Up-regulation of the Homophilic Adhesion Molecule Sidekick-1 in Podocytes Contributes to Glomerulosclerosis
2010; Elsevier BV; Volume: 285; Issue: 33 Linguagem: Inglês
10.1074/jbc.m110.133959
ISSN1083-351X
AutoresLewis Kaufman, Uma Potla, Sarah K. Coleman, Stanislav Dikiy, Yutaka Hata, Hidetake Kurihara, John Cijiang He, Vivette D. D’Agati, Paul E. Klotman,
Tópico(s)Renal and related cancers
ResumoFocal segmental glomerulosclerosis (FSGS) is a leading cause of nephrotic syndrome and end-stage renal disease worldwide. Although the mechanisms underlying this important disease are poorly understood, the glomerular podocyte clearly plays a central role in disease pathogenesis. In the current work, we demonstrate that the homophilic adhesion molecule sidekick-1 (sdk-1) is up-regulated in podocytes in FSGS both in rodent models and in human kidney biopsy samples. Transgenic mice that have podocyte-specific overexpression of sdk-1 develop gradually progressive heavy proteinuria and severe FSGS. We also show that sdk-1 associates with the slit diaphragm linker protein MAGI-1, which is already known to interact with several critical podocyte proteins including synaptopodin, α-actinin-4, nephrin, JAM4, and β-catenin. This interaction is mediated through a direct interaction between the carboxyl terminus of sdk-1 and specific PDZ domains of MAGI-1. In vitro expression of sdk-1 enables a dramatic recruitment of MAGI-1 to the cell membrane. Furthermore, a truncated version of sdk-1 that is unable to bind to MAGI-1 does not induce podocyte dysfunction when overexpressed. We conclude that the up-regulation of sdk-1 in podocytes is an important pathogenic factor in FSGS and that the mechanism involves disruption of the actin cytoskeleton possibly via alterations in MAGI-1 function. Focal segmental glomerulosclerosis (FSGS) is a leading cause of nephrotic syndrome and end-stage renal disease worldwide. Although the mechanisms underlying this important disease are poorly understood, the glomerular podocyte clearly plays a central role in disease pathogenesis. In the current work, we demonstrate that the homophilic adhesion molecule sidekick-1 (sdk-1) is up-regulated in podocytes in FSGS both in rodent models and in human kidney biopsy samples. Transgenic mice that have podocyte-specific overexpression of sdk-1 develop gradually progressive heavy proteinuria and severe FSGS. We also show that sdk-1 associates with the slit diaphragm linker protein MAGI-1, which is already known to interact with several critical podocyte proteins including synaptopodin, α-actinin-4, nephrin, JAM4, and β-catenin. This interaction is mediated through a direct interaction between the carboxyl terminus of sdk-1 and specific PDZ domains of MAGI-1. In vitro expression of sdk-1 enables a dramatic recruitment of MAGI-1 to the cell membrane. Furthermore, a truncated version of sdk-1 that is unable to bind to MAGI-1 does not induce podocyte dysfunction when overexpressed. We conclude that the up-regulation of sdk-1 in podocytes is an important pathogenic factor in FSGS and that the mechanism involves disruption of the actin cytoskeleton possibly via alterations in MAGI-1 function. IntroductionFocal segmental glomerulosclerosis (FSGS) 2The abbreviations used are: FSGSfocal segmental glomerulosclerosisHIVhuman immunodeficiency virusHIVANHIV-associated nephropathyMAGIMAGUK with inverted domain structurePANpuromycin aminonucleosideGSTglutathione S-transferaseMAGUKmembrane-associated guanylate kinase. is an important cause of end-stage renal disease worldwide, accounting for ∼20% of all dialysis patients (1Kitiyakara C. Eggers P. Kopp J.B. Am. J. Kidney Dis. 2004; 44: 815-825Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar). In