The Chloride Channel ClC-4 Co-localizes with Cystic Fibrosis Transmembrane Conductance Regulator and May Mediate Chloride Flux across the Apical Membrane of Intestinal Epithelia

Cystic fibrosis (CF) causing mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) lead to mislocalization of CFTR protein from the brush border membrane of epithelial tissues and/or its dysfunction as a chloride channel. In initial reports, it was proposed that certain channels from the ClC family of chloride channels may provide compensatory or alternative pathways for epithelial chloride secretion in tissues from cystic fibrosis patients. In the present work, we provide the first evidence that ClC-4 protein is functionally expressed on the surface of the intestinal epithelium and hence, is appropriately localized to act as a therapeutic target in this CF-affected tissue. We show using confocal and electron microscopy that ClC-4 co-localizes with CFTR in the brush border membrane of the epithelium lining intestinal crypts in mouse and human tissues. In Caco-2 cells, a cell line thought to model human enterocytes, ClC-4 protein is expressed on the cell surface and also partially co-localizes with EEA1 and transferrin, marker molecules of early and recycling endosomes, respectively. Hence, like CFTR, ClC-4 may cycle between the plasma membrane and endosomal compartment. Furthermore, we show that ClC-4 functions as a chloride channel on the surface of these epithelial cells as antisense ClC-4 cDNA expression reduced the amplitude of endogenous chloride currents by 50%. These studies provide the first evidence that ClC-4 is endogenously expressed and may be functional in the brush border membrane of enterocytes and hence should be considered as a candidate channel to provide an alternative pathway for chloride secretion in the gastrointestinal tract of CF patients. Cystic fibrosis (CF) causing mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) lead to mislocalization of CFTR protein from the brush border membrane of epithelial tissues and/or its dysfunction as a chloride channel. In initial reports, it was proposed that certain channels from the ClC family of chloride channels may provide compensatory or alternative pathways for epithelial chloride secretion in tissues from cystic fibrosis patients. In the present work, we provide the first evidence that ClC-4 protein is functionally expressed on the surface of the intestinal epithelium and hence, is appropriately localized to act as a therapeutic target in this CF-affected tissue. We show using confocal and electron microscopy that ClC-4 co-localizes with CFTR in the brush border membrane of the epithelium lining intestinal crypts in mouse and human tissues. In Caco-2 cells, a cell line thought to model human enterocytes, ClC-4 protein is expressed on the cell surface and also partially co-localizes with EEA1 and transferrin, marker molecules of early and recycling endosomes, respectively. Hence, like CFTR, ClC-4 may cycle between the plasma membrane and endosomal compartment. Furthermore, we show that ClC-4 functions as a chloride channel on the surface of these epithelial cells as antisense ClC-4 cDNA expression reduced the amplitude of endogenous chloride currents by 50%. These studies provide the first evidence that ClC-4 is endogenously expressed and may be functional in the brush border membrane of enterocytes and hence should be considered as a candidate channel to provide an alternative pathway for chloride secretion in the gastrointestinal tract of CF patients. cystric fibrosis cystic fibrosis transmembrane conductance regulator glutathione S-transferase reverse transcriptase phosphate-buffered saline green fluorescent protein The disease cystic fibrosis (CF)1 affects the epithelium lining multiple organs including the respiratory tract, the gastrointestinal