2002 Global Researcher Conference Proceeding
April 26 - 28, 2002
| Conference: | 2002 Global Researcher Conference |
|---|---|
| Title: | Long- and Short-Term Regulation of AQP2 Expression and Function |
| Authors: | Baggaley, Erin; Shaw, Stephen; Marples, David |
| Institutions: | University of Leeds, Nijmegen Centre for Molecular Life Sciences |
The work in our lab has two major foci: the cellular mechanisms underlying the acute shuttling of AQP2 water channels, and the longer-term signals controlling the expression of AQP2 in the cells. Both of these elements are likely to be important in the aetiology of acquired forms of nephrogenic diabetes insipidus.
It is now well-established that vasopressin (VP) causes a rise in cAMP, and hence activation of protein kinase A, and that AQP2 is phosphorylated at serine 256. This seems to be a critical step in the shuttling of AQP2 to the plasma membrane. However, recent evidence has suggested that a calcium signal is also involved (Chou et al, 2000), and it has been shown that protein kinase C (PKC) also phosphorylates AQP2 (Promeneur et al, 2001), albeit probably at a different site (Serine 231). We have therefore looked at the effect of activation or inhibition of PKC on VP-induced shuttling of AQP2, using our recently-developed tubule preparation. We find that activation of PKC prevents AQP2 shuttling, while inhibition of PKC potentiates the response to VP. These results show that the protein kinase C activity modulates the response to VP. This tubule preparation will provide a powerful tool for future studies.
Our other major interest concerns the signals that control AQP2 synthesis. We have substantial, but indirect, evidence that prostaglandins are one of these signals, providing a stimulus for AQP2 production. Shortly before the last NDIF meeting we were awarded a grant from the NDIF to do preliminary studies to try and determine which prostaglandins were responsible for this effect, using in vivo infusions of PGE2 and PGI2, the two principal candidates. These experiments have provided interesting but inconclusive results, probably because of metabolism of the prostaglandins in the bloodstream, so that renal levels were limited. Both PGE2 and PGI2 appeared to decrease urine output, but this result did not reach statistical significance. There was also no significant change in AQP2 expression, although there was a suggestion that PGI2 might increase, and perhaps PGE2 decrease, AQP2 expression. Rather than increasing PG doses further, which would probably have unacceptable systemic effects, we plan to take this work further using the isolated tubule preparation, once we can keep the tissue viable for several hours.
We have also investigated the effect of chlorpropamide, a drug sometimes used in the treatment of central DI, on AQP2 distribution and expression. The mechanism by chlorpropamide acts remains unclear. A recent study by Durr et al. (2000) had shown an increase in both basal and VP-stimulated water permeability, which could have been a consequence of increased AQP2 expression. However, an earlier study (Zusman et al, 1977) had implicated an inhibition of PG production. We found that chlorpropamide increased AQP2 shuttling, without altering its expression.
References
Chou CL et al. (2000) JBC 275:36839-46
Durr J et al. (2000) Am. J. Physiol. 278:F799-808
Promeneur D. et al. (2001) J. Am. Soc. Nephrol. 12: A0110
Zusman RM et al. (1977) J. Clin. Invest. 60:1348-53
Marples, et al., investigated the effect of protein kinase C (PKC) on the movement of aquaporin-2 (AQP2) from inside the cell to the cell wall. They found that when PKC is activated, it prevents AQP2's movement to the cell wall. When PKC is inhibited, it increases the cell's ability to respond to vasopressin, the hormone which initiates the molecular sequence responsible for allowing water to flow out of the urine and across the cell.
Marples, et al. also investigated PGE2 and PGI2, two prostaglandins that they suspected might affect AQP2 production. Their research to date has been suggestive but not conclusive. The research team also investigated the effect on AQP2 of the drug chlorpropamide, which is used to treat central diabetes insipidus, and found it increased AQP2s movement from cell interior to cell wall.