From the Renal Division, University Hospital
Freiburg, Hugstetter Str. 55, 79106 Freiburg, Germany and
§ University Hospital, Hölkeskampring 40, 44625 Herne,
Germany
Received for publication, December 6, 2002, and in revised form, February 10, 2003
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ABSTRACT |
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The PSD95/Dlg/ZO-1 (PDZ) domain-containing
protein zonula occludens-1 (ZO-1) selectively localizes to the
cytoplasmic basis of the slit diaphragm, a specialized cell-cell
contact in between glomerular podocytes necessary to prevent the loss
of protein in the urine. However, the function of ZO-1 at the slit
diaphragm has remained elusive. Deletion of Neph1, a slit diaphragm
protein of the immunoglobulin superfamily with a cytoplasmic PDZ
binding site, causes proteinuria in mice. We demonstrate now that Neph1 binds ZO-1. This interaction was mediated by the first PDZ domain of
ZO-1 and involved the conserved PDZ domain binding motif present in the
carboxyl terminus of the three known Neph family members. Furthermore,
Neph1 co-immunoprecipitates with ZO-1 from lysates of mouse kidneys,
demonstrating that this interaction occurs in vivo. Both
deletion of the PDZ binding motif of Neph1 as well as
threonine-to-glutamate mutation of the threonine within the binding
motif abrogated binding of ZO-1, suggesting that phosphorylation may
regulate this interaction. ZO-1 binding was associated with a strong
increase in tyrosine phosphorylation of the cytoplasmic tail of Neph1
and dramatically accelerated the ability of Neph1 to induce signal
transduction. Thus, our data suggest that ZO-1 may organize Neph
proteins and recruit signal transduction components to the slit
diaphragm of podocytes.
Hereditary nephrotic syndrome is a heterogeneous disease
characterized by heavy proteinuria and renal failure. The recent description of gene defects of the podocyte resulting in hereditary nephrotic syndrome has provided a completely new understanding of the
glomerular filter and unraveled important aspects of the pathogenesis
of proteinuric kidney diseases (1). The most severe hereditary disorder
is the congenital nephrotic syndrome of the Finnish type, caused by
mutations in
NPHS1,1 the gene
encoding for nephrin. Nephrin is an integral membrane protein of the
immunoglobulin superfamily located at opposing sites of the secondary
foot processes formed by podocytes, specialized epithelial cells that
ensure size- and charge-selective ultrafiltration (for reviews, see
Refs. 2 and 3). The precise function of nephrin is unknown; however,
nephrin is a critical structural component of the slit diaphragm, an
ultrathin zipper-like structure that bridges the ~40-nm-wide slit
between interdigitating podocyte foot processes. We have recently shown
that nephrin is a signaling protein and that signal transduction can be
augmented by another podocyte protein called podocin (4). Podocin is a
hairpin-like protein at the slit diaphragm encoded by NPHS2,
the gene disrupted in a steroid-resistant hereditary nephrotic syndrome
(5, 6). In addition to nephrin and podocin, other podocyte proteins
including CD2AP (7, 8), Zonula occludens (ZO) proteins are membrane-associated multidomain
proteins usually localized at sites of intercellular junctions. They
contain three PDZ domains, a SH3 domain, and a guanylate kinase
domain (14). PDZ domains are protein-binding modules that recognize
short peptide motifs within their protein targets (15). In almost all
cases the last three to five residues at the extreme carboxyl terminus
of a transmembrane protein represent the target sequences. Genetic
evidence from invertebrate systems demonstrates a role for ZO proteins
in facilitating signal transduction, and evidence from vertebrate
systems demonstrates a structural role in organizing transmembrane
protein complexes (16, 17). In podocytes, ZO-1 has been shown to
specifically localize to the cytoplasmic surface of the slit diaphragms
(18-20). However, the function of ZO-1 at the slit and its binding
partners have not been characterized.
