Biology subjects

Grunwald, Y.

Publications and source records attributed to Grunwald, Y..

5 recordsLinked to original sources

bioRxiv2023

A tale of two pumps: Blue light and ABA alter Arabidopsis leaf hydraulics via bundle sheath cells H+-pumps and channels

This study focuses on the cellular mechanism underlying the co-regulation of the leaf hydraulic conductance (Kleaf) by blue light (BL) and the stress hormone ABA in Arabidopsis thaliana. Our previous work has demonstrated that (1) Kleaf increased by BL signaling within the leaf bundle sheath cells (BSCs), which activated their plasmalemma (PM) H+-ATPase (AHA2), acidifying the xylem sap; (2) external acidification enhanced the BSCs Kleaf and their osmotic water permeability (Pf); (3) ABA decreased both Kleaf and Pf by reducing the BSCs PM aquaporins activity. We now show, using pH and EM (membrane potential) probes combined with H+-pumps inhibitors and manipulations of cytosolic and external Ca2+ concentrations ([Ca2+]CYT, [Ca2+]EXT, respectively), that, in the BSCs: (a) under BL, ABA inhibits AHA2, depolarizing the BSCs and alkalinizing the xylem sap, (b) ABA stimulates the BSCs vacuolar H+-ATPase (VHA), alkalinizing their cytosol; (c) each pump stimulation, AHA2 by BL and VHA by ABA, requires [Ca2+]CYT elevation. ABA-effect-mimicking conditions in patch-clamp experiments activate the BSCs K+-release channels (SKOR and/or GORK). ABA decreased the Kleaf of skor mutants less than WTs, while during water deprivation stress, skor plants transpired more and their leaves lost relatively less K+ than WT. This suggests a role for SKOR in water conservation under drought.

plant biology↗

bioRxiv2021

Stress is basic: ABA alkalinizes both the xylem sap and the cytosol of Arabidopsis vascular bundle sheath cellsby inhibiting their P-type H+-ATPaseand stimulating their V-type H+-ATPase

BACKGROUND AND HYPOTHESIS* Under water deprivation, in many perennial species, the stress hormone, ABA, appears in the xylem sap in the shoot (including leaf) veins and the xylem sap pH (pHEXT) increases. This study aimed to test the hypothesis that ABA is the signal for an altered proton balance of the leaf-vein-enwrapping bundle sheath cells (BSCs). METHODS* Plant Material. We used a few Arabidopsis thaliana (L.) Heynh. genotypes: wildtype (WT) of two accessions, Landsberg erecta (Ler) and Columbia (Col), and a few mutants and transformants in these backgrounds. * H+-Pumps activities. We monitored ABA effects on the H+-pump activities in the BSCs cytosol-delimiting membranes (plasma membrane and tonoplast) by monitoring the cytosol and the xylem pH, and the membrane potential (EM), by imaging the fluorescence of pH- and membrane potential (EM)-reporting probes: (a) the BSCs pHEXT - with the ratiometric fluorescent dye FITC-dextran petiole-fed into detached leaves in unbuffered xylem perfusion solution (XPS), (b) the BSCs pHCYT - with the ratiometric dye SNARF1 loaded into BSCs isolated protoplasts, and (c) the BSCs EM - with the ratiometric dye di- 8-ANEPPS. RESULTS* ABA increased the pHEXT; this response was abolished in an abi1-1 mutant with impaired signaling via a PP2C (ABI1) and in an aha2-4 mutant with knocked-down AHA2; * ABA depolarized the WT BSCs; * ABA increased pHCYT irrespective of AHA2 activity (i.e., whether or not AHA was inhibited by vanadate, or in the aha2-4 mutant); * The ABA-induced cytosol alkalinization was abolished in the absence of VHA activity (i.e., when VHA was inhibited by bafilomycin A1, or in the vha-a2 vha-a3 double mutant with inactive VHA); * All these results resemble the ABA effect on GCs; * In contrast to GCs, AHA2 and not AHA1 is the ABA major target in BSCs; * Blue light (BL) enabled the response of the BSCs VHA to ABA; * The ABA- and BL-signaling pathways acting on both BSCs pumps, AHA2 and VHA, are likely to be BSCs autonomous, based on (a) the presence in the BSCs of many genes of the ABA- and BL-signaling pathways and (b) ABA responses (depolarization and pHCYT elevation) demonstrated under BL in isolated protoplasts. SIGNIFICANCE STATEMENTWe reveal here an alkalinizing effect of the plant drought-stress hormone ABA on the pH on both sides of the plasmalemma of the vein-enwrapping bundle sheath cells (BSCs), due to ABA inhibition of the BSCs AHA2, the plasmalemma H+- ATPase and stimulation of VHA, their vacuolar H+-ATPase. Since pH affects the BSCs selective regulation of solute and water fluxes into the leaf, these H+- pumps may be attractive targets for manipulations aiming to improve plant drought response.

plant biology↗

bioRxiv2020

A High Throughput Gravimetric Phenotyping Platform for Real Time Physiological Screening of Plant Environment Dynamic Responses

