The testa of higher plant seeds protects the embryo against adverse environmental conditions. Its role is assumed mainly by controlling germination through dormancy imposition and by limiting the detrimental activity of physical and biological agents during seed storage. To analyze the function of the testa in the model plant Arabidopsis, we compared mutants affected in testa pigmentation and/or structure for dormancy, germination, and storability. The seeds of most mutants exhibited reduced dormancy. Moreover, unlike wild-type testas, mutant testas were permeable to tetrazolium salts. These altered dormancy and tetrazolium uptake properties were related to defects in the pigmentation of the endothelium and its neighboring crushed parenchymatic layers, as determined by vanillin staining and microscopic observations. Structural aberrations such as missing layers or a modified epidermal layer in specific mutants also affected dormancy levels and permeability to tetrazolium. Both structural and pigmentation mutants deteriorated faster than the wild types during natural aging at room temperature, with structural mutants being the most strongly affected.

Influence of the Testa on Seed Dormancy, Germination, and Longevity in Arabidopsis

Plant protoplasts show physiological perceptions and responses to hormones, metabolites, environmental cues, and pathogen-derived elicitors, similar to cell-autonomous responses in intact tissues and plants. The development of defined protoplast transient expression systems for high-throughput screening and systematic characterization of gene functions has greatly contributed to elucidating plant signal transduction pathways, in combination with genetic, genomic, and transgenic approaches.

/pdf/signal-transduction-in-maize-and-arabidopsis-mesophyll-4b65wac81t.pdf

Signal Transduction in Maize and Arabidopsis Mesophyll Protoplasts

THE molecular mechanisms by which Ca2+ controls a variety of ion transport-associated cellular functions in higher plant cells1, including movements of stomatal pores, remain unknown. Stomatal pores regulate the gas exchange in leaves. Openings of stomatal pores are mediated by an increase in the intracellular potassium and anion content of guard cells2,3. Voltage-dependent K+ channels4–7 and hyperpolarizing H+ pumps8,9 have been identified as mechanisms controlling stomatal movements. But depolarizing mechanisms required for K+ efflux during stomatal closing remain unknown. Indirect evidence suggests that Ca2+ triggers stomatal closing and inhibits stomatal opening10–12. Using patch-clamp techniques13, we show here that elevated (micromolar) concentrations of cytosolic Ca2+ in guard cells block inward rectifying K+ channels. Furthermore, elevation of cytosolic Ca2+ leads to the activation of a voltage-dependent depolarizing conductance with a permeability to anions. The Ca2+-induced modulation of ion channels reported here could provide a molecular basis for Ca2+-dependent regulation of stomatal movements in leaves.

Cytosolic calcium regulates ion channels in the plasma membrane of Vicia faba guard cells

Article de synthese sur les mecanismes moleculaires du processus de transport des ions dans les cellules des plantes superieures

The Physiology of ION Channels and Electrogenic Pumps in Higher Plants

Biosensors incorporate a biological sensing element that converts a change in an immediate environment to signals conducive for processing. Biosensors have been implemented for a number of applications ranging from environmental pollutant detection to defense monitoring. Biosensors have two intriguing characteristics: (1) they have a naturally evolved selectivity to biological or biologically active analytes; and (2) biosensors have the capacity to respond to analytes in a physiologically relevant manner. In this paper, molecular biosensors, based on antibodies, enzymes, ion channels, or nucleic acids, are briefly reviewed. Moreover, cell-based biosensors are reviewed and discussed. Cell-based biosensors have been implemented using microorganisms, particularly for environmental monitoring of pollutants. Biosensors incorporating mammalian cells have a distinct advantage of responding in a manner that can offer insight into the physiological effect of an analyte. Several approaches for transduction of cellular signals are discussed: these approaches include measures of cell metabolism, impedance, intracellular potentials, and extracellular potentials. Among these approaches, networks of excitable cells cultured on microelectrode arrays are uniquely poised to provide rapid, functional classification of an analyte and ultimately constitute a potentially effective cell-based biosensor technology. Three challenges that constitute barriers to increased cell-based biosensor applications are presented: analytical methods, reproducibility, and cell sources. Possible future solutions to these challenges are discussed.

Development and application of cell-based biosensors.