We demonstrate now that ZO-1 specifically binds to the carboxyl
terminus of Neph1. Neph1 and ZO-1 are co-localized in the kidney
glomerulus. Interaction of ZO-1 with Neph1 alters the ability of Neph1
to stimulate signaling. Our findings suggest that the multiadapter
protein ZO-1 serves as a scaffold for the organization of Neph1
molecules and/or provides a platform for the recruitment of signal
transduction components to the dynamic protein complex at the slit.
Reagents and Plasmids--
Neph1, Neph2, and Neph3 have been
described previously (12). Membrane-bound fusion proteins of the
carboxyl-terminal cytoplasmic domain of Neph1 were generated using a
pCDM8 cassette that contained the leader sequence of CD5 fused to the
CH2 and CH3 domain of human IgG1 followed by the transmembrane region
of CD7 (21). A full-length cDNA clone of ZO-1 was kindly provided
by Dr. Anderson (Yale University School of Medicine, New Haven, CT).
Truncations and mutations of ZO-1 and Neph1 were generated by standard
cloning procedures. The Neph1 antiserum has been described (12);
antibodies were obtained from Sigma (anti-FLAG M2), Santa Cruz
Biotechnology, Inc. (anti-ZO-1 and anti-HA), Roche Molecular
Biochemicals (anti-HA), and Upstate Biotechnology (anti-Tyr(P) 4G10).
Co-immunoprecipitation--
Co-immunoprecipitations were
performed as described previously (22). Briefly, HEK 293T cells were
transiently transfected by the calcium phosphate method. After
incubation for 24 h, cells were washed twice and lysed in a 1%
Triton X-100 lysis buffer. After centrifugation (15,000 × g, 15 min, 4 °C) and ultracentrifugation (100,000 × g, 30 min, 4 °C) cell lysates containing equal amounts of
total protein were incubated for 1 h at 4 °C with the
appropriate antibody followed by incubation with 40 µl of protein
G-Sepharose beads for ~3 h. The beads were washed extensively with
lysis buffer, and bound proteins were resolved by 10% SDS-PAGE. Since
native Neph1 and ZO-1 in kidney cortex may be associated with lipid
rafts, the lysis buffer was supplemented with 20 mM CHAPS.
Sufficient solubilization was monitored by Western blot of different
fractions during the preparation. Before immunoprecipitation cellular
lysates were extensively precleared by ultracentrifugation and
absorption to protein G beads. All kidneys were freshly prepared and
perfused in situ with ice-cold phosphate-buffered saline
before lysis. Densitometric analysis was performed on non-saturated
radiographs using the NIH Image software package.
32P Labeling and Phosphoamino Acid
Analysis--
After labeling of transfected HEK 293T cells with
32P (0.5 mCi/ml for 6 h), Neph1 was immunoprecipitated
with protein G. Immunoprecipitates were washed extensively, subjected
to 10% SDS-PAGE, and transferred onto polyvinylidene difluoride
membranes. After autoradiography membrane pieces containing the
32P-labeled Neph1 were cut out and subjected to
phosphoamino acid analysis.
Pull-down Assay--
HEK 293T cells were transiently transfected
with plasmid DNA as indicated. Cells were lysed in 1% Triton X-100, 20 mM Tris-HCl, pH 7.5, 50 mM NaCl, 50 mM NaF, 15 mM
Na4P2O7, 2 mM
Na3VO4, 1 mM EDTA, and protease
inhibitors for 15 min on ice. Following centrifugation, the supernatant
was incubated for 1 h at 4 °C with 4-8 µg of recombinant purified glutathione S-transferase (GST) or GST·PDZ domain
fusion protein prebound to glutathione-Sepharose beads (Amersham
Biosciences). Bound proteins were separated by 10% SDS-PAGE, and
precipitated proteins were visualized with anti-FLAG antibody. Equal
loading of recombinant proteins was confirmed by Coomassie Blue
staining of the gels.
