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