- Email: [email protected]
Regulation of aquaporin-2 water channel trafficking by vasopressin
560
Regulation of aquaporin-2 vasopressin Mark A Kneppe? Vasopressin
regulates
collecting
water excretion from the kidney
duct. The aquaporin-2
of the renal
water channel has been
to be the target for this action of vasopressin.
Recent studies have demonstrated through
by
and Takeaki lnoue
by increasing the osmotic water permeability demonstrated
water channel trafficking
that vasopressin,
acting
with two asparagine-proline-alanine (NPA) sequences. These NPA sequences have been proposed to form tight turns which interact in the membrane to form the molecular pore for translocation of water across the plasma membrane [S].
cyclic AMP, triggers fusion of aquaporin-2-bearing
vesicles with the apical plasma membrane duct principal cells. The vesicle-targeting synaptobrevin-2
and syntaxin-
of the collecting proteins
are proposed
Figure 1
to play roles
in this process.
E C
Addresses Laboratory of Kidney and Electrolyte Metabolism,
A National Heart,
Lung and Blood Institute, Building 9, Room IN1 05, 9 Memorial Drive, MSC-0951, National Institutes of Health, Bethesda, MD 20692-0951,
Membrane
USA *e-mail: [email protected] Correspondence:
Mark A Knepper
Current
in Cell Biology
Opinion
G-2
Extracellular
Intracellular 1997, 9560-564
http://biomednet.comlelecref/0955067400900560 0 Current Biology Ltd ISSN
0955-0674
C-J
0
Ser256
dD 1997 Current Opinion in Cell Biology
Abbreviations NPA NSF PKA SNARE VAMP
asparagine-proline-alanine N-ethylmaleimide-sensitive protein kinase A
factor
soluble NSF attachment protein receptor vesicle-associated membrane protein
Introduction There are few physiological processes that are as prominent in everyday life as the regulation of water excretion by the kidney. The chief regulator of renal water excretion is vasopressin, a nine amino acid peptide secreted by the posterior pituitary in response to increased plasma osmolality. Vasopressin (the ‘antidiuretic hormone’) acts at various sites along the renal tubule to control solute and water transport in a manner that allows water excretion to be varied without major changes in solute excretion [l]. Among the regulatory actions of vasopressin in the kidney, the most familiar and best understood is the effect of vasopressin to increase the water permeability of the collecting duct principal cells, thus increasing the return of water from the tubule lumen to the blood and decreasing the osmolality of the systemic plasma. Our understanding of this process has been enhanced by the identification and cloning of the genes encoding a family of molecular water channels called ‘aquaporins’ [Zlr]. The aquaporins are integral membrane glycoproteins with six membrane-spanning regions (Figure 1). An individual aquaporin molecule consists of a tandem repeat structure
Aquaporin-2 topology. Aquaporin-2 spans the plasma membrane six times with three extracellular loops (A, C, and E) and two intracellular loops (B and D). Loop C contains Asnl24, the only putative N-glycosylation site. (A!-glycosylation is shown by a branched structure at Asnl24.) Loops B and E contain NPA sequences which are believed to be elements of the water-conducting pore. The carboxy-terminal tail contains the only putative site of phosphorylation by PKA, that is, Ser256. The membrane-spanning domains are shown by thick black vertical lines. N, amino terminus; C, carboxyl terminus.
The chief target for the action of vasopressin in the regulation of water transport is aquaporin-2, whose cDNA was initially cloned by Fushimi et al. [6]. Patients with inactivating mutations in both alleles of the aquaporin-2 gene manifest severe nephrogenic diabetes insipidus, indicating that this water channel is essential for normal water conservation by the kidney [7]. Immunolocalization studies at light and electron microscopic levels have revealed that aquaporin-2 is abundant in the apical plasma membrane of collecting duct principal cells [g], the rate-limiting barrier for transepithelial water transport in the collecting duct and the chief site of vasopressin action. The remainder of this review focuses on this question: how does vasopressin trigger the activation of aquaporin-2 in the collecting duct principal cells?
Vasopressin-mediated
signalling
The vasopressin receptor responsible for the regulation of water permeability in the collecting duct is the so-called Vz receptor. This receptor is an integral membrane protein
Regulation of aquaporin-2 water channel Knepper and lnoue
of -370 amino acids in length (molecular weight of core protein, 40 kDa) with seven hydrophobic domains, a structure that is common to all G-protein-coupled receptors [9,10]. The V2 receptor gene has been localized to the human X chromosome [9,1 I]. Several mutations of this gene have been identified in patients with X-linked nephrogenic diabetes insipidus [ 12’1, thus establishing the essential role of this receptor in the regulation of renal water excretion. Extensive studies have established that the V2 receptor is coupled to adenylyl cyclase through the heterotrimeric increases G protein G,, and that receptor occupation intracellular cyclic AMP levels in collecting duct cells through activation of adenylyl cyclase [13]. Most cellular effects of cyclic AMP are believed to be secondary to the activation of protein kinase A (PKA) [14] and PKAmediated phosphorylation therefore presumably plays a key role in the regulation of aquaporin-2 activation by vasopressin. Consistent with this view, selective inhibitors of PKA, namely H-S [15] and H-89 [16], blocked the ability of vasopressin or cyclic AMP to activate vasopressin-regulated water permeability in renal collecting ducts. A major focus of current studies of the action of vasopressin in renal epithelia is a determination of which proteins are phosphorylated by PKA and what role these proteins play in regulating renal epithelial water permeability. For example, Dousa and colleagues [17] demonstrated in saponin-permeabilized rat collecting ducts that vasopressin alters the phosphorylation of at least two proteins (of 45 and 66 kDa), both of which remain unidentified. More recently, it has been demonstrated that the vasopressin-regulated water channel aquaporin-2 is phosphorylaced in response to cyclic AMP at position Ser256 [l&19] (Figure 1). The significance of this posttranslational modification with regard to the regulation of water permeability in the collecting duct is discussed below.
Long-term and short-term regulation of osmotic water transport by vasopressin Long-term elevations in circulating vasopressin levels increase maximal osmotic water permeability by increasing the expression level of the aquaporin-2 water channel [ZO]. This action, which requires many hours to be fully manifested, is believed to be, in part, a result of transcriptional regulation mediated by phosphorylation of CAMP response element (CRE)-binding protein (CREB) and binding of phospho-CREB to a CRE in the 5’ flanking region of the aquaporin-2 gene [Zl]. However, the chief cellular process responsible for wide-ranging regulation of water transport in the collecting duct is the short-term effect of vasopressin that is initiated within one minute after exposure of the epithelium to vasopressin and is complete within 30-40 minutes [Z&23]. The short-term effect of vasopressin to increase collecting duct osmotic water permeability is mimicked by application of cyclic AhiP analogs, pointing to a direct role of cyclic AhIP in the
561
regulatory process [24]. However, increases in collecting duct water permeability achieved with cyclic AMP analogs are rarely as great as those obtainable with vasopressin itself, raising the possibility that cyclic AMP may not be the sole mediator of the water-permeability response.