Immunofluorescence--
Frozen adult mouse kidneys were embedded
in OCT, sectioned at 5 µm, and fixed with ice-cold acetone.
The sections were incubated with affinity-purified anti-Neph1 antiserum
followed by anti-rabbit rhodamine red and anti-ZO-1 rat monoclonal
antibody (Chemicon, Hofheim, Germany) followed by anti-rat fluorescein
isothiocyanate (DAKO, Hamburg, Germany). An Axiophot 2 microscope
(Zeiss, Jena) equipped with a CCD camera was used for image
documentation. Confocal images were taken using a Zeiss laser scan
microscope equipped with a 100× oil immersion objective.
Luciferase Assay--
HEK 293T cells seeded in 12-well plates
were transiently transfected with a luciferase reporter construct, a
Recent genetic studies of human hereditary disease and genetically
modified animals have led to the identification of new podocyte
proteins and have highlighted the crucial role of proteins at the
filtration slit for the integrity of the glomerular filtration barrier
(23). However, it is unknown how these different protein components of
the slit diaphragm are organized. Since the carboxyl-terminal tail of
Neph1 contains a putative class 1 PDZ domain binding motif (scansite.mit.edu, (24)), we speculated that this motif could mediate interaction with PDZ domain proteins. ZO-1 has been the only
PDZ domain protein demonstrated to localize to the cytoplasmic surface
of the filtration slit. We therefore tested whether ZO-1 interacts with
Neph1. Fig. 1A shows that
Neph1 specifically co-immunoprecipitated with ZO-1. No interaction
could be demonstrated for FAT, another slit diaphragm
transmembrane protein containing a putative PDZ domain binding motif
(Fig. 1E) (9). Binding of Neph1 could be localized to the
first 503 amino acids of ZO-1, a region containing the three PDZ
domains of ZO-1 (Fig. 1, B and D) and the
cytoplasmic tail of Neph1 (Fig. 1C). FLAG-tagged
ZO-1-(1-503) appeared as a double band suggesting premature
termination or post-translational modification of this construct. The
interaction was verified for endogenous proteins from adult mouse
kidney, demonstrating that this interaction occurs in vivo
(Fig. 1F). Since slit diaphragm proteins are partially
Triton X-100-insoluble in vivo, a special CHAPS-containing
buffer was used to solubilize the glomeruli. Confirming the biochemical
data, ZO-1 and Neph1 co-localized in the glomeruli of the native kidney
(Fig. 1G). In addition to the SH3 domain and a guanylate
kinase domain the multiadapter protein ZO-1 contains three PDZ domains.
These PDZ domains are located within the first 503 amino acids, the
region that confers binding to Neph1. We tested therefore which PDZ
domain mediated the interaction with Neph1. Pull-down experiments
revealed that the interaction specifically involved the first PDZ
domain of ZO-1 (Fig. 2A). Deletion of the last three amino acids of the carboxyl-terminal cytoplasmic tail of Neph1 (Thr-His-Val) completely abrogated binding (Fig. 2D). Interestingly mutation of the critical threonine
at the
INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS AND DISCUSSION
REFERENCES
-actinin 4 (10), and Neph1 (11) have recently been associated with the development of proteinuria. Neph1
contains five extracellular immunoglobulin-like domains and is
structurally related to nephrin. It is abundantly expressed in the
kidney, and disruption of the Neph1 gene in mice results in
effacement of glomerular podocytes, heavy proteinuria, and early
postnatal death (11). Neph1 belongs to a family of three closely
related proteins that bind to podocin (12). The interaction of nephrin
with the Src homology 3 (SH3)-containing adaptor protein CD2AP (8),
likely involved in protein trafficking and endocytosis (13), suggests
that the protein complex at the slit diaphragm is highly dynamic and
regulated by protein-protein interactions that organize the filtration
barrier and initiate intracellular signaling. Although many of the key
components of the slit diaphragm have now been identified, the
fundamental question remains how the different components are organized
at this specialized cell-cell junction.