tract, sweat ducts, and the reproductive organs (1Sheppard D.N. Welsh M.J. Physiol. Rev. 1999; 79: S23-45Crossref PubMed Scopus (813) Google Scholar). Normally, the protein product of the CF gene, the cystic fibrosis transmembrane conductance regulator (CFTR) resides on the apical surface of these epithelia. However, the most common disease causing mutation in CFTR (i.e. CFTRΔF508) promotes misfolding, leading to its mislocalization and degradation in intracellular compartments (2Cheng S.H. Gregory R.J. Marshall J. Paul S. Souza D.W. White G.A. O'Riordan C.R. Smith A.E. Cell. 1990; 63: 827-834Abstract Full Text PDF PubMed Scopus (1427) Google Scholar). Alternatively, other mutations lead to alterations in CFTR function as a phosphorylation and nucleotide-regulated anion channel (3Welsh M.J. Smith A.E. Cell. 1993; 73: 1251-1254Abstract Full Text PDF PubMed Scopus (1235) Google Scholar). The anion channel function of CFTR is thought to provide the primary driving force for fluid transport and clearance of mucus and bacteria and lack of this function is the major cause for mucus obstruction in CF-affected organs. As proposed initially by Clarkeet al. (4Clarke L.L. Grubb B.R. Yankaskas J.R. Cotton C.U. McKenzie A. Boucher R.C. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 479-483Crossref PubMed Scopus (312) Google Scholar), non-CFTR chloride channels could compensate for lack of CFTR in certain tissues if they were appropriately localized on the apical, brush border membrane of the epithelium and could be opened under physiological conditions. In fact, we recently reported that increased basal chloride secretion correlates with amelioration of disease severity in intestinal tissues of a subpopulation ofCftr-deficient mice (5Gyomorey K. Rozmahel R. Bear C.E. Pediatr. Res. 2000; 48: 731-734Crossref PubMed Scopus (17) Google Scholar). Therefore, there is a compelling rationale for studying the expression of other chloride channels that may mediate chloride secretion across the gastrointestinal epithelium as these proteins may provide a strategic therapeutic target for treatment of cystic fibrosis. There are nine mammalian members of the superfamily of voltage-gated ClC channels (6Jentsch T.J. Friedrich T. Schriever A. Yamada H. Pflugers Arch. 1999; 437: 783-795Crossref PubMed Scopus (297) Google Scholar). ClC-1, ClC-2, ClCKa, and ClCKb are closely related, ClC-3, ClC-4, and ClC-5 form another arm of the family and finally, ClC-6 and ClC-7 comprise a distinct branch. Some of these family members exhibit quite a restricted tissue expression and hence are unlikely to contribute to chloride transport across the epithelium of the gastrointestinal tract, i.e. ClC-1 is only expressed in muscle tissue and ClCKa and ClCKb are expressed exclusively in the kidney (7Waldegger S. Jentsch T.J. J. Am. Soc. Nephrol. 2000; 11: 1331-1339PubMed Google Scholar, 8Uchida S. Am. J. Physiol. Renal Physiol. 2000; 279: F802-808Crossref PubMed Google Scholar, 9Kieferle S. Fong P. Bens M. Vandewalle A. Jentsch T.J. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 6943-6947Crossref PubMed Scopus (246) Google Scholar). On the other hand, ClC-2, ClC-3, ClC-4, ClC-5, ClC-6, and ClC-7 are relatively widely distributed. However, most of these family members, with the exception of ClC-2 and possibly ClC-3, are thought to be primarily expressed on intracellular membranes (6Jentsch T.J. Friedrich T. Schriever A. Yamada H. Pflugers Arch. 1999; 437: 783-795Crossref PubMed Scopus (297) Google Scholar, 10Vandewalle A. Cluzeaud F. Peng K.C. Bens M. Luchow A. Gunther W. Jentsch T.J. Am. J. Physiol. Cell Physiol. 2001; 280: C373-381Crossref PubMed Google Scholar,11Wang S.S. Devuyst O. Courtoy P.J. Wang X.T. Wang H. Wang Y. Thakker R.V. Guggino S. Guggino W.B. Hum. Mol. Genet. 