Vasopressin rapidly increases collecting duct water permeability through effects on the exocytosis and endocytosis of aquaporin-2 Immunoelectron microscopic localization of aquaporin-2 in collecting duct principal cells using the immunogold technique has demonstrated that aquaporin-2 is abundant both in the apical plasma membrane and in small intracellular vesicles [8,20]. This finding raised the possibility that the vasopressin-induced increase in water permeability of the collecting duct epithelium could result from exocytosis of the water-channel-containing vesicles, as was originally hypothesized several years ago by Wade and colleagues [ZS]. To test this hypothesis in isolated perfused collecting ducts, we measured osmotic water permeability in the presence and absence of vasopressin and we then immediately fixed the perfused collecting ducts to allow immunogold localization of aquaporin-2 [26]. These studies revealed that vasopressin induced a strong redistribution of aquaporin-2 from intracellular vesicles to the apical plasma membrane concomitantly with a marked increase in the epithelial water permeability. When the vasopressin was withdrawn, aquaporin-2 labeling of the plasma membrane was decreased, with of aquaporin-2 labeling in intracellular reappearance vesicles. This decrease was paralleled by a decrease in water permeability. Thus, these studies provide strong support for the view that the vasopressin-induced immediate increase in collecting duct water permeability is a consequence of vasopressin-induced fusion of intracellular vesicles containing aquaporin-2 with the apical plasma membrane. A series of in viva studies in rats has further strengthened this conclusion by showing that systemic injection of vasopressin is associated with a redistribution of aquaporin-2 from the intracellular compartment to the plasma membrane of collecting duct cells [27-291. Thus, the regulation of water permeability in collecting ducts is, in part, a result of regulated exocytosis of aquaporin-Z-bearing vesicles. In addition to regulating the exocytosis of water channels, vasopressin is also believed to have important effects on the rate of endocytosis of water channels in collecting duct principal cells. Experiments by Brown et al. [30] demonstrated that principal cells take up horseradish peroxidase (HRP) from the collecting duct lumen at a decreased rate in vasopressin-deficient rats and that administration of vasopressin normalized the uptake. The increased rate of endocytosis during vasopressin exposure may be a mass action effect associated with the increased apical plasma membrane surface area that results from vasopressin-stimulated exocytosis. In addition, Strange et a/. [31] demonstrated in isolated
562
Membrane
permeability
perfused rabbit cortical collecting ducts that the removal of vasopressin causes a marked transient acceleration of endocytic uptake of HRP from the lumen, presumably associated with the accelerated retrieval of apical water channels. Subsequently, Nielsen et a/. [32] demonstrated that vasopressin withdrawal is associated with increased luminal uptake of cationic ferritin and albumin gold into small apical vesicles and multivesicular bodies in isolated perfused inner medullary collecting ducts from rats. This effect occurred very rapidly [32], and corresponded with the rapid initial phase of decreased permeability to water in the first 3-5 minutes after washing out of vasopressin [33]. Kinetic analysis of changes in permeability to water in isolated perfused rat inner medullary collecting ducts following vasopressin addition and withdrawal supported the view that vasopressin has two effects: it increases the rate of water-channel exocytosis and it decreases the rate of water-channel endocytosis [33,34]. The manner in which a vasopressin-induced rise in intracellular cyclic AMP levels can alter the rates of trafficking of aquaporin-2 to and from the apical plasma membrane is a major focus of current research. As noted above, aquaporin-2 itself is a target for PKA-mediated phosphorylation at a serine residue at position 256 [l&19]. Direct phosphorylation and dephosphorylation of aquaporin-2 in v&-o did not measurably alter the water permeability of isolated aquaporin-Z-bearing vesicles, suggesting the absence of a simple phosphorylation-dependent gating mechanism [19]. It seems possible, instead, that phosphorylation of Ser256 could play a role in the regulation of the rate of exocytosis or endocytosis of aquaporin-2. Consistent with this view, expression of a mutated form of aquaporin-2 in LLC-PKl cells, with an alanine replacing Ser256, eliminated the vasopressin-induced trafficking of the water channel to the plasma membrane that is normally seen with wild-type aquaporin-2 [35*].
Role of vesicle-targeting aquaporin-2 trafficking
receptors in
Important questions remain regarding the mechanism by which the aquaporin-2-bearing vesicles are specifically targeted to the apical plasma membrane and the mechanism by which vasopressin regulates the docking and fusion of aquaporin-2-bearing vesicles. A new concept of how the targeting of water-channel vesicles might take place is based on studies of regulated exocytosis of synaptic vesicles [36]. These findings suggest that vesicle-targeting receptors (or SNARES, for soluble NSF attachment protein receptors) may mediate a specific interaction between a given vesicle and its target membrane. One class of targeting receptors in the vesicles has been called vesicle-associated membrane proteins (VAMPS) or, alternatively, synaptobrevins. Another family of vesicle-targeting receptors that reside in the vesicles is the synaptotagmins. Two families of SNARE proteins that are present in the target membrane are the syntaxins and synaptosome-associated protein of 25 kDa (SNAP-25) and
its homologs. Recent studies have established that several SNARES are expressed in the principal cells of the renal collecting duct (Figure 2).
Fiaure
2
Synaptotagmin6P I 0 1997 Current Opinion in Cell Biology Several vesicle-targeting proteins (SNARES) are expressed in collecting duct principal cells and are postulated to play a role in the targeting of aquaporin-2-bearing vesicles to the apical plasma membrane. Vesicle-associated SNARES include VAMP-2 and possibly synaptotagmin-6; SNAP-23.