fact, the frequency of this disease has dramatically increased over the last 20 years, making it the most common cause of primary nephrotic syndrome in adults (1Kitiyakara C. Eggers P. Kopp J.B. Am. J. Kidney Dis. 2004; 44: 815-825Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar). Multiple cohort studies show progression to end-stage renal disease in 50–70% of cases at 10 years, giving FSGS one of the worst prognoses among primary glomerular diseases (2Rydel J.J. Korbet S.M. Borok R.Z. Schwartz M.M. Am. J. Kidney Dis. 1995; 25: 534-542Abstract Full Text PDF PubMed Scopus (239) Google Scholar, 3Korbet S.M. Schwartz M.M. Lewis E.J. Am. J. Kidney Dis. 1994; 23: 773-783Abstract Full Text PDF PubMed Scopus (234) Google Scholar).The diagnosis of FSGS is based on the clinical findings of proteinuria and specific histopathological changes that include glomerular sclerosis, glomerular tuft collapse, and synechia formation. In the early stages, these changes are both focal, affecting a subset of glomeruli, and segmental, involving a portion of the glomerular tuft. Although the idiopathic form of FSGS is the most common, secondary FSGS occurs in association with other underlying conditions including HIV-associated nephropathy (HIVAN), among others.Although the pathogenic mechanisms underlying this disease are poorly understood, the podocyte, the visceral epithelial cell of the glomerulus, plays a central role. Multiple genetic studies using both human and murine models demonstrate that the development of FSGS is initiated by podocyte dysfunction (4Pollak M.R. Semin. Nephrol. 2003; 23: 141-146Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar). In humans, mutations in the podocyte-specific genes nephrin (5Kestilä M. Lenkkeri U. Männikkö M. Lamerdin J. McCready P. Putaala H. Ruotsalainen V. Morita T. Nissinen M. Herva R. Kashtan C.E. Peltonen L. Holmberg C. Olsen A. Tryggvason K. Mol. Cell. 1998; 1: 575-582Abstract Full Text Full Text PDF PubMed Scopus (1554) Google Scholar), podocin (6Boute N. Gribouval O. Roselli S. Benessy F. Lee H. Fuchshuber A. Dahan K. Gubler M.C. Niaudet P. Antignac C. Nat. Genet. 2000; 24: 349-354Crossref PubMed Scopus (1186) Google Scholar), α-actinin-4 (7Kaplan J.M. Kim S.H. North K.N. Rennke H. Correia L.A. Tong H.Q. Mathis B.J. Rodríguez-Pérez J.C. Allen P.G. Beggs A.H. Pollak M.R. Nat. Genet. 2000; 24: 251-256Crossref PubMed Scopus (1033) Google Scholar), TRPC6 (8Winn M.P. Conlon P.J. Lynn K.L. Farrington M.K. Creazzo T. Hawkins A.F. Daskalakis N. Kwan S.Y. Ebersviller S. Burchette J.L. Pericak-Vance M.A. Howell D.N. Vance J.M. Rosenberg P.B. Science. 2005; 308: 1801-1804Crossref PubMed Scopus (874) Google Scholar, 9Reiser J. Polu K.R. Möller C.C. Kenlan P. Altintas M.M. Wei C. Faul C. Herbert S. Villegas I. Avila-Casado C. McGee M. Sugimoto H. Brown D. Kalluri R. Mundel P. Smith P.L. Clapham D.E. Pollak M.R. Nat. Genet. 2005; 37: 739-744Crossref PubMed Scopus (674) Google Scholar), and others all disrupt podocyte function, leading to inherited forms of FSGS. Two recent landmark studies showed a strong association of non-coding variants in the podocyte-expressed gene myh9 with susceptibility to HIVAN and FSGS (10Kopp J.B. Smith M.W. Nelson G.W. Johnson R.C. Freedman B.I. Bowden D.W. Oleksyk T. McKenzie L.M. Kajiyama H. Ahuja T.S. Berns J.S. Briggs W. Cho M.E. Dart R.A. Kimmel P.L. Korbet S.M. Michel D.M. Mokrzycki M.H. Schelling J.R. Simon E. Trachtman H. Vlahov D. Winkler C.A. Nat. Genet. 2008; 40: 1175-1184Crossref PubMed Scopus (582) Google Scholar, 11Kao W.H. Klag M.J. Meoni L.A. Reich D. Berthier-Schaad Y. Li M. Coresh J. Patterson N. Tandon A. Powe N.R. Fink N.E. Sadler J.H. Weir M.R. Abboud H.E. Adler S.G. Divers J. Iyengar S.K. Freedman B.I. Kimmel P.L. Knowler W.C. Kohn O.F. Kramp K. Leehey D.J. Nicholas S.B. Pahl M.V. Schelling J.R. Sedor J.R. Thornley-Brown D. Winkler C.A. Smith M.W. Parekh R.S. Nat. Genet. 