2000; 9: 2937-2945Crossref PubMed Scopus (271) Google Scholar). We have recently reported that ClC-2 is endogenously expressed at a unique location in intestinal epithelia, in proximity to the tight junctions at the apical boundary between interacting differentiated enterocytes (12Gyomorey K. Yeger H. Ackerley C. Garami E. Bear C.E. Am. J. Physiol. Cell Physiol. 2000; 279: C1787-1794Crossref PubMed Google Scholar, 13Mohammad-Panah R. Gyomorey K. Rommens J. Choudhury M. Li C. Wang Y. Bear C.E. J. Biol. Chem. 2001; 276: 8306-8313Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). Furthermore, we showed using an antisense strategy that endogenous ClC-2 channels in intestinal epithelial cells can mediate chloride flux. Hence, we proposed that ClC-2 contributes to chloride secretion across certain epithelia. These findings were substantiated in a in the chloride across the epithelium was in the C. Jentsch T.J. J. 2001; PubMed Scopus Google Scholar). However, as the the of ClC-2 in intestinal tissue may only be in the and physiological of as studies that functions in intracellular (6Jentsch T.J. Friedrich T. Schriever A. Yamada H. Pflugers Arch. 1999; 437: 783-795Crossref PubMed Scopus (297) Google Scholar, T. T. Jentsch T.J. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google and a for channels on the plasma membrane C. S. J.R. PubMed Scopus Google Scholar, K. Li Am. J. Physiol. Physiol. 2000; 279: PubMed Google Scholar, P. J. A. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). However, in studies of expression in the human intestinal cell we that protein was expressed in intracellular R. Gyomorey K. Rommens J. Choudhury M. Li C. Wang Y. Bear C.E. J. Biol. Chem. 2001; 276: 8306-8313Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). in are with disease in a kidney disease by and (6Jentsch T.J. Friedrich T. Schriever A. Yamada H. Pflugers Arch. 1999; 437: 783-795Crossref PubMed Scopus (297) Google Scholar, S. Jentsch T.J. J. Am. Soc. Nephrol. 2000; 11: 1331-1339PubMed Google Scholar, O. Courtoy P.J. R. Thakker R.V. Hum. Mol. Genet. 1999; PubMed Scopus Google Scholar, Gunther W. S.H. A. M. O. Jentsch T.J. Thakker R.V. Hum. Mol. Genet. PubMed Scopus Google Scholar). of the expression of ClC-5 in differentiated epithelial tissues that this channel resides in intracellular In mice show a in of protein from of the S.S. Devuyst O. Courtoy P.J. Wang X.T. Wang H. Wang Y. Thakker R.V. Guggino S. Guggino W.B. Hum. Mol. Genet. 2000; 9: 2937-2945Crossref PubMed Scopus (271) Google Scholar). al. A. Cluzeaud F. Peng K.C. Bens M. Luchow A. Gunther W. Jentsch T.J. Am. J. Physiol. Cell Physiol. 2001; 280: C373-381Crossref PubMed Google showed that ClC-5 protein in intracellular i.e. and in the intestinal Hence, in of these studies of it unlikely that ClC-5 contribute to chloride currents across the apical membrane of the intestinal ClC-4 is to be expressed in kidney T.J. Gunther W. M. J. Physiol. PubMed Scopus Google Scholar, K. A. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar, A. A. Hum. Mol. Genet. 1994; PubMed Scopus Google Scholar), and K. Garami E. K. Rommens J. Bear C. Pflugers Arch. 2001; PubMed Scopus Google Scholar), the and function of ClC-4 protein in these tissues to be In present work, we provide the first evidence that ClC-4 is endogenously expressed in the apical plasma membrane of and human Furthermore, we show using an antisense that endogenous ClC-4 protein chloride currents across the plasma membrane of Caco-2 cells, cells that model human the of the ClC family of chloride these findings were was that the of ClC-4 be to that of the channel ClC-5 and ClC-3, and primarily in the membranes of intracellular ClC-4 is the first chloride channel protein to with CFTR on the brush border membrane of intestinal epithelial cells endogenously it could mediate chloride currents the Hence, these a for ClC-4 in intestinal chloride secretion