Functional Water Channels Are Present in Clathrin-coated Vesicles from Bovine Kidney but Not from Brain
| Title: | Functional Water Channels Are Present in Clathrin-coated Vesicles from Bovine Kidney but Not from Brain |
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| Authors: | Ausiello, M.D., Dennis A.; Verkman, Alan S.; Weyer, P.; Brown, Dennis |
| Publisher: | Journal of Biological Chemistry |
| Date Published: | December 05, 1989 |
| Reference Number: | 213 |
Targeting of water channels in renal epithelia may involve trafficking of clathrin-coated vesicles. We have isolated and measured the osmotic water permeability (Pf) of purified clathrin-coated vesicles from bovine kidney cortex and inner medulla, and bovine brain, a tissue not expected to contain "water channels." Brain-coated vesicles had a diameter of 80 nm in negatively stained preparations. Pf was measured by a stopped-flow light scattering technique. In brain-coated vesicles, water transport was functionally homogeneous with a low Pf of 0.0016 +/- 0.0001 cm/s (seven preparations, 23 degrees C). Pf was independent of osmotic gradient size (25-300 mOsm), not inhibited by mercurials, and not altered by removal of the clathrin coat. The activation energy (Ea) for Pf was high (11 +/- 1 kcal/mol less than 34 degrees C, 17 +/- 2 kcal/mol greater than 34 degrees C). Therefore, water channels are absent from brain-coated vesicles. In contrast, there were two functional populations of vesicles in coated vesicle preparations from both kidney cortex and medulla. One population of vesicles had low water permeability and no water channels, whereas a second population had high Pf (0.02 cm/s, 21 degrees C) that was inhibited by HgCl2, and low Ea (2-3 kcal/mol). The fraction of vesicles with high Pf was 52 +/- 3% (S.D., n = 3, cortical vesicles) and 26 +/- 3% (medullary vesicles). These results provide evidence that functional water channels are not present in clathrin-coated vesicles from the brain, whereas they are found in a population of coated vesicles from kidney cortex and medulla, tissues in which water channels are recycled between the plasma membrane, and an intracellular compartment.
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This translation by the NDI Foundation is to assist the lay reader. To provide a clear, accessible interpretation of the original article, we eliminated or simplified some technical detail and complicated scientific language. We concentrated our translation on those aspects of the article dealing directly with NDI. The NDI Foundation thanks the researchers for their work toward understanding and more effectively treating this disorder.
© Copyright NDI Foundation 2007 (JC)
There is a pathway the AQP2 takes to perform its task, and a vehicle to take it along its pathway. The AQP2 sits within a little sac called a vesicle. The AQP2-bearing vesicle waits within the cell in its holding site beneath the apex of the cell membrane (the apical membrane) till signaled by the antidiuretic hormone, vasopressin (VP). Upon receiving the signal, the vesicle travels to and fuses with the apical membrane. When the VP signal ends, the vesicle, AQP2 inside, returns to its holding site.
A protein called clathrin plays an important role in getting the AQP2-bearing vesicle back from the apical membrane to its holding site within the cell. Clathrin coats the surface of pits in the membrane, the AQP2-bearing vesicles moves into the pits then get a coating of clathrin. The cell membrane then infolds the clathrin-coated vesicle and brings it within the cell.
Researchers did not know if clathrin-coated vesicles remain permeable. Nor did they know if the AQP2s within the cathrin-coated vesicles remained functional. Verkman, et al, designed a study to help answer these questions. They investigated the degree of osmotic water permeability of clathrin-coated vesicles from the bovine brain and kidney collecting duct. The brain tissue was not expected to have water channels and therefore was not expected to have a high level of water permeability. The kidney collecting duct cells contain high levels of the water channel, AQP2, and therefore were expected to be more water permeable than the brain tissue.
Water channels generally possess the following characteristics:
- They have a high degree of water permeability (Pf). That is, they let a relatively large volume of water pass through the membranes.
- They have a low activation energy (Ea). That means it does not take much energy for the water channels to transport water across cell membranes.
- Their function is inhibited by mercurial compounds. That is, they cannot perform their task in the presence of mercurial compounds.
- They have a high osmotic to diffusional permeability ratio. This means more water crosses membranes through the water channels than by means of diffusing through the membranes.
Water permeability in clathrin-coated vesicles from bovine brain was low. The Ea was high and water transport was not inhibited by a mercurial compound (HgCl2). Thus, the researchers concluded there were no water channels in the brain tissue. The authors found no change in Ea or Pf when they removed the coating of clathrin from the brain vesicles.
In the kidney collecting tissue, specifically, the outermost part of the kidney (the kidney cortex), the researchers found two distinct functional classes of vesicles. One population of vesicles had low water permeability and no water channels. The other population of vesicles had water channels, as indicated by high water permeability, low Ea, and the fact that HgCl2 inhibited their performance.
In conclusion, Verkman, et al., determined that functional water channels are present in clathrin-coated vesicles from bovine kidney but not from bovine brain. This finding strengthens the notion that water channels are only present in tissues which require high levels of rapid water transport across cell membranes.