2008; 40: 1185-1192Crossref PubMed Scopus (532) Google Scholar), suggesting an important connection with the genetic predisposition of African-Americans to developing nephropathy and confirming the central role of podocyte genetics to many forms of renal failure.Sidekick was first described in Drosophila melanogaster as a critical determinant of cell fate in retinal photoreceptors (12Nguyen D.N. Liu Y. Litsky M.L. Reinke R. Development. 1997; 124: 3303-3312Crossref PubMed Google Scholar). Later, sidekick-1 (sdk-1) and its ortholog sidekick-2 (sdk-2) were found to localize to the neurological synapse and to function as guidance molecules targeting synapses to specific layers (13Yamagata M. Weiner J.A. Sanes J.R. Cell. 2002; 110: 649-660Abstract Full Text Full Text PDF PubMed Scopus (280) Google Scholar, 14Yamagata M. Sanes J.R. Nature. 2008; 451: 465-469Crossref PubMed Scopus (312) Google Scholar). sdk-1 and sdk-2 each consist of a large extracellular domain containing six Ig motifs followed by 13 fibronectin type III repeats, a single transmembrane domain, and a short cytoplasmic tail containing a highly conserved PDZ binding domain at its carboxyl terminus. sdk-1 and sdk-2 function as homophilic adhesion molecules such that cells expressing sdk-1 or sdk-2 strongly prefer to interact exclusively with cells expressing the same sidekick isoform (13Yamagata M. Weiner J.A. Sanes J.R. Cell. 2002; 110: 649-660Abstract Full Text Full Text PDF PubMed Scopus (280) Google Scholar, 15Hayashi K. Kaufman L. Ross M.D. Klotman P.E. FASEB J. 2005; 19: 614-616Crossref PubMed Scopus (19) Google Scholar).We identified sdk-1 as being massively overexpressed in podocytes in HIVAN (16Kaufman L. Hayashi K. Ross M.J. Ross M.D. Klotman P.E. J. Am. Soc. Nephrol. 2004; 15: 1721-1730Crossref PubMed Scopus (36) Google Scholar, 17Kaufman L. Yang G. Hayashi K. Ashby J.R. Huang L. Ross M.J. Klotman M.E. Klotman P.E. FASEB J. 2007; 21: 1367-1375Crossref PubMed Scopus (27) Google Scholar) and reported that this up-regulation was a maladaptive response resulting in increased intercellular adhesion and loss of cytoskeletal integrity (17Kaufman L. Yang G. Hayashi K. Ashby J.R. Huang L. Ross M.J. Klotman M.E. Klotman P.E. FASEB J. 2007; 21: 1367-1375Crossref PubMed Scopus (27) Google Scholar). In the current work, we first demonstrate that sdk-1 up-regulation in podocytes also occurs in idiopathic FSGS. We then validate the physiological relevance of this up-regulation by generating novel transgenic mice that develop severe FSGS when sdk-1 is expressed specifically in podocytes. Next, we begin to dissect the mechanisms by which sdk-1 up-regulation contributes to disease. We show that sdk-1 directly interacts with the podocyte linker protein MAGI-1 and that this interaction is necessary for sdk-1 overexpression to induce podocyte dysfunction. MAGUK with inverted domain structure-1 (MAGI-1) is localized specifically at the slit diaphragm in podocytes and is composed of one guanylate kinase, two WW domains, and six PDZ domains (18Dobrosotskaya I. Guy R.K. James G.L. J. Biol. Chem. 1997; 272: 31589-31597Abstract Full Text Full Text PDF PubMed Scopus (172) Google Scholar, 19Ide N. Hata Y. Nishioka H. Hirao K. Yao I. Deguchi M. Mizoguchi A. Nishimori H. Tokino T. Nakamura Y. Takai Y. Oncogene. 