target-membrane
SNARES
include syntaxin-
and
Syntaxins are integral membrane proteins of 3040 kDa that are broadly distributed among mammalian tissues [37]. Recent studies using reverse transcriptase (RT)-PCR and peptide-directed polyclonal antibodies have established that one of these syntaxins, namely, syntaxin4, is expressed in principal cells of the mammalian collecting duct [38*]. Immunolocalization studies in the kidney showed that syntaxinis predominantly present in the apical plasma membrane, the target membrane for vasopressin-regulated aquaporin-2 trafficking [38*]. The VAMPS (or synaptobrevins) are vesicle-associated targeting receptors that are postulated to interact with syntaxins in the target membrane to determine docking specificity. In mammals, there are presently three known VAMP isoforms [39], namely VAMP-l, VAMP-2, and cellubrevin. Two of these isoforms, VAMP-2 and cellubrevin, have been localized to the collecting duct principal cells [40-43]. Immunoelectron microscopic studies using a dual labeling approach have demonstrated the colocalization of aquaporin-2 and VAMP-2 in the same intracellular vesicles in collecting duct principal cells [40]. Thus, studies up to now have demonstrated the presence of two putative SNARES in collecting duct principal cells, as summarized in Figure 2. VAMP-2 is present in intracellular vesicles that contain aquaporin-2, and syntaxinis present in the apical plasma membrane of principal cells. In v&o binary binding assays have demonstrated that VAMP-2 avidly binds syntaxin[44,45]. Thus, it seems possible that VAMP-2 and syntaxinplay vital roles in the targeting of aquaporin-2-bearing vesicles to the apical plasma membrane. However, a functional role for these targeting proteins remains to be demonstrated in the
Regulation
renal collecting duct. Furthermore, preliminary evidence exists indicating that members of the other two classes of SNARE proteins, namely synaptotagmins and SNAPZ-like proteins, are also expressed in the renal collecting duct. Expression of synaptotagmind has been demonstrated in collecting duct cells by RT-PCR and immunocytochemistry, although association with aquaporin-Z-bearing vesicles has not been demonstrated [46]. In addition, we have recently prepared polyclonal antibodies to a recently characterized SNAP-25 homolog called SNAP-23 (MA Knepper, T Inoue, unpublished data). Preliminary immunolocalization studies have demonstrated significant labeling of the apical region of collecting duct cells with this antibody. If the SNARES are involved in the vasopressin-induced trafficking of aquaporin-2 to the apical plasma membrane, then one of these SNARES, or an ancillary protein that binds to them, might be modified, via PKA-induced phosphorylation, at a site that is critical for protein-protein binding. Thus far, there is no evidence for such a modification in the collecting duct.
References
and recommended
. ..
of special interest of outstanding interest
1.
Knepper M, Burg M: Organization Physiol 1963, 244:F579-F569.
2.
Knepper MA: The aquaporin family of molecular water channels. Proc Nat/ Acad Sci USA 1994, 91:6255-6256.
3.
Agre P, Sasaki S, Chrispeels MJ: Aquaporins: a family of water channel proteins. Am J Physioll993, 265:F461.
4.
Nielsen S, Agre P: The aquaporin family of water channels kidney. Kidney lnt 1995, 48:1057-l 066.
5.
Jung JS, Preston GM, Smith BL, Guggino WB, Agre P: Molecular structure of the water channel through aquaporin CHIP. The hour glass model. J Biol Chem 1994, 269:14646-l 4654.
Knepper and lnoue
563
z
Deen PM, Verdijk MA, van OS CH. van Oost channel e&aporin-2 of the urine. Science
6.
Nielsen S, DiGiovanni SR, Christensen El, Knepper MA, Harris HW: Cellular and subcellular immunolocalizetion of vasopressin-regulated water channel in rat kidney. Proc Nat/ Acad Sci USA 1993, 90:11663-l 1667.
9.
Lolait SJ, O’Carroll A-M, McBride OW, Konig M, Morel A, Brownstein MJ: Cloning and characterization of a vasooressin V2 receptor and possible link to nephrogenic diabete; insipidus. Nature 1992, 357:336-339.
10.
Birnbaumer M, Seibold A, Gilbert S, lshido M, Barbaris C, Antaramian A, Brabet P, Rosenthal W: Molecular cloning of the receptor for human antidiuretic hormone. Nature 1992, 357:333-335.
11.
Seibold A, Brabet F’, Rosenthal W, Birnbaumer M: Structure and chromosomal localization of the human antidiuretic hormone receptor gene. Am J Hum Genet 1992, 51 :1076-l 063.
Knoers NV, Wieringa B, Monnens LA, BA: Reauirement of human renal water for vasbpressin-dependent concentration 1994, 264:92-95.
12. Bichet DG: Vasopressin receptors in health and disease. . Kidney lnt 1996, 49:1706-l 711. . .. A thorough revnew ot mutations associated with X-llnkecl diabetes Inslplcius. 13.
Knepper MA, Nielsen S, Chou C-L, DiGiovanni SR: Mechanism of vasopressin action in the renal collecting duct. Semin Nephrol 1994, 141302-321.
14.
Miyamoto E, Kuo JF, Greengard P: Cyclic nucleotide-dependent protein kinases. Ill. Purification and properties of adenosine 3’,5’-monophosphate-dependent protein kinase from bovine brain. J Biol Chem 1969, 244:6395-6402.
15.
Zhang R, Verkman AS: Urea transport in freshly isolated and cultured cells from rat inner medullary collecting duct J Membr Bioll990, 117:253-261.
16.
Snyder HM, Noland TD, Breyer MD: CAMP-dependent protein kinase mediates hydroosmotic effect of vasopressin in collecting duct Am J Physioll992, 263:C147-C153.
1 7.
Homma S, Gapstur SM, Yusufi ANK, Dousa TP: In situ phosphorylation of proteins in MCTs microdissected from rat kidney: effect of AVP. Am J Physiol 1966, 254:F512F520.
16.
Kuwahara M, Fushimi K, Terada Y, Bai L, Marumo F, Sasaki S: CAMP-dependent phosphorylation stimulates water permeability of aquaporin-collecting duct water channel protein expressed in Xenopus oocytes. J Biol Chem 1995, 270:10364-l 0307.
19.
Lande MB, Jo I, Zeidel ML, Somers M, Harris HW Jr: Phosphorylation of aquaporin-2 does not alter the membrane water permeability of rat papillary water channel-containing vesicles. J Biol Chem 1996, 271:5552-5557.
20.
DiGiovanni SR, Nielsen S, Christensen El, Knepper MA: Regulation of collecting duct water channel expression by vasopressin in Brattleboro rat Proc Nat/ Acad Sci USA 1994, 91:6964-6966.
21.
Matsumura Y, Uchida S, Rai T, Sasaki S, Marumo F: Transcriptional regulation of aquaporin-2 water channel gene by CAMP. J Am Sot Nephrol 1997, in press.
22.
Wall SM, Han JS, Chou C-L, Knepper MA: Kinetics of urea and water permeability activation by vasopressin in rat terminal IMCD. Am J Physioll992, 262:F969-F996.
23.
Kuwahara M, Verkman AS: Pre-steady-state analysis of the turnon and turn-off of water permeability in the kidney collecting tubule. J Membr Biol 1969, 11057-65.
24.
