Heat Shock Protein 70 Interacts with Aquaporin-2 and Regulates Its Trafficking

The trafficking of aquaporin-2 (AQP2) involves multiple complex pathways, including regulated, cAMP-, and cGMP-mediated pathways, as well as a constitutive recycling pathway. Although several accessory proteins have been indirectly implicated in AQP2 recycling, the direct protein-protein interactions that regulate this process remain largely unknown. Using yeast two-hybrid screening of a human kidney cDNA library, we have identified the 70-kDa heat shock proteins as AQP2-interacting proteins. Interaction was confirmed by mass spectrometry of proteins pulled down from rat kidney papilla extract using a GST-AQP2 C-terminal fusion protein (GST-A2C) as a bait, by co-immunoprecipitation (IP) assays, and by direct binding assays using purified hsc70 and the GST-A2C. The direct interaction of AQP2 with hsc70 is partially inhibited by ATP, and the Ser-256 residue in the AQP2 C terminus is important for this direct interaction. Vasopressin stimulation in cells enhances the interaction of hsc70 with AQP2 in IP assays, and vasopressin stimulation in vivo induces an increased co-localization of hsc70 and AQP2 on the apical membrane of principal cells in rat kidney collecting ducts. Functional knockdown of hsc70 activity in AQP2 expressing cells results in membrane accumulation of AQP2 and reduced endocytosis of rhodamine-transferrin. Our data also show that AQP2 interacts with hsp70 in multiple in vitro binding assays. Finally, in addition to hsc70 and hsp70, AQP2 interacts with several other key components of the endocytotic machinery in co-IP assays, including clathrin, dynamin, and AP2. To summarize, we have identified the 70-kDa heat shock proteins as a AQP2 interactors and have shown for hsc70 that this interaction is involved in AQP2 trafficking. The trafficking of aquaporin-2 (AQP2) involves multiple complex pathways, including regulated, cAMP-, and cGMP-mediated pathways, as well as a constitutive recycling pathway. Although several accessory proteins have been indirectly implicated in AQP2 recycling, the direct protein-protein interactions that regulate this process remain largely unknown. Using yeast two-hybrid screening of a human kidney cDNA library, we have identified the 70-kDa heat shock proteins as AQP2-interacting proteins. Interaction was confirmed by mass spectrometry of proteins pulled down from rat kidney papilla extract using a GST-AQP2 C-terminal fusion protein (GST-A2C) as a bait, by co-immunoprecipitation (IP) assays, and by direct binding assays using purified hsc70 and the GST-A2C. The direct interaction of AQP2 with hsc70 is partially inhibited by ATP, and the Ser-256 residue in the AQP2 C terminus is important for this direct interaction. Vasopressin stimulation in cells enhances the interaction of hsc70 with AQP2 in IP assays, and vasopressin stimulation in vivo induces an increased co-localization of hsc70 and AQP2 on the apical membrane of principal cells in rat kidney collecting ducts. Functional knockdown of hsc70 activity in AQP2 expressing cells results in membrane accumulation of AQP2 and reduced endocytosis of rhodamine-transferrin. Our data also show that AQP2 interacts with hsp70 in multiple in vitro binding assays. Finally, in addition to hsc70 and hsp70, AQP2 interacts with several other key components of the endocytotic machinery in co-IP assays, including clathrin, dynamin, and AP2. To summarize, we have identified the 70-kDa heat shock proteins as a AQP2 interactors and have shown for hsc70 that this interaction is involved in AQP2 trafficking. Aquaporin 2 is expressed in principal cells of the kidney collecting duct and mediates water absorption and urinary concentration in response to vasopressin (VP). 4The abbreviations used are: VPvasopressinAQP2aquaporin-2GSTglutathione S-transferaseco-IPco-immunoprecipitationPBSphosphate-buffered salineNBDnucleotide binding domainWTwild type. 4The abbreviations used are: VPvasopressinAQP2aquaporin-2GSTglutathione S-transferaseco-IPco-immunoprecipitationPBSphosphate-buffered salineNBDnucleotide binding domainWTwild type. Upon stimulation by VP, AQP2 accumulates on the plasma membrane of collecting duct principal cells (1Brown D. Am. J. Physiol. 2003; 284: F893-F901Crossref PubMed Scopus (219) Google Scholar, 2Knepper M.A. Wade J.B. Terris J. Ecelbarger C.A. Marples D. Mandon B. Chou C.L. Kishore B.K. Nielsen S. Kidney Int. 1996; 49: 1712-1717Abstract Full Text PDF PubMed Scopus (151) Google Scholar, 3Nielsen S. Kwon T.H. Christensen B.M. Promeneur D. Frokiaer J. Marples D. J. Am Soc. Nephrol. 