Spatial Organization of Calcium Signaling Involved in Cell Volume Control in the Fucus Rhizoid

Abstract

Subprotoplasts prepared from different regions of rhizoid and thallus cells of Fucus zygotes displayed mechanosensitive plasma membrane channels in cell-attached patch-clamp experiments by using laser microsurgery. In excised patches, this channel was found to be voltage gated, carrying K+ outward and Ca2+ inward, with a relative permeability of Ca2+/K+ of 0.35 to 0.5, and an increased open probability at membrane potentials more positive than -80 mV. No significant difference was found in the density of this channel type from different regions of rhizoid or thallus cells. Hypoosmotic treatment of intact zygotes induced dramatic transient elevations of cytoplasmic Ca2+, initiating at the rhizoid apex and propagating in a wavelike manner to subapical regions. Localized initiation of the Ca2+ transient correlated with greater osmotic swelling at the rhizoid apex compared with other regions of the zygote. Ca2+ transients exhibited a refractory period between successive hypoosmotic shocks, during which additional transients could not be elicited and the ability to osmoregulate was impaired. Buffering the Ca2+ transients with microinjected Br2BAPTA similarly reduced the ability of rhizoid cells to osmoregulate. Ca2+ influx was associated with the initiation of the Ca2+ transient in apical regions, whereas intracellular sources contributed to its propagation. Thus, localized signal transduction is patterned by interactions of the cell wall, plasma membrane, and intracellular Ca2+ stores.

Laser ablation of the cell wall and localized patch clamping of the plasma membrane in the filamentous fungus Aspergillus: characterization of an anion‐selective efflux channel

“…Silanisation of the pipettes enhanced seal formation. Treating pipette tips with 0-001 % (w/v) poly-L-lysine has been reported to enhance seal formation to the plasma membrane of Fucus (Taylor et al, 1996) though this treatment did not enhance seal formation in the present study. To reduce pipette capacitance, electrodes were coated with a 50 % (w/w) mixture of mineral oil (Sigma, Poole, UK) and Parafilm® (American National Can., Chicago, IL).…”

“…Laser ablation of the cell wall to expose the plasma membrane has been used as a tool for investigations of algal cells (Taylor & Brownlee, 1992;DeBoer et al, 1994;Taylor et al, 1996) and higher plant cells (Kurkdjian et al, 1993;Henricksen et al, 1995); though only two studies have then successfully applied the patch-clamp technique to characterize ion channels in the exposed plasma membrane (Henricksen et al, 1995;Taylor et al, 1996). In the present study, we have shown that it is feasible to use the laser ablation technique to produce plasma membrane-bound blebs at defined locations along the hyphae of Aspergillus.…”

“…laser is used to ablate the cell wall and expose the plasma membrane without the use of cell walldegrading enzymes circumvents these problems (Taylor & Brownlee, 1992). To date, the laser ablation approach has been successfully used in conjunction with the patch clamp technique to study plasma membrane ion channels in Fucus rhizoid cells (Taylor et al., 1996) and guard cells (Henricksen et al, 1995). Significantly, the study on guard cells revealed K"^ channel activity previously unreported in guard cell protoplasts derived by enzymaticdegradation of the cell wall.…”

Imaging of the Yellow Cameleon 3.6 Indicator Reveals That Elevations in Cytosolic Ca2+ Follow Oscillating Increases in Growth in Root Hairs of Arabidopsis

“…The relationship between the ROS/ hyperpolarization-activated Ca 21 channel thought to support the gradient in root hairs (Foreman et al, 2003) and such a mechanical response remains to be defined. It is probable that more than one Ca 21 -permeable channel exists at the tips of root hairs, similar to the tip-growing rhizoids of Fucus, which contain two Ca 21 -permeable channels, one with and one without mechanosensitivity (Taylor et al, 1996). Integration of the activity of these channels may well lie at the heart of the system that must precisely balance the promotion and restriction of turgor-driven expansion to permit root hair elongation without runaway expansion and the associated catastrophic failure of the apical wall.…”

Spatial characteristics to calcium signalling; the calcium wave as a basic unit in plant cell calcium signalling

“…This initiation site is an important model for other plant cells. Taylor et al (1996) used laser microsurgery and patchclamping on the Fucus rhizoid tip plasma membrane. Voltage-regulated and mechano-sensitive channels that carry Ca 2+ were detected in the plasma membrane of the tip.…”