Food security for the growing global population is a major concern. The data provided by genomic tools far exceeds the supply of phenotypic data, creating a knowledge gap. To meet the challenge of improving crops to feed the growing global population, this gap must be bridged. Physiological traits are considered key functional traits in the context of responsiveness or sensitivity to environmental conditions. Many recently introduced high-throughput phenotyping techniques are based on remote sensing or imaging and are capable of directly measuring morphological traits, but measure physiological parameters only indirectly. This paper describes a method for direct physiological phenotyping that has several advantages for the functional phenotyping of plant-environment interactions. It aims to help users overcome the many challenges encountered in the use of load-cell gravimetric systems and pot experiments. The suggested techniques will enable users to distinguish between soil weight, plant weight and soil water content, providing a method for continuous and simultaneous measurement of dynamic soil, plant and atmosphere conditions, alongside key physiological traits. This method allows researchers to closely mimic field stress scenarios while taking into consideration the environments effect on the plants physiology. This method also minimizes pot effects, which are one of the major problems in pre-field phenotyping. It includes a feedback fertigation system that enables a truly randomized experimental design with a field-like plant density. This system detects the soil-water content limiting threshold ({theta}) and allows for the translation of data into knowledge through the use of a real-time analytic tool and an online statistical resource. This method for the rapid and direct measurement of the physiological responses of multiple plants to a dynamic environment has great potential for use in screening for beneficial traits associated with responses to abiotic stress, in the context of pre-field breeding and crop improvement. SUMMARYThis high-throughput, whole-plant water relations gravimetric phenotyping method enables direct and simultaneous real-time measurements and analysis of multiple yield-related physiological traits involved in dynamic plant-environment interactions.

plant biology↗

bioRxiv2019

Out of the blue; Phototropins of the leaf vascular bundle sheath mediate the blue light regulation of the leaf hydraulic conductance

The Arabidopsis leaf veins bundle sheath cells (BSCs) - a selective barrier to water and solutes entering the mesophyll - increase the leaf radial hydraulic conductance (Kleaf) by acidifying the xylem sap by their plasma membrane H+-ATPase, AHA2. Based on this and on the BSCs expression of PHOT1 and PHOT2, and the known blue-light (BL)-induced Kleaf increase, we hypothesized that, resembling the guard cells, BL perception by the BSCs phots activates its H+-ATPase, which, consequently, upregulates Kleaf. Indeed, under BL, the Kleaf of the knockout mutant lines phot1-5, phot2-1, phot1-5phot2-1, and aha2-4 was lower than that of the WT. BSC-only-directed complementation of phot1-5 or aha2-4 by PHOT1 or AHA2, respectively, restored the BL-induced Kleaf increase. BSC-specific silencing of PHOT1 or PHOT2 prevented such Kleaf increase. A xylem-fed kinase inhibitor (tyrphostin 9) replicated this also in WT plants. White light - ineffective in the phot1-5 mutant - acidified the xylem sap (relative to darkness) in WT and in the PHOT1-complemented phot1-5. These results, supported by BL increase of BSC protoplasts water permeability and cytosolic pH and their hyperpolarization by BL, identify the BSCs as a second phot-controlled water conductance element in leaves, in series with stomatal conductance. Through both, BL regulates the leaf water balance. One-Sentence summaryBlue light regulates the leaf hydraulic conductance via the bundle-sheath cells blue light PHOT receptors which, via an autonomous signaling pathway, activate the BSCs AHA2 H+-pump.

plant biology↗

bioRxiv2017

Bundle-sheath cells are internal leaf "water valves" controlled via xylem acidification by H+-ATPase

O_LIThe leaf vascular bundle sheath cells (BSCs) that tightly envelop the leaf veins, are a selective and dynamic barrier to xylem-sap water and solutes radially entering the mesophyll cells. Under normal conditions, xylem-sap pH of <6 is presumably important for driving and regulating the transmembranal solute transport. Having discovered recently a differentially high expression of a BSCs proton pump, AHA2, we now test the hypothesis that it regulates this pH and leaf radial water fluxes. C_LIO_LIWe monitored the xylem-sap pH in the veins of detached leaves of WT Arabidopsis, AHA mutants, and aha2 mutants complemented with AHA2 gene solely in BSCs. We tested an AHA inhibitor and stimulator, and different pH buffers. We monitored their impact on the xylem-sap pH and the whole leaf hydraulic conductance (Kleaf), and the effect of pH on the water osmotic permeability (Pf) of isolated BSCs protoplasts. C_LIO_LIOur results demonstrated that AHA2 is necessary for xylem-sap acidification, and in turn, for elevating Kleaf. Conversely, knocking out AHA2 alkalinized the xylem-sap. Also, elevating xylem sap pH to 7.5 reduced Kleaf and elevating external pH to 7.5 decreased the BSCs Pf. C_LIO_LIAll these demonstrate a causative link between AHA2 activity in BSCs and leaf radial water conductance. C_LI One-sentence summaryBundle-sheath cells can control the leaf hydraulic conductance by proton-pump-regulated xylem sap pH

plant biology↗