1999; 10: 647-663PubMed Google Scholar, 4Sasaki S. Kuwahara M. Yamashita Y. Marumo F. Nephrol. Dial. Transplant. 2000; 15: 21-22Crossref PubMed Scopus (17) Google Scholar). Activation of both cAMP-dependent, VP/forskolin-stimulated and cAMP-independent, cGMP-responsive signaling pathways stimulates membrane accumulation of AQP2 (5van Balkom B.W. Savelkoul P.J. Markovich D. Hofman E. Nielsen S. van der Sluijs P. Deen P.M. J. Biol. Chem. 2002; 277: 41473-41479Abstract Full Text Full Text PDF PubMed Scopus (197) Google Scholar). Recent evidence also indicates that cAMP- and cGMP-independent membrane accumulation of AQP2 can also occur in the absence of hormonal stimulation (6Sun T.X. Van Hoek A. Huang Y. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. 2002; 282: F998-F1011Crossref PubMed Scopus (101) Google Scholar, 7Katsura T. Verbavatz J.M. Farinas J. Ma T. Ausiello D.A. Verkman A.S. Brown D. S. A. PubMed Scopus Google Scholar, T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google as by endocytosis (6Sun T.X. Van Hoek A. Huang Y. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. 2002; 282: F998-F1011Crossref PubMed Scopus (101) Google Scholar, T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google and by the E. M. Am. J. Physiol. PubMed Scopus Google Scholar, E. D. B. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). vasopressin aquaporin-2 co-immunoprecipitation binding type. vasopressin aquaporin-2 co-immunoprecipitation binding type. Although the pathways of AQP2 trafficking remain to is the direct interaction of AQP2 with a of accessory that involved in interactions is a in the of as well as and of AQP2 that to the response to in of the proteins to involved in trafficking in have also been implicated in AQP2 including components as and and as well as protein and proteins (1Brown D. Am. J. Physiol. 2003; 284: F893-F901Crossref PubMed Scopus (219) Google Scholar, E. Physiol. 2000; PubMed Google Scholar, S. Marples D. M. M. M. J. PubMed Scopus Google Scholar, M. M. PubMed Scopus Google Scholar, D. A. M.A. Nielsen S. Am. J. Physiol. PubMed Google Scholar, T. Nielsen S. Mandon B. Terris J. Kishore B.K. M.A. Am. J. Physiol. Google Scholar). direct evidence the interaction of AQP2 with components of the trafficking machinery is in this a identified AQP2 as a of a including and the protein Y. S. T. Y. T. Kuwahara M. T. M. S. PubMed Scopus Google Scholar). AQP2 was several that water in the collecting duct a endocytotic process D. PubMed Scopus Google Scholar). we have shown that AQP2 this endocytotic in a a constitutive the process of endocytosis with using results in and plasma membrane accumulation of AQP2 in (6Sun T.X. Van Hoek A. Huang Y. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. 2002; 282: F998-F1011Crossref PubMed Scopus (101) Google Scholar, T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google and in the rat kidney M. Brown D. Am. J. Physiol. PubMed Scopus Google Scholar). The endocytotic is of the of endocytosis in cells and is by the of proteins from the complex of protein-protein interactions is involved in the and of this is and and involves of protein that the activity of both and Biol. PubMed Scopus Google Scholar, 2003; PubMed Scopus Google Scholar). have the well of heat shock protein as in protein and and to a in endocytosis D. B. Chem. PubMed Scopus Google Scholar, R. B. E. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, B. Full Text Full Text PDF PubMed Scopus Google Scholar, M. J. Biol. 2002; PubMed Scopus Google Scholar). hsc70 an important with in the of endocytosis in vitro and in vivo M. J. Biol. 2002; PubMed Scopus Google Scholar, Full Text PDF PubMed Scopus Google Scholar, S. Full Text Full Text PDF PubMed Scopus Google Scholar, J. Biol. PubMed Scopus Google Scholar). hsc70 been to to membrane as and and to F. J. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar, J.M. Biol. PubMed Scopus Google Scholar, P. S. Full Text Full Text PDF PubMed Scopus Google Scholar, A. J. 2002; PubMed Scopus Google Scholar, Y. S. Biol. PubMed Scopus Google Scholar). data have also shown that heat shock protein is in the papilla of and a that hsp70 was identified in kidney cells Kwon Biol. 2002; PubMed Scopus Google Scholar). of the kidney water an of hsp70 and a of the collecting duct of vasopressin that the of heat shock protein is increased in response to vasopressin Balkom B.W. Chou C.L. M.A. Am. J. Physiol. PubMed Scopus Google Scholar, Balkom B.W. van M. S. Bouley R. van der Sluijs P. Brown D. Deen P.M. J. Biol. Chem. 2003; Full Text Full Text PDF PubMed Scopus Google Scholar). data and the of the hsc70 in trafficking the that the 70-kDa heat shock proteins also a in AQP2 this we to show that both hsc70 and hsp70 with AQP2 and that the interaction with hsc70 is important for AQP2 and was from was by and protein from hsc70 and purified hsp70 from hsp70 was using a from was from the of the was from the and 2 by S. J. Biol. 2000; PubMed Scopus Google Scholar). Our the AQP2 C terminus been T. Verbavatz J.M. Brown D. J. Biol. PubMed Scopus Google Scholar). hsc70 and hsp70 and from from from and of expressing to as in and and in the of the and been T. Verbavatz J.M. Farinas J. Ma T. Ausiello D.A. Verkman A.S. Brown D. S. A. PubMed Scopus Google Scholar, T. Ausiello D.A. Brown D. Am. J. Physiol. PubMed Google Scholar). in with with with hsc70 and co-IP of AQP2 and hsc70 from AQP2 cells using an and kidney papilla extract by protein to co-IP hsc70 is in the co-IP complex pulled down with the AQP2 was also using as co-IP of hsc70 and AQP2 using and show that AQP2 hsc70 to co-IP binding from and this co-IP an interaction AQP2 and C that AQP2 is to co-IP with hsc70 also co-IP with hsp70 from rat kidney papilla hsc70 and hsp70 used in this co-IP The of AQP2 from cells is that from kidney papilla extract of the of in proteins The rat aquaporin-2 was a as a for yeast two-hybrid protein was as a fusion of the for from the terminus of from the of and from the of The of this C-terminal was also the as a fusion protein with a on terminus R. S. T. McLaughlin M. Ausiello D.A. Brown D. J. 2000; PubMed Scopus Google Scholar). the to and The kidney cDNA in the yeast was from expressing hsc70 an hsc70 and a by J. Biol. PubMed Scopus Google Scholar). of the of cells with in (6Sun T.X. Van Hoek A. Huang Y. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. 2002; 282: F998-F1011Crossref PubMed Scopus (101) Google Scholar). of of was used to for cells and used for endocytosis assays with rhodamine-transferrin. and yeast two-hybrid screening was in that and The and the The and the and was to both and to The the was with of using a The cells and for and was and that of the in the The the protein was from yeast cells and to The on and the was and for two-hybrid assays to the interaction of and as from the used to with to of The also to with to for cells on that with with by of and Kidney in in to cells with and and and a of on for and a and for to The used for down and co-immunoprecipitation assays. Kidney papilla extract was from rat The from with and in a a with a the in and rat kidney papilla extract was used for and co-IP and of AQP2 was used for fusion protein GST-AQP2 C-terminal fusion proteins using of in E. was by for by and in was to to the for with with with and and with the fusion protein was with reduced and and GST-AQP2 fusion proteins to and and papilla on a for of the protein was with of and to of rat kidney papilla extract with for 2 the with and to proteins. Finally, the pulled down was by and mass the pulled down proteins from rat kidney papilla with and and for mass spectrometry by purified fusion proteins also used to down purified hsc70 as of and was with of and of hsc70 for 2 the pulled down was to and using as a to the hsc70 The from and was using The of the of the of AQP2 using purified with hsc70 hsp70, in the absence of from using the of the of the of hsc70 AQP2 from and rat kidney papilla extract on protein for to co-IP with of and of rat kidney papilla extract with for 2 with and the co-IP to and using a of with protein used as a using from cells was also used as a in and of and kidney was by with as R. S. T. McLaughlin M. Ausiello D.A. Brown D. J. 2000; PubMed Scopus Google with to vasopressin for in the of the and for in in in and for The kidney with for to D. J. McLaughlin M. A. R. S. Biol. 1996; PubMed Scopus Google and with in AQP2 hsc70 to kidney and for with and for with in and by and cells and as on with in for for with with for and to for AQP2 hsc70 as T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google Scholar). of of AQP2 to the plasma membrane was by of AQP2 in from cells with and cells with expressing hsc70 and the The by AQP2 in a of of the in of cells was using as R. S. T. McLaughlin M. Ausiello D.A. Brown D. J. 2000; PubMed Scopus Google and was with the The of was shown to an in membrane the AQP2 trafficking by with of a of and in on a and on the for with and for with a concentration of rat a concentration