MATERIALS AND METHODS
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS AND DISCUSSION
REFERENCES
-galactosidase expression vector (kindly provided by C. Cepko), and
vectors directing the expression of the proteins as indicated. The
total DNA amount was 1.5-2.0 µg/well. Cells were serum-starved for
12 h, harvested in cold phosphate-buffered saline, and lysed in
100 µl of reporter lysis buffer (Applied Biosystems, Norwalk, CT) for
10 min at 4 °C. Lysates were centrifuged at 14,000 rpm for 5 min to
remove insoluble material. Luciferase activity was determined using a commercial assay system (Applied Biosystems) and normalized for
-galactosidase activity to correct for transfection efficiency. Equal expression of proteins was ensured by Western blot analysis.
RESULTS AND DISCUSSION
TOP
ABSTRACT
INTRODUCTION
MATERIALS AND METHODS
RESULTS AND DISCUSSION
REFERENCES
2 position of the PDZ domain binding motif to glutamate
disrupted binding to ZO-1 (Fig. 2C). Since
threonine-to-glutamate mutation may mimic phosphorylation of the
threonine, this finding suggests that phosphorylation of the threonine
at the
2 position regulates the interaction between ZO-1 and Neph1 as
has been reported for other PDZ-dependent interactions
(25-27). However, this hypothesis needs further clarification. The PDZ
binding motif is highly conserved in all currently known Neph proteins
(Fig. 2B), and as demonstrated in Fig. 2E, all
Nephs bind to ZO-1 in transiently transfected HEK 293T cells; all
interactions were mediated by the first PDZ domain of ZO-1 (data not
shown). These data suggest that ZO-1 binding may be a general mechanism
in the regulation of Neph proteins.
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Fig. 1.
The cytoplasmic tail of Neph1 interacts with
ZO-1. A and B, HEK 293T cells were
transfected with the expression plasmids as indicated. Lysates were
subjected to immunoprecipitation with anti-Myc (A) or
anti-FLAG (B) antisera, resolved by SDS-PAGE, and
immunoblotted for Neph1. Neph1 specifically interacts with the first
503 amino acids of ZO-1. C-E, the cytoplasmic,
carboxyl-terminal domain of Neph1 or FAT was fused to human
immunoglobulin and the transmembrane domain of CD7
(sIg.7.Neph1) and co-transfected into HEK 293T cells with
FLAG-tagged ZO-1 constructs (F.ZO-1). After
immunoprecipitation with protein G, ZO-1 is detectable in the
precipitate containing Neph1 but not the control protein
(sIg.7) lacking the cytoplasmic domain of Neph1 or a fusion
protein with the cytoplasmic tail of FAT. Western blot analysis was
performed using the FLAG-specific M2 monoclonal antibody. F,
co-immunoprecipitation of Neph1 with ZO-1 from mouse kidneys. Mouse
kidneys were perfused in situ with ice-cold
phosphate-buffered saline, collected, and homogenized in a
CHAPS-containing buffer. After extensive preclearing the lysate was
subjected to immunoprecipitation with a control antibody (anti-HA,
lane 1) or anti-ZO-1 antiserum (lane 2). Neph1
could only be detected in the anti-ZO-1 precipitate. Lane 3 shows Neph1 in the kidney lysate. HEK 293T cell lysates untransfected
(lane 4) and transfected with a Neph1 cDNA (lane
5) served as control for antibody specificity. G,
co-localization of ZO-1 and Neph1 in the adult mouse kidney. Double
staining of ZO-1 (green) and Neph1 (red) shows a
basement membrane-like staining, a staining pattern typical for the
podocyte foot processes in glomeruli. An overlay technique using
conventional immunofluorescence microscopy (upper panel) and
confocal microscopy (lower panel) demonstrate
co-localization of ZO-1 and Neph1 in a basement membrane-like
appearance. WT, wild type; IF,
immunofluorescence.