and that it may be of functionally CFTR in Caco-2 cells were from the were in with of and at in an of cells were of ClC-4 in differentiated epithelia, Caco-2 cells were on and for reported to a differentiated R. Bear C. W. E. T. J.R. M. J. 11: PubMed Scopus Google Scholar). Caco-2 cell at were with antisense ClC-4 cDNA or using and the was as R. Gyomorey K. Rommens J. Choudhury M. Li C. Wang Y. Bear C.E. J. Biol. Chem. 2001; 276: 8306-8313Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). were from a of were in and at of a human by a were for electron of ClC-4 protein in mouse tissues and Caco-2 cells was by as in (12Gyomorey K. Yeger H. Ackerley C. Garami E. Bear C.E. Am. J. Physiol. Cell Physiol. 2000; 279: C1787-1794Crossref PubMed Google Scholar, 13Mohammad-Panah R. Gyomorey K. Rommens J. Choudhury M. Li C. Wang Y. Bear C.E. J. Biol. Chem. 2001; 276: 8306-8313Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). In tissues from mouse were and at to the The was at to a membrane Caco-2 cells were and at for at to a membrane of was by using at a of was a of mouse ClC-4 cDNA from E. The was to a to the the was with a of the from ClC-4, or a protein of at cDNA by T. protein was using the was from Caco-2 cell by in and a R.J. A.E. PubMed Scopus Google Scholar). was on and to membranes were with and with ClC-4 cDNA by PubMed Scopus Google Scholar). of with at The were to for at at with on of ClC-4 was by reverse using the with human ClC-4, and antisense a product of The of the was by The antisense ClC-4 was by the ClC-4 and the that the on this reverse the of the to the antisense The ClC-4 with the human Caco-2 cells were with with in and with in was by using in for to primary were in as early mouse of H. P. of T. and The cells were for at in primary and in and with or or and primary in were by the using and In we Caco-2 cells for at with to and in cells we with the ClC-4 of of were in for to were for as were with a on a with an confocal of from a human from and Caco-2 cells on were and for and J. PubMed Scopus Google Scholar). The were with the ClC-4 in were as (12Gyomorey K. Yeger H. Ackerley C. Garami E. Bear C.E. Am. J. Physiol. Cell Physiol. 2000; 279: C1787-1794Crossref PubMed Google Scholar). the of the primary or or a In of human were first with the ClC-4 and with a the of CFTR ClC-4 the were for an in the CFTR with The were with with and with and lead to in a electron were with an Caco-2 cells were with at on for as R. Gyomorey K. Rommens J. Choudhury M. Li C. Wang Y. Bear C.E. J. Biol. Chem. 2001; 276: 8306-8313Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). were to a of for antisense ClC-4 and antisense was also to the to Caco-2 cell membrane currents were using cell as R. Gyomorey K. Rommens J. Choudhury M. Li C. Wang Y. Bear C.E. J. Biol. Chem. 2001; 276: 8306-8313Abstract Full Text Full Text PDF PubMed Scopus (69) Google Scholar). were and with an and The the and and were to The was with the are as the were using the of or were considered ClC-4 protein we a a protein of the of mouse As the ClC-4 expression in the K. A. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar), a in of mouse The to a protein of in to the ClC-4 from the primary of is for ClC-4 as it is by the of the of ClC-4 a protein the of the closely channel ClC-5 this we a in of mouse that ClC-4 protein is also endogenously expressed in intestinal tissue ClC-4 and protein could also be in the human intestinal epithelial cell ClC-4 in Caco-2 cells was as a by using a mouse ClC-4 as of was also in Caco-2 ClC-4 expression in Caco-2 cells was using with to human ClC-4 ClC-4 protein expression in Caco-2 cells was by using the As in the of mouse ClC-4 protein was as a protein in Caco-2 cells, that ClC-4 is also expressed in human intestinal epithelial cells The of ClC-5 protein in intestinal tissue by Vandewalle al. A. Cluzeaud F. Peng K.C. Bens M. Luchow A. Gunther W. Jentsch T.J. Am. J. Physiol. Cell Physiol. 