Changes in Surface Area of Intact Guard Cells Are Correlated with Membrane Internalization

Abstract

Guard cells must maintain the integrity of the plasma membrane as they undergo large, rapid changes in volume. It has been assumed that changes in volume are accompanied by changes in surface area, but mechanisms for regulating plasma membrane surface area have not been identified in intact guard cells, and the extent to which surface area of the guard cells changes with volume has never been determined. The alternative hypothesis-that surface area remains approximately constant because of changes in shape-has not been investigated. To address these questions, we determined surface area for intact guard cells of Vicia faba as they underwent changes in volume in response to changes in the external osmotic potential. We also estimated membrane internalization for these cells. Epidermal peels were subjected to external solutions of varying osmotic potential to shrink and swell the guard cells. A membrane-specific fluorescent dye was used to identify the plasma membrane, and confocal microscopy was used to acquire a series of optical paradermal sections of the guard cell pair at each osmotic potential. Solid digital objects representing the guard cells were created from the membrane outlines identified in these paradermal sections, and surface area, volume, and various linear dimensions were determined for these solid objects. Surface area decreased by as much as 40% when external osmotic potential was increased from 0 to 1.5 MPa, and surface area varied linearly with volume. Membrane internalization was approximated by determining the amount of the fluorescence in the cell's interior. This value was shown to increase approximately linearly with decreases in the cell's surface area. The changes in surface area, volume, and membrane internalization were reversible when the guard cells were returned to a buffer solution with an osmotic potential of approximately zero. The data show that intact guard cells undergo changes in surface area that are too large to be accommodated by plasma membrane stretching and shrinkage and suggest that membrane is reversibly internalized to maintain cell integrity.Guard cells regulate stomatal pore size to allow CO 2 uptake while controlling water loss. To accomplish this, they respond to environmental factors such as light and CO 2 concentration and to chemical signals such as ABA, which may originate in other parts of the plant. Guard cells respond to these factors by regulating ion fluxes across the plasma membrane, and the resulting movement of water causes changes in cell volume, turgor pressure, and shape, leading to changes in the pore aperture. The changes in guard cell turgor pressure and volume caused by these processes can be quite large. In Vicia faba, guard cell turgor pressure has been shown to vary between 1.0 and 4.0 MPa (Franks et al., 1998(Franks et al., , 2001) as stomata go from closed to wide open, and volume has been shown to change by as much as 40% (Raschke and Dickerson, 1973;Franks et al., 2001). These changes can occur over a time period of minutes.An often-un...

Binding of SEC11 Indicates Its Role in SNARE Recycling after Vesicle Fusion and Identifies Two Pathways for Vesicular Traffic to the Plasma Membrane

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Mechanics of surface area regulation in cells examined with confined lipid membranes

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Effects of hydrostatic pressure on mouse sperm

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