Leaflet movements in Samanea saman are driven by the shrinking and swelling of cells in opposing (extensor and flexor) regions of the motor organ (pulvinus). Changes in cell volume, in turn, depend upon large changes in motor cell content of K+, Cl− and other ions. We performed patch-clamp experiments on extensor and flexor protoplasts, to determine whether their plasma membranes contain channels capable of carrying the large K+ currents that flow during leaflet movement. Recordings in the “whole-cell” mode reveal depolarization-activated K+ currents in extensor and flexor cells that increase slowly (t½ = ca. 2 seconds) and remain active for minutes. Recordings from excised patches reveal a single channel conductance of ca. 20 picosiemens in both cell types. The magnitude of the K+ currents is adequate to account quantitatively for K+ loss, previously measured in vivo during cell shrinkage. The K+ channel blockers tetraethylammonium (5 millimolar) or quinine (1 millimolar) blocked channel opening and decreased light- and dark-promoted movements of excised leaflets. These results provide evidence for the role of potassium channels in leaflet movement.

Potassium channels in motor cells of Samanea saman: a patch-clamp study

The patch-clamp technique was used to study passive movements of ions through the plasmalemma of wheat leaf protoplasts. This method overcomes the problems inherent in conventional electrophysiological study of plant cells. Changes in conductance were recorded in patches excised from the plasmalemma. Two types of patches were observed: (i) regions of low channel density, where discrete single-channel currents could be resolved and conductance ranged from 10 to 200 picosiemens and (ii) regions of high channel density, where single-channel currents could not be resolved and conductance was on the order of a few nanosiemens. The results indicate a striking similarity between animal and plant cell membranes in the basic phenomena of transport. Moreover, the approach used constitutes a new degree of refinement in the study of processes of regulation, pathology, and toxicity in plants.

http://science.sciencemag.org/content/sci/226/4676/835.full.pdf

Ion channels in plasmalemma of wheat protoplasts.

By using both optical and mechano-electric detectors, we have shown that the squid giant axon swells when an action potential is generated. The maximum swelling is reached at the peak of the action potential. The undershoot of the membrane potential is associated with a marked shrinkage of the axon. We have also demonstrated these mechanical changes in axons from which a major portion of the axoplasm has been removed. We have examined the effects of changing the tonicity of the external medium and of applying several chemical reagents.

/pdf/rapid-pressure-changes-and-surface-displacements-in-the-52cut617yc.pdf

Rapid pressure changes and surface displacements in the squid giant axon associated with production of action potentials.

A nerve impulse travelling along a crustacean nerve was found to be accompanied by a small, rapid movement of the nerve surface.The movement was 10-20nm in amplitude and was concurrent with a rise in the “swelling pressure” of the order of 5mg/cm2 for a nerve bundle.Initiation of an action potential at the site of cathodal polarization was preceded by a small, slow mechanical change in the nerve fiber.Anodal polarization produced a large mechanical change of the opposite sign.Tetrodotoxin and procaine suppressed rapid mechanical changes.

/pdf/mechanical-changes-in-crab-nerve-fibers-during-action-2mpdb070bv.pdf

Mechanical changes in crab nerve fibers during action potentials.

Mechanical changes in the squid giant axon associated with the production of an action potential are examined further by using piezoelectric and optical methods. The peak of swelling of the axon coincides with the peak of the action potential recorded internally at the site of mechanical recording. Mechanical changes produced by a train of action potentials do not summate. Repetitively fired action potentials induced by lowering the external Ca-ion concentration are preceded by a gradual swelling of the axon. An inward current through the membrane causes shrinkage and an outward current produces swelling of the axon. An inward current enhances and an outward current depresses the mechanical changes associated with the action potential. There is a transient shortening followed by an elongation of the axon when an action potential travels along the axon. It is argued that the experimental results obtained are consistent with the colloid chemical, or macromolecular, theory of excitation.

https://www.jstage.jst.go.jp/article/jjphysiol1950/32/4/32_4_505/_pdf

Kunihiko Iwasa

Papers

Potassium channels in motor cells of Samanea saman: a patch-clamp study

Ion channels in plasmalemma of wheat protoplasts.

Rapid pressure changes and surface displacements in the squid giant axon associated with production of action potentials.

Mechanical changes in crab nerve fibers during action potentials.

Further studies of rapid mechanical changes in squid giant axon associated with action potential production.