2001; 280: C373-381Crossref PubMed Google using a by Gunther al. W. Luchow A. Cluzeaud F. Vandewalle A. Jentsch T.J. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). We also that ClC-5 was primarily in of intestinal a of ClC-5 also to be localized to the membrane of the intestinal was reported that this membrane of ClC-5 may be localized in early A. Cluzeaud F. Peng K.C. Bens M. Luchow A. Gunther W. Jentsch T.J. Am. J. Physiol. Cell Physiol. 2001; 280: C373-381Crossref PubMed Google Scholar). Therefore, we of for using the ClC-5 and a of the most early endosomal M. Mol. Biol. Cell. 2000; 11: PubMed Scopus Google The ClC-5 protein to with EEA1 protein in a distinct the apical membrane These provide a for studies of ClC-4 in this tissue. the for of the intestinal is from that for of intestinal crypts were most the membrane of the the of ClC-4 in the apical membrane and/or membrane The apical or membrane of the can be by microscopy and if ClC-4 protein is in early endosomes, we a intestinal were with ClC-4 and the EEA1 These show that the partially that ClC-4 like ClC-5 is localized in early Furthermore, of intestinal crypts with ClC-4 and a marker the lack of and the apical of ClC-4 studies on intestinal tissue that ClC-4 is localized in proximity to the apical membrane of intestinal epithelial The confocal have the to ClC-4 protein is in the apical plasma Therefore, we of mouse and human intestinal tissues with ClC-4 by electron that in mouse ClC-4 is localized primarily the brush border membrane and the apical of an cell from the of the In cells, ClC-4 was in in the apical of a of human with ClC-4 and CFTR that of these proteins primarily on the brush border membrane of human enterocytes ClC-4 was also in the apical and membrane of cells in human tissue The ClC-4 in mouse enterocytes and cells as as in human enterocytes and cells was with the ClC-4 protein the ClC-4 was its as ClC-4 was in the apical membrane of the cells in proximity to CFTR, it was to ClC-4 is functional as a chloride channel in the plasma membrane of We that the Caco-2 cell line is an model for of ClC-4 function in intestinal epithelia, as ClC-4 in Caco-2 is to that in the epithelia. an electron of an of Caco-2 cells for on a using ClC-4 with electron from intestinal we to ClC-4 localized primarily to and brush border was with the ClC-4 protein the ClC-4 was to its confocal microscopy in the of ClC-4 in Caco-2 on a with ClC-4 that the ClC-4 is to the apical membranes of the In Caco-2 cells, ClC-4 the cell surface and intracellular The intracellular and ClC-4 partially with the with EEA1 These that ClC-4 may be between the plasma membrane and the endosomal compartment. this we Caco-2 cells with to and marker to from the plasma membrane to the endosomal its with the S. Physiol. Rev. PubMed Scopus Google Scholar, S. J. Cell Biol. 1993; PubMed Scopus Google Scholar). that the of ClC-4 protein partially with the intracellular of as it the recycling the of ClC-4 and the using this we the of antisense ClC-4 cDNA expression on the by and confocal microscopy of Caco-2 for green protein was with or as a Caco-2 cells to cells We the to the ClC-4 in and in Caco-2 cells We that the of the to the expression of ClC-4 was reduced by in Caco-2 cells to the of the ClC-4 in cells We the of the of by its on ClC-5 and as these proteins and with ClC-4, In cells with ClC-5 was primarily expressed in intracellular membranes with studies in intestinal tissue and Caco-2 cells and reported by Vandewalle al. A. Cluzeaud F. Peng K.C. Bens M. Luchow A. Gunther W. Jentsch T.J. Am. J. Physiol. Cell Physiol. 