of in the with and and of in for several with the on for and with water and Finally, in a and in endocytosis using as a of endocytosis was in a as we for a endocytosis using T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google Scholar). cells on with for cells with for by with cells with in for with and The of was using a and of the of in the in vitro binding assays, co-IP and assays, was as the of on was using and the of of hsc70 the of AQP2 in the the an a from the in the for was for The of from was using a of data from using of Finally, the of hsc70 AQP2 was in in the the and the indicates an AQP2-interacting and of the by two-hybrid screening was to for AQP2-interacting proteins as identified binding the data in using cDNA and an of both human hsp70 and hsc70 from an of both human hsp70 and hsc70 from The of cDNA the from of heat shock protein and is to hsp70 to the hsc70 in this purified was used to down from rat kidney papilla The purified fusion protein was by and by using AQP2 The AQP2 the fusion protein and proteins by and the of the proteins also in of with kidney several proteins in the and in the The was identified by mass spectrometry as Interaction of hsc70 and AQP2 by and was using purified and with and rat kidney papilla a of the pulled down with an hsc70 and hsp70 is to down hsc70 from both rat kidney papilla extract and and 2 show that hsc70 is in from cells and kidney is also to down hsp70 from hsc70 hsp70 was in down using with kidney The interaction of AQP2 and was by co-IP using AQP2 and hsc70 that the AQP2 C terminus was to AQP2 and hsc70 from from from with protein was to AQP2 from cells from cells co-IP was also using rat kidney papilla extract show that AQP2 with also with the protein hsp70 from kidney papilla and AQP2 in this co-immunoprecipitation in to cells that the of The of AQP2 from cells is that from kidney papilla extract of the of the on the C terminus of AQP2 in hsc70 and hsp70 with hsc70 can with AQP2 C-terminal in purified hsc70 and purified used to that hsc70 is pulled down by by hsc70 and interacts with binding in an the interaction of hsc70 and AQP2 C terminus was in the of and the of the binding of hsc70 and was to that in the absence of in the of ATP, the binding of hsc70 to was reduced The of of the of hsc70 is in and that is a of the binding of hsc70 to in the of data that the interaction of hsc70 and AQP2 in vitro is the interaction of hsc70 and was also using hsp70 and that to hsp70 can to AQP2 in and this interaction is also the interaction of hsp70 and AQP2 show on the binding of hsp70 with GST-A2C. from To the of the of Ser-256 on the interaction of AQP2 with and and that of Ser-256 and purified GST-AQP2 fusion proteins show a in and as R. T. R. PubMed Scopus Google and of The was using hsc70 with GST-AQP2 proteins is important for direct in vitro interaction of hsc70 and GST-AQP2 fusion hsc70 was with purified and proteins and in this show that the binding of hsc70 is with with hsc70 with 2 and hsc70 with hsc70 with hsc70 with hsc70 was using and fusion proteins using in the of the of of the of of hsc70 GST-AQP2 fusion proteins the the of the hsc70 pulled down by the was and the of the hsc70 pulled down by the AQP2 was also for the the in the for was from and and from a the of the with the from the The was in from indicates Ser-256 with the constitutive with a constitutive the interaction of AQP2 and The of binding of hsc70 by GST-AQP2 was in B. from and a in binding of hsc70 Ser-256 was to that an Ser-256 residue is important for the interaction of AQP2 with hsc70 in Interaction of AQP2 with hsc70 by been well that vasopressin AQP2 trafficking both in vitro and in have using that AQP2 interacts with and we the interaction of AQP2 and hsc70 in The interaction of AQP2 and hsc70 in the and absence of stimulation was by co-IP using cells with for The results show an increased of hsc70 in the co-IP complex to of with VP, 2 that the interaction of AQP2 and hsc70 is The of hsc70 in the of stimulation was also using and that is of the of hsc70 the using and show that is also a increased of hsp70 in co-IP with for and 2 shown in a the of hsp70 the of the hsc70 and AQP2 of the interaction of hsc70 and AQP2 was we that hsc70 is with AQP2 in principal cells of kidney collecting by of rat kidney using AQP2 and hsc70 was the of AQP2 was on and vasopressin of for was an accumulation of AQP2 on the plasma membrane as was also a increased apical membrane for hsc70 in the principal cells of the collecting duct in the was a co-localization