting and modification in eukaryotic cells. pH homeostasis is maintained through the activity of vacuolar H+-ATPases (V-ATPases) pumping protons (H+) into the endomembrane lumen, and counter-action by cation/proton exchangers such as the NHX family of Na+(K+)/H+ exchangers. In plants, disturbing V-ATPase activity at the trans-Golgi network/early endosome (TGN/EE) impairs secretory and endocytic trafficking. However, it is unclear if the endosomal NHX-type antiporters NHX5 and NHX6 play functionally similar roles in endomembrane trafficking through maintaining ion and pH homeostasis. Here we show through genetic, pharmacological, and live-cell imaging approaches that double knockout of endosomal isoforms NHX5 and NHX6 results in impairment of endosome motility, protein recycling at the TGN/EE, but not in the secretion of integral membrane proteins. Furthermore, we report that nhx5 nhx6 mutants are partially insensitive to osmotic swelling of TGN/EE induced by the monovalent cation ionophore monensin. Similarly, nhx5 nhx6 cells are unresponsive to late endosomal swelling by the phosphatidylinositol 3/4-kinase inhibitor wortmannin, demonstrating that NHX5 and NHX6 are required for maintaining endosomal cation balance. Lastly, we report that the distal region of the cytosolic tail of NHX6 is required for mediating NHX6 localisation to late endosomes, but does not appear to be essential for NHX6 function.

plant biology↗