1999; 18: 7810-7815Crossref PubMed Scopus (106) Google Scholar). MAGI-1 is a linker protein that directly interacts with other critical podocyte proteins including α-actinin-4, synaptopodin, nephrin, JAM4, and β-catenin and is linked to the actin cytoskeleton through these proteins (20Hirabayashi S. Tajima M. Yao I. Nishimura W. Mori H. Hata Y. Mol. Cell. Biol. 2003; 23: 4267-4282Crossref PubMed Scopus (149) Google Scholar, 21Patrie K.M. Drescher A.J. Welihinda A. Mundel P. Margolis B. J. Biol. Chem. 2002; 277: 30183-30190Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar, 22Hirabayashi S. Mori H. Kansaku A. Kurihara H. Sakai T. Shimizu F. Kawachi H. Hata Y. Lab. Invest. 2005; 85: 1528-1543Crossref PubMed Scopus (55) Google Scholar, 23Sakurai A. Fukuhara S. Yamagishi A. Sako K. Kamioka Y. Masuda M. Nakaoka Y. Mochizuki N. Mol. Biol. Cell. 2006; 17: 966-976Crossref PubMed Scopus (127) Google Scholar). We conclude that the up-regulation of sdk-1 in podocytes is an important pathogenic factor in the development of FSGS and that the mechanism involves alterations in the actin cytoskeleton possibly mediated by changes in MAGI-1 scaffolding.DISCUSSIONIn the current work, we have identified a novel mouse model of chronic FSGS mediated by a unique cellular mechanism that is relevant to human disease. De novo expression of sdk-1 in podocytes was sufficient to induce slowly progressive glomerulosclerosis in transgenic mice. Affected animals develop relatively late onset proteinuria, without renal failure, that does not begin until well into adulthood and then progresses over many months. This is in stark contrast to existing models of FSGS that typically involve early onset proteinuria with rapid progression to kidney failure and death. These rapidly progressive FSGS models include knock-out models (CD2AP (27Shih N.Y. Li J. Karpitskii V. Nguyen A. Dustin M.L. Kanagawa O. Miner J.H. Shaw A.S. Science. 1999; 286: 312-315Crossref PubMed Scopus (693) Google Scholar), neph1 (28Donoviel D.B. Freed D.D. Vogel H. Potter D.G. Hawkins E. Barrish J.P. Mathur B.N. Turner C.A. Geske R. Montgomery C.A. Starbuck M. Brandt M. Gupta A. Ramirez-Solis R. Zambrowicz B.P. Powell D.R. Mol. Cell. Biol. 2001; 21: 4829-4836Crossref PubMed Scopus (346) Google Scholar), α-actinin-4 (29Kos C.H. Le T.C. Sinha S. Henderson J.M. Kim S.H. Sugimoto H. Kalluri R. Gerszten R.E. Pollak M.R. J. Clin. Invest. 2003; 111: 1683-1690Crossref PubMed Scopus (210) Google Scholar), podocin (30Roselli S. Heidet L. Sich M. Henger A. Kretzler M. Gubler M.C. Antignac C. Mol. Cell. Biol. 2004; 24: 550-560Crossref PubMed Scopus (222) Google Scholar), nephrin (31Putaala H. Soininen R. Kilpeläinen P. Wartiovaara J. Tryggvason K. Hum. Mol. Genet. 2001; 10: 1-8Crossref PubMed Scopus (419) Google Scholar), and others) and podocyte-specific overexpression models such as Notch-1 (32Niranjan T. Bielesz B. Gruenwald A. Ponda M.P. Kopp J.B. Thomas D.B. Susztak K. Nat. Med. 2008; 14: 290-298Crossref PubMed Scopus (319) Google Scholar, 33Waters A.M. Wu M.Y. Onay T. Scutaru J. Liu J. Lobe C.G. Quaggin S.E. Piscione T.D. J. Am. Soc. Nephrol. 2008; 19: 1139-1157Crossref PubMed Scopus (98) Google Scholar). Overexpression of sdk-1 in podocytes also induced changes that are typical for the collapsing variant of FSGS including podocyte hyperplasia, hypertrophy, pseudocrescent formation, and expression of the cell cycle marker Ki-67. Several other mouse models of collapsing glomerulopathy have been described including: α-actinin-4 knock-in and knock-out models (34Henderson J.M. Al-Waheeb S. Weins A. Dandapani S.V. Pollak M.R. Kidney Int. 2008; 73: 741-750Abstract Full Text Full Text PDF PubMed Scopus (28) Google Scholar), transgenic mice with podocyte-specific overexpression of vascular endothelial growth factor (35Eremina V. Sood M. Haigh J. Nagy A. Lajoie G. Ferrara N. Gerber H.P. Kikkawa Y. Miner J.H. Quaggin S.E. J. Clin. Invest. 