Grantham JJ, Burg MB: Effect of vasopressin and cyclic AMP on permeability of isolated collecting tubules. Am J Physiol 1966, 211:255-259.
25.
Wade JB. Stetson DL. Lewis SA: ADH action: evidence for a membrane shuffle ‘mechanism. Ann N Y Acad Sci 1961, 372:106-l 17.
26.
Nielsen S, Chou C-L, Marples D, Christensen El, Kishore BK, Knepper MA: Vasopressin increases water permeability of kidney collecting duct by inducing translocation of aquaporin-CD water channels to plasma membrane. Pioc Nat/ Acad Sci USA 1995, 92:1013-l 017.
of nephron function. Am J
in
channel
Fushimi K. Uchida S. Hara Y. Hirata Y. Marumo F. Sasaki S: Cloning and expreision of apical membrane water channel of rat kidney collecting tubule. Nature 1993, 361:549-552.
reading
Papers of particular interest, published within the annual period of review, have been highlighted as:
water
6.
Conclusions The availability of antibodies to the aquaporin-2 water channel has led to studies demonstrating that vasopressin increases the water permeability of collecting duct cells by triggering the fusion of aquaporin-2-bearing vesicles with the apical plasma membrane of principal cells. Additional evidence indicates that vasopressin also regulates the endocytosis of water channels. Although these responses are presumably a result of PKA-mediated phosphorylation of various proteins, the critical proteins in these regulatory processes are largely unidentified. However, aquaporin-2 itself appears to be a target for PKA-mediated phosphorylation and this process could play a role in regulating aquaporin-2 trafficking in the cell. Several vesicle-targeting proteins (syntaxin-4, VAMP-2, synaptotagmin-6 and SNAP-23) have been immunolocalized to collecting duct principal cells and have been proposed to play a role in targeting aquaporin-2-bearing vesicles to the apical plasma membrane.
of aquaporin-2
564
Membrane
permeability
27.
Marpies D, Knepper MA, Christensen El, Nielsen S: Redistribution of aquaporin-2 water channels inducad by vaooprerrin in rat kidney inner medullary collecting duct Am J Physiol 1995, 269:C655-C664.
29.
Sabolic I, Katsura T, Verbabatz JM, Brown D: The AQPP water channel: effect of vasopressin treatment microtubule disruption, end distribution in neonatal rats. J Membr Bioll995, 143:165-l 77.
29.
Yamamoto N, Sasaki S, Fushimi K, lshibashi K, Yaiota E, Kawasaki K, Marumo F, Kihara I: Vasopressin increases AQP-CD water channel in the apical membrane of collecting duct cells without affecting AQP3 distribution in Brattleboro rat. Am J Physioll995, 266:C1546-Cl551.
37
Bennett MK, Garcia-Arraras JE, Elferink LA, Peterson K, Fleming AM, Haxuka CD, Scheller RH: The syntaxin family of vesicular transport receptors. Cell 1993, 74:863-873.
38. .
Mandon B, Chou C-L, Nielsen S, Knepper MA: Syntaxinis localized to the apical plasma membrane of rat renal collecting duct cells: possible role in aquaporin-2 trafficking. J C/in invest 1996, 98:906-913. This paper is the first to demonstrate expression of a syntaxin protein in an epithelium manifesting hormonally regulated exocytosis. 39.
Sijdhof TC, De Camilli P, Niemann H, Jahn R: Membrane fusion machinery: insights from synaptic proteins. Cell 1993, 75:1-4.
40.
Brown D, Weyer P, Orci L: Vasopressin stimulates endocytosis in kidney collecting duct principal cells. Eur J Cell Biol 1988, 46:336-341.
Nielsen S, Marples D, Mohtashami M, Dalby NO, Thimble W, Knepper M: Expression of VAMP2-like protein in kidney collecting duct intracellular vesicles: co-localization with aquaporin-2 water channels. J C/in Invest 1995, 96:1634-l
41.
31.
Strange K, Willingham MC, Handler JS, Harris HW Jr: Apical membrane endocytosis via coated pits is stimulated by removal of antidiuretic hormone from isolated, perfused rabbit cortical collecting tubule. J Membr Biol 1988, 103:17-26.
Liebenhofl U, Rosenthal W: Identification of RabS-. Rab5a and synaptobrevin II-like proteins in a preparation of rat kidney vesicles containing the vasopressin-regulated water channel. FEBS Lett 1995, 366:209-213.
42.
32.
Nielsen S, Muller J, Knepper MA: Vasopressin- and cAMPinduced changes in ultrastructure of isolated perfused inner medullary collecting ducts. Am J Physiol 1993, 265:F225-F239.
33.
Nielsen S, Knepper MA: Vasopressin activates collecting duct urea transporters and water channels by distinct physical processes. Am J Physioll993,265:F204-F213.
Jo I, Harris HW, Amendt-Raduege AM, Majewski RR, Hammond TG: Rat kidney papilla contains abundant svnaotobrevin orotein that oarticioates in the fusion of antidiuretic hormone-regulated water channel-containing endosomes in vitro. Proc Nat/ Acad Sci USA 1995, 92:1876-l 880.
43.
Knepper MA, Nielsen S: Kinetic model of water end urea permeability regulation by vasopressin in collecting duct Am J Physioll993,266:F214-F224.
Franki N, Macaluso F, Schubert W, Gunther L, Hays RM: Water channel-carrying vesicles in the rat IMCD contain cellubrevin. Am J Physioll995,269:C797-C801.
44.
Calakos N. Bennett MK. Peterson KE. Scheller RH: Proteinprotein interactions contributing to ihe specificity of intracellular vesicular trafficking. Science 1994, 263:1146-l 149.
45.
Pevsner J, Hsu S-C, Braun JEA, Calakos N, Ting AE, Bennett MK, Scheller RH: Specificity and regulation of a synaptic vesicle docking complex. Neuron 1994, 13:353-361.
46.
Kishore BK, Mandon B, Nielsen S, Knepper MA: Synaptotagmin6 is expressed in collecting duct principal cell: a possible molecular switch in vasopressin-induced water channel trafficking [abstractl. J Am Sot Nephrol 1996. 7:1310.
30.
34.
Katsura T, Gustafson CE. Ausiello DA, Brown D: Protein kinase A phosphorylation is involved in regulated exocytosis of aquaporin-2 in transfected LLC-PKI cells. Am J Physiol 1997, In press. Provides strong evidence in support of the view that regulated trafficking of aquaporin-2 is dependent on PKA-mediated phosphotylation of Ser256 in aquaporin-2. 35. .
36.
Bajjalieh SM, Scheller RH: The biochemistry of neurotransmitter secretion. J Biol Chem 1995, 270:1971-l 974.