of hsc70 and AQP2 in this apical this co-localization was in the with hsp70 and AQP2 using the kidney that hsp70, and partially with AQP2 The of hsc70 to the apical membrane was also by using for AQP2 and hsc70 of AQP2 and hsc70 on the apical plasma membrane of principal cells from in with using for and hsc70 in this kidney of hsc70 and in cells on the apical membrane with AQP2 on apical membrane in kidney by of AQP2 and hsc70 was in both and rat kidney and hsc70 was with the and is by AQP2 was with and is by vasopressin hsc70 and AQP2 and in A. stimulation (AQP2) on to the apical and with a on the to on the plasma membrane shown as in C that in the kidney with with hsc70 with in of hsc70 of AQP2 and in AQP2 the of hsc70 activity on AQP2 we the trafficking of AQP2 down hsc70 by an cells with hsc70 hsc70 and and with to AQP2 membrane accumulation of AQP2 in cells with expressing the and a with expressing the was as we have (6Sun T.X. Van Hoek A. Huang Y. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. 2002; 282: F998-F1011Crossref PubMed Scopus (101) Google Scholar). was of AQP2 trafficking in cells with of the of AQP2 in cells with hsc70 a in with an in membrane using a that we have R. S. T. McLaughlin M. Ausiello D.A. Brown D. J. 2000; PubMed Scopus Google Scholar). data of hsc70 activity results in membrane accumulation of To this membrane accumulation of AQP2 to the of hsc70 in we a endocytosis using cells with and to a Our data show that is in cells with and cells expressing hsc70 is reduced and a membrane accumulation of the in cells expressing the the hsc70 indicates that endocytosis is inhibited by the was with the The of hsp70 in cells was also is an of hsp70 in cells with both hsc70 and the is in cells with hsc70 was a in hsp70 in cells with the hsc70 a in cells in hsc70 been the of hsp70 the of hsc70 on AQP2 trafficking in cells is knockdown of hsc70 activity with endocytosis in a to the well of on this process (6Sun T.X. Van Hoek A. Huang Y. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. 2002; 282: F998-F1011Crossref PubMed Scopus (101) Google Scholar, T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google Scholar). The of hsc70 in cells with is also shown in the of hsc70 was in cells with both hsc70 and hsc70 of hsp70 and hsc70 in cells with The of hsp70 and hsc70 was in cells with expressing hsc70 hsc70 of cells to and using and was using is a of hsp70 in cells with hsc70 and a in cells with hsc70 with cells with The of hsc70 was using the the and was to in cells with expressing both and was AQP2 with of the we co-immunoprecipitation to AQP2-interacting proteins in the endocytosis pathway. that in addition to AQP2 also in a complex with clathrin, dynamin, and the protein AP2. the of endocytotic complex proteins in AQP2 as from data that AQP2 is this (6Sun T.X. Van Hoek A. Huang Y. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. 2002; 282: F998-F1011Crossref PubMed Scopus (101) Google Scholar, T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google Scholar, D. PubMed Scopus Google Scholar). AQP2 and the plasma membrane (1Brown D. Am. J. Physiol. 2003; 284: F893-F901Crossref PubMed Scopus (219) Google this can in addition by a of including to the of water and concentration (1Brown D. Am. J. Physiol. 2003; 284: F893-F901Crossref PubMed Scopus (219) Google Scholar, T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google Scholar, E. D. B. F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, R. S. T. McLaughlin M. Ausiello D.A. Brown D. J. 2000; PubMed Scopus Google Scholar, M. Y. Marumo F. S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). complex recycling AQP2 with a of accessory proteins that involved in the trafficking of have been implicated in AQP2 trafficking by and indirectly by co-localization on in to with AQP2 the protein is unknown. show that heat shock protein interacts with was using a of including yeast two-hybrid and co-immunoprecipitation assays from and and direct interaction assays using purified proteins. The cDNA that we have identified from the yeast two-hybrid the C-terminal of the binding of hsp70 and hsc70 an of the data using we to for and proteins in AQP2 C-terminal as in proteins that to the of protein and other proteins J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, M. A. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar, S. D.A. J. S. E. J. M. Biol. PubMed Scopus Google Scholar, S. S. PubMed Scopus Google Scholar, J. S. J. PubMed Scopus Google Scholar). we for the binding as in the the C terminus of protein and the human protein in the AQP2 C-terminal E. 1999; PubMed Scopus Google Scholar, T. E. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, E. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, C.A. P. Y. Biol. 1999; PubMed Scopus Google Scholar, A. D. B.K. T. PubMed Scopus Google Scholar). the and of the interaction of hsc70 and AQP2 to data that the Ser-256 residue in the AQP2 C terminus is involved in this interaction. Ser-256 is to the protein and protein on and is increased vasopressin to cells and with increased plasma membrane accumulation of AQP2 (1Brown D. Am. J. Physiol. 2003; 284: F893-F901Crossref PubMed Scopus (219) Google Scholar, S. Marumo F. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Our in vitro binding data that vasopressin enhances the interaction of AQP2 and heat shock protein in key is this interaction is the and this have on AQP2 hsc70 is a and was shown to and fusion with a in the R. B. E. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, M. J. Biol. 2002; PubMed Scopus Google Scholar, Full Text PDF PubMed Scopus Google Scholar). hsc70 have been shown to the endocytosis of M. J. Biol. 2002; PubMed Scopus Google Scholar, S. Full Text Full Text PDF PubMed Scopus Google Scholar, J. Biol. PubMed Scopus Google and is that hsc70 is of the endocytotic machinery that endocytosis R. B. E. J. Biol. Chem. 2000; Full Text Full Text PDF PubMed Scopus Google Scholar, J. Biol. PubMed Scopus Google Scholar). we that hsc70 is for AQP2 the of the vasopressin as well as the AQP2 constitutive recycling pathway. show that knockdown of hsc70 activity induces membrane accumulation of AQP2 in cells and a of of reduced that involved in the interaction the recycling process is of residue Ser-256 in the AQP2 C we that stimulation enhances the interaction of AQP2 and this of a in the response is and A. J. T. J. R. M. and D. interaction was the AQP2 is and accumulates on the plasma and data show that induces co-localization of hsc70 and AQP2 on the apical plasma membrane of collecting duct principal cells in with that AQP2 in accumulates the and with hsc70 an increased of hsc70 with AQP2 the of activity M. A. R. M. J. 2000; Google Scholar). of we that the interaction of hsc70 and as the was in that a residue this for the interaction of hsc70 and is the is and to that hsc70 binding to and membrane accumulation of AQP2 by have shown that of endocytosis is to membrane accumulation of AQP2 in T.X. Bouley R. McLaughlin M. Brown D. Am. J. Physiol. PubMed Scopus Google Scholar). the also hsc70 binding membrane accumulation of other also been that constitutive of Ser-256 in the AQP2 to to for M. A. R. Nielsen S. Kwon T.H. M. J. 2003; PubMed Scopus Google Scholar). shock protein been well to involved in Y. S. Biol. PubMed Scopus Google Scholar, E. P.J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google and the of in yeast R. T. R. PubMed Scopus Google is also that the and the of Ser-256 and and to Ser-256 in vivo a Finally, is that other in the AQP2 C including a protein C and a (5van Balkom B.W. Savelkoul P.J. Markovich D. Hofman E. Nielsen S. van der Sluijs P. Deen P.M. J. Biol. Chem. 2002; 277: 41473-41479Abstract Full Text Full Text PDF PubMed Scopus (197) Google also involved in heat shock protein interaction and of AQP2 trafficking in the AQP2 recycling pathway. hsc70 and hsp70 have of as well as and have been to in and data have shown that have D. A. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar, Y. Y. PubMed Scopus Google Scholar). both hsc70 and hsp70 identified by a yeast two-hybrid and shown to with AQP2 by multiple in vitro assays. the interaction of and can by and hsc70 and hsp70 to interaction with have on AQP2 trafficking is by data that hsc70 to with AQP2 the plasma membrane of principal cells in response to hsp70 in the hsc70 is an important protein trafficking of other membrane as a and multiple that endocytosis and F. J. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar, J.M. Biol. PubMed Scopus Google Scholar, P. S. Full Text Full Text PDF PubMed Scopus Google Scholar, A. J. 2002; PubMed Scopus Google Scholar). the that the interaction of AQP2 and hsc70 occur multiple co-localization show that important of interaction is the plasma on AQP2 endocytosis is by co-IP data the of other components involved in endocytosis dynamin, and in the of the interaction in other for AQP2 recycling, and is that heat shock protein is involved in multiple of AQP2 trafficking and for the of the in is from of for the of and of and