2003; 111: 707-716Crossref PubMed Scopus (1061) Google Scholar), alterations in mitochondrial function (36Barisoni L. Madaio M.P. Eraso M. Gasser D.L. Nelson P.J. J. Am. Soc. Nephrol. 2005; 16: 2847-2851Crossref PubMed Scopus (47) Google Scholar), and expression of HIV genes (37Kopp J.B. Klotman M.E. Adler S.H. Bruggeman L.A. Dickie P. Marinos N.J. Eckhaus M. Bryant J.L. Notkins A.L. Klotman P.E. Proc. Natl. Acad. Sci. U.S.A. 1992; 89: 1577-1581Crossref PubMed Scopus (277) Google Scholar). Again, these other FSGS models are characterized by relatively rapid disease progression. The late onset and more gradual disease progression evident in the sdk-1 transgenic model are more consistent with the usual clinical course of human FSGS patients. We propose our model to be one of the first genetic models of chronic, slowly progressive FSGS.We do note significant differences in the relative amounts of observed glomerulosclerosis between animals and some differences in age of onset of proteinuria and rapidity of progression; this is likely explained by observed differences in expression levels of sdk-1 both between animals and even in different glomeruli within a given animal. The reason for these differences is uncertain but seems to be related to variable expression of cre recombinase between glomeruli.To investigate the mechanisms by which sdk-1 up-regulation in the podocyte contributes to FSGS pathogenesis, we began to investigate how sdk-1 interacts with other important podocyte proteins. The cytoplasmic domains of sdk-1 and sdk-2 contain no clear signaling domains other than a highly conserved carboxyl-terminal PDZ binding domain. In fact, the six terminal amino acids (GFSSFV) are completely conserved in all sidekicks in all available species from Caenorhabditis elegans to humans, suggesting that binding to this conserved domain is critical to sidekick function. In this way, we identified the slit diaphragm linker protein MAGI-1 as binding specifically to this conserved motif.The MAGI proteins, with their multiple protein-protein interacting domains, have a large number of known binding partners. In podocytes, for example, MAGI-1 is known to interact with synaptopodin via its WW domain (21Patrie K.M. Drescher A.J. Welihinda A. Mundel P. Margolis B. J. Biol. Chem. 2002; 277: 30183-30190Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar), with α-actinin-4 via PDZ domain 5 (21Patrie K.M. Drescher A.J. Welihinda A. Mundel P. Margolis B. J. Biol. Chem. 2002; 277: 30183-30190Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar), with nephrin via PDZ domains 2 and 3 (22Hirabayashi S. Mori H. Kansaku A. Kurihara H. Sakai T. Shimizu F. Kawachi H. Hata Y. Lab. Invest. 2005; 85: 1528-1543Crossref PubMed Scopus (55) Google Scholar),with JAM4 via PDZ domains 1 and 4 (20Hirabayashi S. Tajima M. Yao I. Nishimura W. Mori H. Hata Y. Mol. Cell. Biol. 2003; 23: 4267-4282Crossref PubMed Scopus (149) Google Scholar), and with β-catenin via PDZ domain 5 (38Dobrosotskaya I.Y. James G.L. Biochem. Biophys. Res. Commun. 2000; 270: 903-909Crossref PubMed Scopus (138) Google Scholar). Presumably, MAGI-1 functions to stabilize and organize this complex of proteins at the slit diaphragm and to link them to the actin cytoskeleton. Because the phenotype of MAGI-1 null mice has not yet been reported, the importance of MAGI-1 as a stabilizing or signaling molecule in the podocyte has not been fully established. Furthermore, at this time, no human familial nephrotic syndrome cases have been linked to mutations in MAGI-1.In this work, we show that the interaction between sdk-1 and MAGI-1 is promiscuous in that three different PDZ domains of MAGI-1 are each capable of binding to the PDZ binding domain of sdk-1. Expression