2001; 280: C373-381Crossref PubMed Google Scholar). In to the of expression on ClC-4 protein we that there was only a on ClC-5 protein expression in to expression of ClC-3, a family was by or was primarily expressed in intracellular membranes there was a at the cell surface in a subpopulation of cells with studies in Caco-2 cells R. Gyomorey K. Rommens J. Choudhury M. Li C. Wang Y. Bear C.E. J. Biol. Chem. 2001; 276: 8306-8313Abstract Full Text Full Text PDF PubMed Scopus (69) Google and reported by al. K. Li Am. J. Physiol. Physiol. 2000; 279: PubMed Google Scholar). that the in Caco-2 cells were to that in and cells Therefore, these show that ClC-4 protein in studies of these intestinal epithelial we the ClC-4 cDNA antisense strategy to if ClC-4 functions endogenously as a chloride channel on the surface membrane of Caco-2 studies in or in T. T. Jentsch T.J. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google showed that of ClC-4 chloride currents were by Therefore, we functionally currents by using intracellular and as the and a T. T. Jentsch T.J. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). from a of the membrane was by from to As in currents of with ClC-4 i.e. with and an were in Caco-2 the membrane of these currents a of These currents to the of chloride in We an antisense strategy to that the currents were by ClC-4 this channel is by of the of chloride channels T. T. Jentsch T.J. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar). the relatively we ClC-4 expression using the R. Gyomorey K. Rommens J. Choudhury M. Li C. Wang Y. Bear C.E. J. Biol. Chem. 2001; 276: 8306-8313Abstract Full Text Full Text PDF PubMed Scopus (69) Google as this of and hence cell to cell in antisense was with the to of We that of Caco-2 cells the endogenous currents The cell at from in cells to in cells, respectively. We the and of the of antisense ClC-4 expression by ClC-4, ClC-5, and protein by of antisense ClC-4 ClC-4 protein The of the to the expression of ClC-4 was reduced by in Caco-2 cells to the of the ClC-4 in cells We the of the of by its on ClC-5 and proteins As in we only a of on ClC-5 protein expression in cells, and of on protein expression in Therefore, these show the of the antisense strategy using the In of chloride currents by ClC-4 antisense expression was a as of antisense ClC-2, a distinct of the ClC chloride channel family (6Jentsch T.J. Friedrich T. Schriever A. Yamada H. Pflugers Arch. 1999; 437: 783-795Crossref PubMed Scopus (297) Google Scholar), these endogenous currents The currents in antisense Caco-2 cells at were from that in cells these that ClC-4 expressed in Caco-2 cells contributes to the endogenous currents in these Jentsch and (6Jentsch T.J. Friedrich T. Schriever A. Yamada H. Pflugers Arch. 1999; 437: 783-795Crossref PubMed Scopus (297) Google in a that the mammalian ClC channel proteins ClC-7 function in intracellular studies of ClC-3, ClC-5 and ClC-7 mice this The primary with of to function and of by these in mice T. S. M. K. H. A. R. Jentsch T.J. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). The primary in mice and to by of proteins from the of the of the kidney S.S. Devuyst O. Courtoy P.J. Wang X.T. Wang H. Wang Y. Thakker R.V. Guggino S. Guggino W.B. Hum. Mol. Genet. 2000; 9: 2937-2945Crossref PubMed Scopus (271) Google Scholar, Gunther W. M. Jentsch T.J. 2000; PubMed Scopus Google Scholar). in mice the of in mutations in Gunther W. S.H. A. M. O. Jentsch T.J. Thakker R.V. Hum. Mol. Genet. PubMed Scopus Google Scholar). of primarily affects secretion across the membrane of the with the and U. E. M. A. Friedrich W. Jentsch T.J. Cell. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). The membrane of the is to membranes with to its these studies show at in the tissues that these members of the ClC family function in intracellular its with and ClC-5, it was that ClC-4 also function in intracellular The present the first of the and function of the ClC-4 channel In to the evidence that at in the intestinal epithelium of and ClC-4 is expressed primarily on the apical brush border membrane it functions to chloride We showed by and confocal microscopy that ClC-4 is primarily localized in proximity to the apical, brush border membrane of intestinal enterocytes and ClC-4 partially with that of that ClC-4 also in apical early ClC-4 can be in the in the is primarily localized in the apical plasma membrane of these Furthermore, we that ClC-4 is functionally expressed in the plasma membrane of intestinal epithelial cells using an antisense strategy with As ClC-4 antisense expression reduced endogenous ClC-4 protein expression in Caco-2 intestinal cells, and this in protein with a in an endogenous chloride we that ClC-4 this As we have the that target ClC-4 to the brush border of the intestinal ClC-4 with ClC-5 at the the of ClC-4 and ClC-5 are quite distinct in intestinal epithelial as in As ClC-4 protein apical and in apical and brush border ClC-5 is the cell and in and endosomal al. M. Friedrich T. Jentsch T.J. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google a on the of ClC-5 is in ClC-4 These showed that a in ClC-5 may be in of ClC-5 channels from the cell surface an with the M. Friedrich T. Jentsch T.J. J. Biol. Chem. 2001; 276: Full Text Full Text PDF PubMed Scopus Google Scholar). was to the surface expression of the epithelial this O. Yeger H. P.J. H. Am. J. Physiol. PubMed Google Scholar, H. J. O. J. 1999; PubMed Scopus Google Scholar). Hence, this may for the of ClC-5 from the cell surface of intestinal epithelial However, there are to be as ClC-4 that a apical of this protein in intestinal epithelia. We it that ClC-4 in chloride secretion by the as it the tissue as CFTR, the chloride channel thought to be primarily for this function in was as a candidate for intestinal disease severity in mice R. M. A. S. M. W. A. J. P. J. Bear C. Genet. PubMed Scopus Google Scholar). In the mouse is on in proximity to a to with a intestinal in However, in another mouse is on the an exception to of to the K. A. Proc. Natl. Acad. Sci. U. S. A. PubMed Scopus Google Scholar). is also on human hence, ClC-4 is considered a candidate for intestinal disease for cystic fibrosis A. A. Hum. Mol. Genet. 1994; PubMed Scopus Google Scholar). However, it that of ClC-4 function as a chloride channel on the brush border membrane may partially compensate for the of CFTR function in the of the of ClC-4 channel is quite it is to its to chloride secretion under and/or conditions. from the of and M. with M. mice exhibit that other chloride CFTR, may a function in the the primary of ClC-4 for phosphorylation by and as there is in evidence for a for of channel function by to a on ClC-5 and ClC-4 channel function and a on ClC-5 there is of on The only of ClC-4 is membrane However, as for the protein ClC-5, of ClC-4 at to as a physiological Jentsch and T. T. Jentsch T.J. J. Biol. Chem. 1999; Full Text Full Text PDF PubMed Scopus Google have for ClC-5 that there be proteins could with these chloride channels and and the of ClC-4 and ClC-5 channel CFTR could provide a as it is localized on the brush border of to ClC-4 and in the of other chloride channels S.S. Guggino W.B. Cell. Full Text PDF PubMed Scopus Google Scholar, L.L. Boucher R.C. M.J. 1993; PubMed Scopus Google Scholar, M. T. S. R. Guggino W.B. PubMed Scopus Google Scholar). studies on the of ClC-4 and proteins are for its function in we have evidence to that ClC-4 may provide a in with CFTR, to mediate the of chloride across the brush border membrane of the epithelium lining intestinal crypts in and Therefore, we that ClC-4 protein may provide a therapeutic target for treatment of intestinal disease in cystic fibrosis. We are to T. Jentsch for of ClC-5 for of and H. P. for of We for

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