844.
Regulation of aquaporin-2 vasopressin Mark A Kneppe? Vasopressin
regulates
collecting
water excretion from the kidney
duct. The aquaporin-2
of the renal
water channel has been
to be the target for this action of vasopressin.
Recent studies have demonstrated through
by
and Takeaki lnoue
by increasing the osmotic water permeability demonstrated
water channel trafficking
that vasopressin,
acting
with two asparagine-proline-alanine (NPA) sequences. These NPA sequences have been proposed to form tight turns which interact in the membrane to form the molecular pore for translocation of water across the plasma membrane [S].
cyclic AMP, triggers fusion of aquaporin-2-bearing
vesicles with the apical plasma membrane duct principal cells. The vesicle-targeting synaptobrevin-2
and syntaxin-
of the collecting proteins
are proposed
Figure 1
to play roles
in this process.
E C
Addresses Laboratory of Kidney and Electrolyte Metabolism,
A National Heart,
Lung and Blood Institute, Building 9, Room IN1 05, 9 Memorial Drive, MSC-0951, National Institutes of Health, Bethesda, MD 20692-0951,
Membrane
USA *e-mail: [email protected] Correspondence:
Mark A Knepper
Current
in Cell Biology
Opinion
G-2
Extracellular
Intracellular 1997, 9560-564
http://biomednet.comlelecref/0955067400900560 0 Current Biology Ltd ISSN
0955-0674
C-J
0
Ser256
dD 1997 Current Opinion in Cell Biology
Abbreviations NPA NSF PKA SNARE VAMP
asparagine-proline-alanine N-ethylmaleimide-sensitive protein kinase A
factor
soluble NSF attachment protein receptor vesicle-associated membrane protein
Introduction There are few physiological processes that are as prominent in everyday life as the regulation of water excretion by the kidney. The chief regulator of renal water excretion is vasopressin, a nine amino acid peptide secreted by the posterior pituitary in response to increased plasma osmolality. Vasopressin (the ‘antidiuretic hormone’) acts at various sites along the renal tubule to control solute and water transport in a manner that allows water excretion to be varied without major changes in solute excretion [l]. Among the regulatory actions of vasopressin in the kidney, the most familiar and best understood is the effect of vasopressin to increase the water permeability of the collecting duct principal cells, thus increasing the return of water from the tubule lumen to the blood and decreasing the osmolality of the systemic plasma. Our understanding of this process has been enhanced by the identification and cloning of the genes encoding a family of molecular water channels called ‘aquaporins’ [Zlr]. The aquaporins are integral membrane glycoproteins with six membrane-spanning regions (Figure 1). An individual aquaporin molecule consists of a tandem repeat structure
Aquaporin-2 topology. Aquaporin-2 spans the plasma membrane six times with three extracellular loops (A, C, and E) and two intracellular loops (B and D). Loop C contains Asnl24, the only putative N-glycosylation site. (A!-glycosylation is shown by a branched structure at Asnl24.) Loops B and E contain NPA sequences which are believed to be elements of the water-conducting pore. The carboxy-terminal tail contains the only putative site of phosphorylation by PKA, that is, Ser256. The membrane-spanning domains are shown by thick black vertical lines. N, amino terminus; C, carboxyl terminus.
The chief target for the action of vasopressin in the regulation of water transport is aquaporin-2, whose cDNA was initially cloned by Fushimi et al. [6]. Patients with inactivating mutations in both alleles of the aquaporin-2 gene manifest severe nephrogenic diabetes insipidus, indicating that this water channel is essential for normal water conservation by the kidney [7]. Immunolocalization studies at light and electron microscopic levels have revealed that aquaporin-2 is abundant in the apical plasma membrane of collecting duct principal cells [g], the rate-limiting barrier for transepithelial water transport in the collecting duct and the chief site of vasopressin action. The remainder of this review focuses on this question: how does vasopressin trigger the activation of aquaporin-2 in the collecting duct principal cells?
Vasopressin-mediated
signalling
The vasopressin receptor responsible for the regulation of water permeability in the collecting duct is the so-called Vz receptor. This receptor is an integral membrane protein
Regulation of aquaporin-2 water channel Knepper and lnoue
of -370 amino acids in length (molecular weight of core protein, 40 kDa) with seven hydrophobic domains, a structure that is common to all G-protein-coupled receptors [9,10]. The V2 receptor gene has been localized to the human X chromosome [9,1 I]. Several mutations of this gene have been identified in patients with X-linked nephrogenic diabetes insipidus [ 12’1, thus establishing the essential role of this receptor in the regulation of renal water excretion. Extensive studies have established that the V2 receptor is coupled to adenylyl cyclase through the heterotrimeric increases G protein G,, and that receptor occupation intracellular cyclic AMP levels in collecting duct cells through activation of adenylyl cyclase [13]. Most cellular effects of cyclic AMP are believed to be secondary to the activation of protein kinase A (PKA) [14] and PKAmediated phosphorylation therefore presumably plays a key role in the regulation of aquaporin-2 activation by vasopressin. Consistent with this view, selective inhibitors of PKA, namely H-S [15] and H-89 [16], blocked the ability of vasopressin or cyclic AMP to activate vasopressin-regulated water permeability in renal collecting ducts. A major focus of current studies of the action of vasopressin in renal epithelia is a determination of which proteins are phosphorylated by PKA and what role these proteins play in regulating renal epithelial water permeability. For example, Dousa and colleagues [17] demonstrated in saponin-permeabilized rat collecting ducts that vasopressin alters the phosphorylation of at least two proteins (of 45 and 66 kDa), both of which remain unidentified. More recently, it has been demonstrated that the vasopressin-regulated water channel aquaporin-2 is phosphorylaced in response to cyclic AMP at position Ser256 [l&19] (Figure 1). The significance of this posttranslational modification with regard to the regulation of water permeability in the collecting duct is discussed below.
Long-term and short-term regulation of osmotic water transport by vasopressin Long-term elevations in circulating vasopressin levels increase maximal osmotic water permeability by increasing the expression level of the aquaporin-2 water channel [ZO]. This action, which requires many hours to be fully manifested, is believed to be, in part, a result of transcriptional regulation mediated by phosphorylation of CAMP response element (CRE)-binding protein (CREB) and binding of phospho-CREB to a CRE in the 5’ flanking region of the aquaporin-2 gene [Zl]. However, the chief cellular process responsible for wide-ranging regulation of water transport in the collecting duct is the short-term effect of vasopressin that is initiated within one minute after exposure of the epithelium to vasopressin and is complete within 30-40 minutes [Z&23]. The short-term effect of vasopressin to increase collecting duct osmotic water permeability is mimicked by application of cyclic AhiP analogs, pointing to a direct role of cyclic AhIP in the
561
regulatory process [24]. However, increases in collecting duct water permeability achieved with cyclic AMP analogs are rarely as great as those obtainable with vasopressin itself, raising the possibility that cyclic AMP may not be the sole mediator of the water-permeability response.