of sdk-1 was also able to change the subcellular localization of MAGI-1 by inducing dramatic recruitment to the cell membrane. Furthermore, a truncated version of sdk-1 that lacks its terminal four amino acids is unable to interact with MAGI-1 and does not induce podocyte dysfunction when overexpressed. These data imply, but do not definitively prove, that the interaction between the two proteins is important for induction of disease. It is possible that sdk-1 interacts with other molecules besides MAGI-1 and that these other interactions are also important in inducing podocyte dysfunction. Ongoing work is focused on clarifying these possibilities.The interaction between sdk-1 and MAGI-1 may also be important to the process of neuronal synapse formation. In brain development, both proteins are highly expressed specifically at neurological synapses, and both affect the processes of synaptic connectivity and neuronal migration (13Yamagata M. Weiner J.A. Sanes J.R. Cell. 2002; 110: 649-660Abstract Full Text Full Text PDF PubMed Scopus (280) Google Scholar, 14Yamagata M. Sanes J.R. Nature. 2008; 451: 465-469Crossref PubMed Scopus (312) Google Scholar, 26Wright G.J. Leslie J.D. Ariza-McNaughton L. Lewis J. Development. 2004; 131: 5659-5669Crossref PubMed Scopus (46) Google Scholar, 39Emtage L. Chang H. Tiver R. Rongo C. PLoS ONE. 2009; 4: e4613Crossref PubMed Scopus (32) Google Scholar, 40Mizuhara E. Nakatani T. Minaki Y. Sakamoto Y. Ono Y. Takai Y. J. Biol. Chem. 2005; 280: 26499-26507Abstract Full Text Full Text PDF PubMed Scopus (55) Google Scholar). sdk-2 is also important in this process, and because its PDZ binding domain is identical to that of sdk-1, it is also likely to interact with MAGI-1, although this has not been shown.Collectively, our data identify a novel mouse model of chronic FSGS that implicates the sdk-1 genetic pathway as important to the pathogenesis of glomerular sclerosis. These findings suggest that the up-regulation of sdk-1 by podocytes in FSGS is a maladaptive response to injury that is independently capable of leading to progressive podocyte dysfunction and ultimately to glomerular sclerosis. IntroductionFocal segmental glomerulosclerosis (FSGS) 2The abbreviations used are: FSGSfocal segmental glomerulosclerosisHIVhuman immunodeficiency virusHIVANHIV-associated nephropathyMAGIMAGUK with inverted domain structurePANpuromycin aminonucleosideGSTglutathione S-transferaseMAGUKmembrane-associated guanylate kinase. is an important cause of end-stage renal disease worldwide, accounting for ∼20% of all dialysis patients (1Kitiyakara C. Eggers P. Kopp J.B. Am. J. Kidney Dis. 2004; 44: 815-825Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar). In fact, the frequency of this disease has dramatically increased over the last 20 years, making it the most common cause of primary nephrotic syndrome in adults (1Kitiyakara C. Eggers P. Kopp J.B. Am. J. Kidney Dis. 2004; 44: 815-825Abstract Full Text Full Text PDF PubMed Scopus (224) Google Scholar). Multiple cohort studies show progression to end-stage renal disease in 50–70% of cases at 10 years, giving FSGS one of the worst prognoses among primary glomerular diseases (2Rydel J.J. Korbet S.M. Borok R.Z. Schwartz M.M. Am. J. Kidney Dis. 1995; 25: 534-542Abstract Full Text PDF PubMed Scopus (239) Google Scholar, 3Korbet S.M. Schwartz M.M. Lewis E.J. Am. J. Kidney Dis. 1994; 23: 773-783Abstract Full Text PDF PubMed Scopus (234) Google Scholar).The diagnosis of FSGS is based on the clinical findings of proteinuria and specific histopathological changes that include glomerular sclerosis, glomerular tuft collapse, and synechia formation. In the early stages, these changes are both focal, affecting a