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Focus on "Effect of expressing the water channel aquaporin-1 on the CO₂ permeability of Xenopus oocytes"

Department of Physiology and Biophysics, University of Texas Medical Branch, Galveston, Texas 77555-0641

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OVER TWENTY YEARS of biophysical studies have provided strong evidence that, in some biological membranes, water transport is in part through proteinaceous water-conducting pores (reviewed in Ref. 1). This evidence includes the demonstration of 1) a high ratio of osmotic-to-diffusive water permeability coefficients, 2) a low activation energy for water permeation (expected for permeation by a watery pathway, in contrast with the high activation energy typical of lipid permeation), and 3) inhibition by mercury compounds. The major breakthrough in this field occurred in 1992, when Agre and his co-workers (5) demonstrated that the red blood cell membrane protein CHIP28 was a water pore. Soon thereafter, it was shown that CHIP28, now called aquaporin-1 (AQP1), is expressed in epithelial cells of high osmotic water permeability, such as proximal tubule and descending limb of Henle's loop in the kidney. Now a family of aquaporins has been identified, which spans species of the plant and animal kingdoms. Several of these aquaporins are expressed in a tissue-specific fashion in mammals and have distinct properties. AQP1 is a tetramer whose structure has been established at 6 Å resolution by cryo-electron microscopy (6). The amino acid residues in the permeation pathway have been identified and include a cysteine that is clearly the target site of the effect of mercury. Details of these elegant studies can be found in several recent reviews (e.g., Ref. 3).

Functional studies of AQP1 have revealed a high permeability to water, impermeability to small ions, and, at most, very low permeabilities to small nonelectrolytes. This is consistent with the fact that this channel behaves as if water moved in single file, i.e., the permeating path is so narrow that water molecules cannot pass each other. Therefore, the flux of individual molecules within the pore depends on the flux of other water molecules, and this fact explains the high ratio of (osmotic/diffusive) permeability coefficients (see Ref. 1). Higher-resolution crystallographic data are needed to establish the structural bases of the high water selectivity of AQP1.

Because of the lack of small-ion permeation (including protons), likely related to charges in the pore wall or pore-access regions, studies of nonelectrolyte fluxes through AQP1 might be quite informative. Two interesting questions are, first, whether neutral solutes of dimensions similar to that of water can be used to ascertain the dimensions of the permeation pathway, and second, whether permeation of one or more of these solutes can have physiological significance. Nakhoul et al. (Ref. 4; see p. C543 in this issue) address this problem by testing directly the effect of exogenous expression of AQP1 on CO₂ permeation in Xenopus laevis oocytes, cells that have a low baseline CO₂ permeability. To eliminate CO₂ hydration as the rate-limiting step in the intracellular acidification elicited by elevating extracellular PCO₂, the oocytes were injected with AQP1 cRNA and also carbonic anhydrase. The CO₂ permeability was estimated from the rate of change of intracellular pH in response to changes in bathing medium PCO₂. The result was a significant increase in CO₂ permeability that, according to cited preliminary experiments, is mercury sensitive. As stated by the authors, these results could be explained by CO₂ permeation via AQP1 itself or by a stimulatory effect of AQP1 expression on CO₂ permeability via the lipid bilayer or another, endogenous water channel. The mercury sensitivity supports but does not prove the first hypothesis. Hence additional studies are needed to prove that CO₂ permeation is in fact via AQP1.

The CO₂ molecule is linear, its axial cross-sectional area being smaller than that of the water molecule. Hence CO₂ permeation of AQP1 is not surprising if water is permeable. However, the result is important from a biophysical point of view, because it provides a marker for AQP1-mediated transport that, under certain conditions, would permit more rapid or convenient measurements. In addition, it suggests that the lipid bilayer is not the sole permeation pathway for this gas.

Perhaps the most important contribution of the work of Nakhoul et al. (4) is the hypothesis that CO₂ transport via AQP1 is physiologically important. This remains to be tested in cell types with high expression of this protein but is suggested by the fact that CO₂ permeability of sterol-containing lipid membranes is of the order of 0.35 cm/s (2). In the oocyte, Nakhoul et al. (4) estimated that the CO₂ permeability increased by ~2 cm/s. Comparison of these two numbers supports the notion that a parallel, AQP1-mediated CO₂ permeation pathway may increase significantly the CO₂ permeability coefficient of at least some biological membranes. Admittedly, teleological arguments are not conducive to proof by themselves. However, it is of interest to note that AQP1 is highly expressed in cell types that "need" a high water permeability, such as those in the proximal nephron, and also in cells that need a high CO₂ permeability, such as the apical membrane of the proximal tubule, the capillary endothelium, and the red blood cell membrane. The latter two barriers are involved in CO₂ permeation in alveolus-capillary and tissue-capillary exchange. Additional studies are needed to prove or disprove this hypothesis, but the elegant work of Nakhoul et al. provides an exciting beginning.

	REFERENCES

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1.   Finkelstein, A. Water Movement Through Lipid Bilayers, Pores, and Plasma Membranes: Theory and Reality. New York: Wiley, 1987. (Soc. Gen. Physiol. Distinguished Lecture Ser., vol. 4)

2.   Gutknecht, J., M. A. Bisson, and F. C. Tosteson. Diffusion of carbon dioxide through lipid bilayer membranes: effects of carbonic anhydrase, bicarbonate, and unstirred layers. J. Gen. Physiol. 69: 779-794, 1977[Abstract/Free Full Text].

3.   King, L. S., and P. Agre. Pathophysiology of the aquaporin water channels. Annu. Rev. Physiol. 58: 619-648, 1996[Medline].

4.   Nakhoul, N. L., B. A. Davis, M. F. Romero, and W. F. Boron. Effect of expressing the water channel aquaporin-1 on the CO₂ permeability of Xenopus oocytes. Am. J. Physiol. 274 (Cell. Physiol. 43): C543-C548, 1998[Abstract/Free Full Text].

5.   Preston, G. M., T. P. Carroll, W. B. Guggino, and P. Agre. Appearance of water channels in Xenopus oocytes expressing red cell CHIP28 protein. Science 256: 385-387, 1992[Abstract/Free Full Text].

6.   Walz, T., T. Hirai, K. Murata, J. B. Heymann, K. Mitsuoka, Y. Fujiyoshi, B. L. Smith, P. Agre, and A. Engel. The three-dimensional structure of aquaporin-1. Nature 387: 624-626, 1997[Medline].