Vasopressin rapidly increases collecting duct water permeability through effects on the exocytosis and endocytosis of aquaporin-2 Immunoelectron microscopic localization of aquaporin-2 in collecting duct principal cells using the immunogold technique has demonstrated that aquaporin-2 is abundant both in the apical plasma membrane and in small intracellular vesicles [8,20]. This finding raised the possibility that the vasopressin-induced increase in water permeability of the collecting duct epithelium could result from exocytosis of the water-channel-containing vesicles, as was originally hypothesized several years ago by Wade and colleagues [ZS]. To test this hypothesis in isolated perfused collecting ducts, we measured osmotic water permeability in the presence and absence of vasopressin and we then immediately fixed the perfused collecting ducts to allow immunogold localization of aquaporin-2 [26]. These studies revealed that vasopressin induced a strong redistribution of aquaporin-2 from intracellular vesicles to the apical plasma membrane concomitantly with a marked increase in the epithelial water permeability. When the vasopressin was withdrawn, aquaporin-2 labeling of the plasma membrane was decreased, with of aquaporin-2 labeling in intracellular reappearance vesicles. This decrease was paralleled by a decrease in water permeability. Thus, these studies provide strong support for the view that the vasopressin-induced immediate increase in collecting duct water permeability is a consequence of vasopressin-induced fusion of intracellular vesicles containing aquaporin-2 with the apical plasma membrane. A series of in viva studies in rats has further strengthened this conclusion by showing that systemic injection of vasopressin is associated with a redistribution of aquaporin-2 from the intracellular compartment to the plasma membrane of collecting duct cells [27-291. Thus, the regulation of water permeability in collecting ducts is, in part, a result of regulated exocytosis of aquaporin-Z-bearing vesicles. In addition to regulating the exocytosis of water channels, vasopressin is also believed to have important effects on the rate of endocytosis of water channels in collecting duct principal cells. Experiments by Brown et al. [30] demonstrated that principal cells take up horseradish peroxidase (HRP) from the collecting duct lumen at a decreased rate in vasopressin-deficient rats and that administration of vasopressin normalized the uptake. The increased rate of endocytosis during vasopressin exposure may be a mass action effect associated with the increased apical plasma membrane surface area that results from vasopressin-stimulated exocytosis. In addition, Strange et a/. [31] demonstrated in isolated
562
Membrane
permeability
perfused rabbit cortical collecting ducts that the removal of vasopressin causes a marked transient acceleration of endocytic uptake of HRP from the lumen, presumably associated with the accelerated retrieval of apical water channels. Subsequently, Nielsen et a/. [32] demonstrated that vasopressin withdrawal is associated with increased luminal uptake of cationic ferritin and albumin gold into small apical vesicles and multivesicular bodies in isolated perfused inner medullary collecting ducts from rats. This effect occurred very rapidly [32], and corresponded with the rapid initial phase of decreased permeability to water in the first 3-5 minutes after washing out of vasopressin [33]. Kinetic analysis of changes in permeability to water in isolated perfused rat inner medullary collecting ducts following vasopressin addition and withdrawal supported the view that vasopressin has two effects: it increases the rate of water-channel exocytosis and it decreases the rate of water-channel endocytosis [33,34]. The manner in which a vasopressin-induced rise in intracellular cyclic AMP levels can alter the rates of trafficking of aquaporin-2 to and from the apical plasma membrane is a major focus of current research. As noted above, aquaporin-2 itself is a target for PKA-mediated phosphorylation at a serine residue at position 256 [l&19]. Direct phosphorylation and dephosphorylation of aquaporin-2 in v&-o did not measurably alter the water permeability of isolated aquaporin-Z-bearing vesicles, suggesting the absence of a simple phosphorylation-dependent gating mechanism [19]. It seems possible, instead, that phosphorylation of Ser256 could play a role in the regulation of the rate of exocytosis or endocytosis of aquaporin-2. Consistent with this view, expression of a mutated form of aquaporin-2 in LLC-PKl cells, with an alanine replacing Ser256, eliminated the vasopressin-induced trafficking of the water channel to the plasma membrane that is normally seen with wild-type aquaporin-2 [35*].
Role of vesicle-targeting aquaporin-2 trafficking
receptors in
Important questions remain regarding the mechanism by which the aquaporin-2-bearing vesicles are specifically targeted to the apical plasma membrane and the mechanism by which vasopressin regulates the docking and fusion of aquaporin-2-bearing vesicles. A new concept of how the targeting of water-channel vesicles might take place is based on studies of regulated exocytosis of synaptic vesicles [36]. These findings suggest that vesicle-targeting receptors (or SNARES, for soluble NSF attachment protein receptors) may mediate a specific interaction between a given vesicle and its target membrane. One class of targeting receptors in the vesicles has been called vesicle-associated membrane proteins (VAMPS) or, alternatively, synaptobrevins. Another family of vesicle-targeting receptors that reside in the vesicles is the synaptotagmins. Two families of SNARE proteins that are present in the target membrane are the syntaxins and synaptosome-associated protein of 25 kDa (SNAP-25) and
its homologs. Recent studies have established that several SNARES are expressed in the principal cells of the renal collecting duct (Figure 2).
Fiaure
2
Synaptotagmin6P I 0 1997 Current Opinion in Cell Biology Several vesicle-targeting proteins (SNARES) are expressed in collecting duct principal cells and are postulated to play a role in the targeting of aquaporin-2-bearing vesicles to the apical plasma membrane. Vesicle-associated SNARES include VAMP-2 and possibly synaptotagmin-6; SNAP-23.