subset of glomeruli, and segmental, involving a portion of the glomerular tuft. Although the idiopathic form of FSGS is the most common, secondary FSGS occurs in association with other underlying conditions including HIV-associated nephropathy (HIVAN), among others.Although the pathogenic mechanisms underlying this disease are poorly understood, the podocyte, the visceral epithelial cell of the glomerulus, plays a central role. Multiple genetic studies using both human and murine models demonstrate that the development of FSGS is initiated by podocyte dysfunction (4Pollak M.R. Semin. Nephrol. 2003; 23: 141-146Abstract Full Text Full Text PDF PubMed Scopus (67) Google Scholar). In humans, mutations in the podocyte-specific genes nephrin (5Kestilä M. Lenkkeri U. Männikkö M. Lamerdin J. McCready P. Putaala H. Ruotsalainen V. Morita T. Nissinen M. Herva R. Kashtan C.E. Peltonen L. Holmberg C. Olsen A. Tryggvason K. Mol. Cell. 1998; 1: 575-582Abstract Full Text Full Text PDF PubMed Scopus (1554) Google Scholar), podocin (6Boute N. Gribouval O. Roselli S. Benessy F. Lee H. Fuchshuber A. Dahan K. Gubler M.C. Niaudet P. Antignac C. Nat. Genet. 2000; 24: 349-354Crossref PubMed Scopus (1186) Google Scholar), α-actinin-4 (7Kaplan J.M. Kim S.H. North K.N. Rennke H. Correia L.A. Tong H.Q. Mathis B.J. Rodríguez-Pérez J.C. Allen P.G. Beggs A.H. Pollak M.R. Nat. Genet. 2000; 24: 251-256Crossref PubMed Scopus (1033) Google Scholar), TRPC6 (8Winn M.P. Conlon P.J. Lynn K.L. Farrington M.K. Creazzo T. Hawkins A.F. Daskalakis N. Kwan S.Y. Ebersviller S. Burchette J.L. Pericak-Vance M.A. Howell D.N. Vance J.M. Rosenberg P.B. Science. 2005; 308: 1801-1804Crossref PubMed Scopus (874) Google Scholar, 9Reiser J. Polu K.R. Möller C.C. Kenlan P. Altintas M.M. Wei C. Faul C. Herbert S. Villegas I. Avila-Casado C. McGee M. Sugimoto H. Brown D. Kalluri R. Mundel P. Smith P.L. Clapham D.E. Pollak M.R. Nat. Genet. 2005; 37: 739-744Crossref PubMed Scopus (674) Google Scholar), and others all disrupt podocyte function, leading to inherited forms of FSGS. Two recent landmark studies showed a strong association of non-coding variants in the podocyte-expressed gene myh9 with susceptibility to HIVAN and FSGS (10Kopp J.B. Smith M.W. Nelson G.W. Johnson R.C. Freedman B.I. Bowden D.W. Oleksyk T. McKenzie L.M. Kajiyama H. Ahuja T.S. Berns J.S. Briggs W. Cho M.E. Dart R.A. Kimmel P.L. Korbet S.M. Michel D.M. Mokrzycki M.H. Schelling J.R. Simon E. Trachtman H. Vlahov D. Winkler C.A. Nat. Genet. 2008; 40: 1175-1184Crossref PubMed Scopus (582) Google Scholar, 11Kao W.H. Klag M.J. Meoni L.A. Reich D. Berthier-Schaad Y. Li M. Coresh J. Patterson N. Tandon A. Powe N.R. Fink N.E. Sadler J.H. Weir M.R. Abboud H.E. Adler S.G. Divers J. Iyengar S.K. Freedman B.I. Kimmel P.L. Knowler W.C. Kohn O.F. Kramp K. Leehey D.J. Nicholas S.B. Pahl M.V. Schelling J.R. Sedor J.R. Thornley-Brown D. Winkler C.A. Smith M.W. Parekh R.S. Nat. Genet. 2008; 40: 1185-1192Crossref PubMed Scopus (532) Google Scholar), suggesting an important connection with the genetic predisposition of African-Americans to developing nephropathy and confirming the central role of podocyte genetics to many forms of renal failure.Sidekick was first described in Drosophila melanogaster as a critical determinant of cell fate in retinal photoreceptors (12Nguyen D.N. Liu Y. Litsky M.L. Reinke R. Development. 1997; 124: 3303-3312Crossref PubMed Google Scholar). Later, sidekick-1 (sdk-1) and its ortholog sidekick-2 (sdk-2) were found to localize to the neurological synapse and to function as guidance molecules targeting synapses to specific layers (13Yamagata M. Weiner J.A. Sanes J.R. Cell. 2002; 110: 649-660Abstract Full Text Full Text PDF PubMed Scopus (280) Google Scholar, 14Yamagata M. Sanes J.R. Nature. 