target-membrane
SNARES
include syntaxin-
and
Syntaxins are integral membrane proteins of 3040 kDa that are broadly distributed among mammalian tissues [37]. Recent studies using reverse transcriptase (RT)-PCR and peptide-directed polyclonal antibodies have established that one of these syntaxins, namely, syntaxin4, is expressed in principal cells of the mammalian collecting duct [38*]. Immunolocalization studies in the kidney showed that syntaxinis predominantly present in the apical plasma membrane, the target membrane for vasopressin-regulated aquaporin-2 trafficking [38*]. The VAMPS (or synaptobrevins) are vesicle-associated targeting receptors that are postulated to interact with syntaxins in the target membrane to determine docking specificity. In mammals, there are presently three known VAMP isoforms [39], namely VAMP-l, VAMP-2, and cellubrevin. Two of these isoforms, VAMP-2 and cellubrevin, have been localized to the collecting duct principal cells [40-43]. Immunoelectron microscopic studies using a dual labeling approach have demonstrated the colocalization of aquaporin-2 and VAMP-2 in the same intracellular vesicles in collecting duct principal cells [40]. Thus, studies up to now have demonstrated the presence of two putative SNARES in collecting duct principal cells, as summarized in Figure 2. VAMP-2 is present in intracellular vesicles that contain aquaporin-2, and syntaxinis present in the apical plasma membrane of principal cells. In v&o binary binding assays have demonstrated that VAMP-2 avidly binds syntaxin[44,45]. Thus, it seems possible that VAMP-2 and syntaxinplay vital roles in the targeting of aquaporin-2-bearing vesicles to the apical plasma membrane. However, a functional role for these targeting proteins remains to be demonstrated in the
Regulation
renal collecting duct. Furthermore, preliminary evidence exists indicating that members of the other two classes of SNARE proteins, namely synaptotagmins and SNAPZ-like proteins, are also expressed in the renal collecting duct. Expression of synaptotagmind has been demonstrated in collecting duct cells by RT-PCR and immunocytochemistry, although association with aquaporin-Z-bearing vesicles has not been demonstrated [46]. In addition, we have recently prepared polyclonal antibodies to a recently characterized SNAP-25 homolog called SNAP-23 (MA Knepper, T Inoue, unpublished data). Preliminary immunolocalization studies have demonstrated significant labeling of the apical region of collecting duct cells with this antibody. If the SNARES are involved in the vasopressin-induced trafficking of aquaporin-2 to the apical plasma membrane, then one of these SNARES, or an ancillary protein that binds to them, might be modified, via PKA-induced phosphorylation, at a site that is critical for protein-protein binding. Thus far, there is no evidence for such a modification in the collecting duct.
References
and recommended
. ..
of special interest of outstanding interest
1.
Knepper M, Burg M: Organization Physiol 1963, 244:F579-F569.
2.
Knepper MA: The aquaporin family of molecular water channels. Proc Nat/ Acad Sci USA 1994, 91:6255-6256.
3.
Agre P, Sasaki S, Chrispeels MJ: Aquaporins: a family of water channel proteins. Am J Physioll993, 265:F461.
4.
Nielsen S, Agre P: The aquaporin family of water channels kidney. Kidney lnt 1995, 48:1057-l 066.
5.
Jung JS, Preston GM, Smith BL, Guggino WB, Agre P: Molecular structure of the water channel through aquaporin CHIP. The hour glass model. J Biol Chem 1994, 269:14646-l 4654.
Knepper and lnoue
563
z
Deen PM, Verdijk MA, van OS CH. van Oost channel e&aporin-2 of the urine. Science
6.
Nielsen S, DiGiovanni SR, Christensen El, Knepper MA, Harris HW: Cellular and subcellular immunolocalizetion of vasopressin-regulated water channel in rat kidney. Proc Nat/ Acad Sci USA 1993, 90:11663-l 1667.
9.
Lolait SJ, O’Carroll A-M, McBride OW, Konig M, Morel A, Brownstein MJ: Cloning and characterization of a vasooressin V2 receptor and possible link to nephrogenic diabete; insipidus. Nature 1992, 357:336-339.
10.
Birnbaumer M, Seibold A, Gilbert S, lshido M, Barbaris C, Antaramian A, Brabet P, Rosenthal W: Molecular cloning of the receptor for human antidiuretic hormone. Nature 1992, 357:333-335.
11.
Seibold A, Brabet F’, Rosenthal W, Birnbaumer M: Structure and chromosomal localization of the human antidiuretic hormone receptor gene. Am J Hum Genet 1992, 51 :1076-l 063.
Knoers NV, Wieringa B, Monnens LA, BA: Reauirement of human renal water for vasbpressin-dependent concentration 1994, 264:92-95.
12. Bichet DG: Vasopressin receptors in health and disease. . Kidney lnt 1996, 49:1706-l 711. . .. A thorough revnew ot mutations associated with X-llnkecl diabetes Inslplcius. 13.
Knepper MA, Nielsen S, Chou C-L, DiGiovanni SR: Mechanism of vasopressin action in the renal collecting duct. Semin Nephrol 1994, 141302-321.
14.
Miyamoto E, Kuo JF, Greengard P: Cyclic nucleotide-dependent protein kinases. Ill. Purification and properties of adenosine 3’,5’-monophosphate-dependent protein kinase from bovine brain. J Biol Chem 1969, 244:6395-6402.
15.
Zhang R, Verkman AS: Urea transport in freshly isolated and cultured cells from rat inner medullary collecting duct J Membr Bioll990, 117:253-261.
16.
Snyder HM, Noland TD, Breyer MD: CAMP-dependent protein kinase mediates hydroosmotic effect of vasopressin in collecting duct Am J Physioll992, 263:C147-C153.
1 7.
Homma S, Gapstur SM, Yusufi ANK, Dousa TP: In situ phosphorylation of proteins in MCTs microdissected from rat kidney: effect of AVP. Am J Physiol 1966, 254:F512F520.
16.
Kuwahara M, Fushimi K, Terada Y, Bai L, Marumo F, Sasaki S: CAMP-dependent phosphorylation stimulates water permeability of aquaporin-collecting duct water channel protein expressed in Xenopus oocytes. J Biol Chem 1995, 270:10364-l 0307.
19.
Lande MB, Jo I, Zeidel ML, Somers M, Harris HW Jr: Phosphorylation of aquaporin-2 does not alter the membrane water permeability of rat papillary water channel-containing vesicles. J Biol Chem 1996, 271:5552-5557.
20.
DiGiovanni SR, Nielsen S, Christensen El, Knepper MA: Regulation of collecting duct water channel expression by vasopressin in Brattleboro rat Proc Nat/ Acad Sci USA 1994, 91:6964-6966.
21.
Matsumura Y, Uchida S, Rai T, Sasaki S, Marumo F: Transcriptional regulation of aquaporin-2 water channel gene by CAMP. J Am Sot Nephrol 1997, in press.
22.
Wall SM, Han JS, Chou C-L, Knepper MA: Kinetics of urea and water permeability activation by vasopressin in rat terminal IMCD. Am J Physioll992, 262:F969-F996.
23.
Kuwahara M, Verkman AS: Pre-steady-state analysis of the turnon and turn-off of water permeability in the kidney collecting tubule. J Membr Biol 1969, 11057-65.
24.