2008; 451: 465-469Crossref PubMed Scopus (312) Google Scholar). sdk-1 and sdk-2 each consist of a large extracellular domain containing six Ig motifs followed by 13 fibronectin type III repeats, a single transmembrane domain, and a short cytoplasmic tail containing a highly conserved PDZ binding domain at its carboxyl terminus. sdk-1 and sdk-2 function as homophilic adhesion molecules such that cells expressing sdk-1 or sdk-2 strongly prefer to interact exclusively with cells expressing the same sidekick isoform (13Yamagata M. Weiner J.A. Sanes J.R. Cell. 2002; 110: 649-660Abstract Full Text Full Text PDF PubMed Scopus (280) Google Scholar, 15Hayashi K. Kaufman L. Ross M.D. Klotman P.E. FASEB J. 2005; 19: 614-616Crossref PubMed Scopus (19) Google Scholar).We identified sdk-1 as being massively overexpressed in podocytes in HIVAN (16Kaufman L. Hayashi K. Ross M.J. Ross M.D. Klotman P.E. J. Am. Soc. Nephrol. 2004; 15: 1721-1730Crossref PubMed Scopus (36) Google Scholar, 17Kaufman L. Yang G. Hayashi K. Ashby J.R. Huang L. Ross M.J. Klotman M.E. Klotman P.E. FASEB J. 2007; 21: 1367-1375Crossref PubMed Scopus (27) Google Scholar) and reported that this up-regulation was a maladaptive response resulting in increased intercellular adhesion and loss of cytoskeletal integrity (17Kaufman L. Yang G. Hayashi K. Ashby J.R. Huang L. Ross M.J. Klotman M.E. Klotman P.E. FASEB J. 2007; 21: 1367-1375Crossref PubMed Scopus (27) Google Scholar). In the current work, we first demonstrate that sdk-1 up-regulation in podocytes also occurs in idiopathic FSGS. We then validate the physiological relevance of this up-regulation by generating novel transgenic mice that develop severe FSGS when sdk-1 is expressed specifically in podocytes. Next, we begin to dissect the mechanisms by which sdk-1 up-regulation contributes to disease. We show that sdk-1 directly interacts with the podocyte linker protein MAGI-1 and that this interaction is necessary for sdk-1 overexpression to induce podocyte dysfunction. MAGUK with inverted domain structure-1 (MAGI-1) is localized specifically at the slit diaphragm in podocytes and is composed of one guanylate kinase, two WW domains, and six PDZ domains (18Dobrosotskaya I. Guy R.K. James G.L. J. Biol. Chem. 1997; 272: 31589-31597Abstract Full Text Full Text PDF PubMed Scopus (172) Google Scholar, 19Ide N. Hata Y. Nishioka H. Hirao K. Yao I. Deguchi M. Mizoguchi A. Nishimori H. Tokino T. Nakamura Y. Takai Y. Oncogene. 1999; 18: 7810-7815Crossref PubMed Scopus (106) Google Scholar). MAGI-1 is a linker protein that directly interacts with other critical podocyte proteins including α-actinin-4, synaptopodin, nephrin, JAM4, and β-catenin and is linked to the actin cytoskeleton through these proteins (20Hirabayashi S. Tajima M. Yao I. Nishimura W. Mori H. Hata Y. Mol. Cell. Biol. 2003; 23: 4267-4282Crossref PubMed Scopus (149) Google Scholar, 21Patrie K.M. Drescher A.J. Welihinda A. Mundel P. Margolis B. J. Biol. Chem. 2002; 277: 30183-30190Abstract Full Text Full Text PDF PubMed Scopus (113) Google Scholar, 22Hirabayashi S. Mori H. Kansaku A. Kurihara H. Sakai T. Shimizu F. Kawachi H. Hata Y. Lab. Invest. 2005; 85: 1528-1543Crossref PubMed Scopus (55) Google Scholar, 23Sakurai A. Fukuhara S. Yamagishi A. Sako K. Kamioka Y. Masuda M. Nakaoka Y. Mochizuki N. Mol. Biol. Cell. 2006; 17: 966-976Crossref PubMed Scopus (127) Google Scholar). We conclude that the up-regulation of sdk-1 in podocytes is an important pathogenic factor in the development of FSGS and that the mechanism involves alterations in the actin cytoskeleton possibly mediated by changes in MAGI-1 scaffolding.
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