Grantham JJ, Burg MB: Effect of vasopressin and cyclic AMP on permeability of isolated collecting tubules. Am J Physiol 1966, 211:255-259.
25.
Wade JB. Stetson DL. Lewis SA: ADH action: evidence for a membrane shuffle ‘mechanism. Ann N Y Acad Sci 1961, 372:106-l 17.
26.
Nielsen S, Chou C-L, Marples D, Christensen El, Kishore BK, Knepper MA: Vasopressin increases water permeability of kidney collecting duct by inducing translocation of aquaporin-CD water channels to plasma membrane. Pioc Nat/ Acad Sci USA 1995, 92:1013-l 017.
of nephron function. Am J
in
channel
Fushimi K. Uchida S. Hara Y. Hirata Y. Marumo F. Sasaki S: Cloning and expreision of apical membrane water channel of rat kidney collecting tubule. Nature 1993, 361:549-552.
reading
Papers of particular interest, published within the annual period of review, have been highlighted as:
water
6.
Conclusions The availability of antibodies to the aquaporin-2 water channel has led to studies demonstrating that vasopressin increases the water permeability of collecting duct cells by triggering the fusion of aquaporin-2-bearing vesicles with the apical plasma membrane of principal cells. Additional evidence indicates that vasopressin also regulates the endocytosis of water channels. Although these responses are presumably a result of PKA-mediated phosphorylation of various proteins, the critical proteins in these regulatory processes are largely unidentified. However, aquaporin-2 itself appears to be a target for PKA-mediated phosphorylation and this process could play a role in regulating aquaporin-2 trafficking in the cell. Several vesicle-targeting proteins (syntaxin-4, VAMP-2, synaptotagmin-6 and SNAP-23) have been immunolocalized to collecting duct principal cells and have been proposed to play a role in targeting aquaporin-2-bearing vesicles to the apical plasma membrane.
of aquaporin-2
564
Membrane
permeability
27.
Marpies D, Knepper MA, Christensen El, Nielsen S: Redistribution of aquaporin-2 water channels inducad by vaooprerrin in rat kidney inner medullary collecting duct Am J Physiol 1995, 269:C655-C664.
29.
Sabolic I, Katsura T, Verbabatz JM, Brown D: The AQPP water channel: effect of vasopressin treatment microtubule disruption, end distribution in neonatal rats. J Membr Bioll995, 143:165-l 77.
29.
Yamamoto N, Sasaki S, Fushimi K, lshibashi K, Yaiota E, Kawasaki K, Marumo F, Kihara I: Vasopressin increases AQP-CD water channel in the apical membrane of collecting duct cells without affecting AQP3 distribution in Brattleboro rat. Am J Physioll995, 266:C1546-Cl551.
37
Bennett MK, Garcia-Arraras JE, Elferink LA, Peterson K, Fleming AM, Haxuka CD, Scheller RH: The syntaxin family of vesicular transport receptors. Cell 1993, 74:863-873.
38. .
Mandon B, Chou C-L, Nielsen S, Knepper MA: Syntaxinis localized to the apical plasma membrane of rat renal collecting duct cells: possible role in aquaporin-2 trafficking. J C/in invest 1996, 98:906-913. This paper is the first to demonstrate expression of a syntaxin protein in an epithelium manifesting hormonally regulated exocytosis. 39.
Sijdhof TC, De Camilli P, Niemann H, Jahn R: Membrane fusion machinery: insights from synaptic proteins. Cell 1993, 75:1-4.
40.
Brown D, Weyer P, Orci L: Vasopressin stimulates endocytosis in kidney collecting duct principal cells. Eur J Cell Biol 1988, 46:336-341.
Nielsen S, Marples D, Mohtashami M, Dalby NO, Thimble W, Knepper M: Expression of VAMP2-like protein in kidney collecting duct intracellular vesicles: co-localization with aquaporin-2 water channels. J C/in Invest 1995, 96:1634-l
41.
31.
Strange K, Willingham MC, Handler JS, Harris HW Jr: Apical membrane endocytosis via coated pits is stimulated by removal of antidiuretic hormone from isolated, perfused rabbit cortical collecting tubule. J Membr Biol 1988, 103:17-26.
Liebenhofl U, Rosenthal W: Identification of RabS-. Rab5a and synaptobrevin II-like proteins in a preparation of rat kidney vesicles containing the vasopressin-regulated water channel. FEBS Lett 1995, 366:209-213.
42.
32.
Nielsen S, Muller J, Knepper MA: Vasopressin- and cAMPinduced changes in ultrastructure of isolated perfused inner medullary collecting ducts. Am J Physiol 1993, 265:F225-F239.
33.
Nielsen S, Knepper MA: Vasopressin activates collecting duct urea transporters and water channels by distinct physical processes. Am J Physioll993,265:F204-F213.
Jo I, Harris HW, Amendt-Raduege AM, Majewski RR, Hammond TG: Rat kidney papilla contains abundant svnaotobrevin orotein that oarticioates in the fusion of antidiuretic hormone-regulated water channel-containing endosomes in vitro. Proc Nat/ Acad Sci USA 1995, 92:1876-l 880.
43.
Knepper MA, Nielsen S: Kinetic model of water end urea permeability regulation by vasopressin in collecting duct Am J Physioll993,266:F214-F224.
Franki N, Macaluso F, Schubert W, Gunther L, Hays RM: Water channel-carrying vesicles in the rat IMCD contain cellubrevin. Am J Physioll995,269:C797-C801.
44.
Calakos N. Bennett MK. Peterson KE. Scheller RH: Proteinprotein interactions contributing to ihe specificity of intracellular vesicular trafficking. Science 1994, 263:1146-l 149.
45.
Pevsner J, Hsu S-C, Braun JEA, Calakos N, Ting AE, Bennett MK, Scheller RH: Specificity and regulation of a synaptic vesicle docking complex. Neuron 1994, 13:353-361.
46.
Kishore BK, Mandon B, Nielsen S, Knepper MA: Synaptotagmin6 is expressed in collecting duct principal cell: a possible molecular switch in vasopressin-induced water channel trafficking [abstractl. J Am Sot Nephrol 1996. 7:1310.
30.
34.
Katsura T, Gustafson CE. Ausiello DA, Brown D: Protein kinase A phosphorylation is involved in regulated exocytosis of aquaporin-2 in transfected LLC-PKI cells. Am J Physiol 1997, In press. Provides strong evidence in support of the view that regulated trafficking of aquaporin-2 is dependent on PKA-mediated phosphotylation of Ser256 in aquaporin-2. 35. .
36.
Bajjalieh SM, Scheller RH: The biochemistry of neurotransmitter secretion. J Biol Chem 1995, 270:1971-l 974.
844.