View larger version (28K):
[in a new window]
Fig. 2.
The first PDZ domain of ZO-1 interacts with a
PDZ domain binding motif conserved in all Neph family members.
A, lysates of HEK 293T cells transfected with Neph1 cDNA
were subjected to a pull-down assay with recombinant affinity-purified
PDZ domains of ZO-1 fused to GST. Neph1 specifically interacted with
the first PDZ domain of ZO-1. The lower panel shows equal
expression levels of GST fusion proteins on a Coomassie Blue-stained
gel. B, the PDZ domain binding motif is highly conserved
among all Neph family members. Shown is the alignment of the last 18 amino acids of Neph1-3. Identical residues are highlighted
(yellow). The PDZ domain binding motif is shown in
red. C and D, mutation of the
threonine at the 2 position in the PDZ domain binding motif of Neph1
to glutamate (T787E) (C) or deletion of the last three amino
acids of Neph1 (D) abrogates binding of ZO-1. HEK 293T cells
were transfected with the expression plasmids as indicated. Lysates
were precipitated with protein G, resolved by SDS-PAGE, and
immunoblotted for ZO-1 with an anti-FLAG antibody (M2). E,
all Neph family members interact with ZO-1 in HEK 293T cells. Lysates
of cells transfected with the plasmid DNA as indicated were
precipitated with protein G. ZO-1 was detected with the M2 anti-FLAG
antiserum. WT, wild type.
It has been shown that ZO-1 may organize signal transduction, and
multiple evidences demonstrate a structural role in organizing transmembrane protein complexes (16). We examined therefore whether
ZO-1 expression modulates Neph1 signaling in HEK 293T cells. HEK 293T
cells contain only very low levels of ZO-1 and Neph1 and represent an
ideal model system to examine ZO-1 effects on Neph1 signaling.
AP-1 activity, measured by luciferase assays, is a very
sensitive parameter for the activation of several pathways including
the c-Jun NH2-terminal kinase, p38, extracellular
signal-regulated kinase 1/2, or phosphatidylinositol 3-kinase/AKT
signaling cascades. Since ZO-1 can recruit signal transduction
components including G proteins and kinases to cell-cell contacts and
may interact with a variety of additional signal transduction
components including Fyn, Grb-2, Crk, or glycogen synthase
kinase-3 (scansite.mit.edu/), we analyzed the effect of ZO-1
on Neph1-mediated AP-1 activation (Fig.
3A). Co-expression of ZO-1
significantly accelerated the Neph1-mediated AP-1 activity, whereas
ZO-1 alone had no effect (Fig. 3A). Augmentation of Neph1
signaling by ZO-1 requires direct binding of Neph1. Neph1-induced AP-1
activity was not modulated by ZO-1 after deletion of the PDZ binding
motif in Neph1 or mutation of ZO-1 (data not shown). We have previously
shown that the cytoplasmic tail of Neph1 is tyrosine-phosphorylated
(12). Interestingly ZO-1 co-expression strongly augmented tyrosine
phosphorylation of the carboxyl terminus of Neph1 (Fig. 3,
B-F) but did not influence tyrosine phosphorylation of
Neph1 T787E, a mutant that fails to interact with ZO-1 (data not
shown). Of note, ZO-1 did not only enhance tyrosine phosphorylation but
also augmented serine phosphorylation of the cytoplasmic tail of Neph1
as demonstrated by in vivo labeling and phosphoamino acid
analysis (Fig. 3F). These findings suggest that ZO-1
organizes Neph1 complexes and helps to recruit signal transduction
components to facilitate Neph1 signaling.
|
Recent work has highlighted the exquisite role of the slit diaphragm
for the integrity of the glomerular filter (1, 2, 28, 29). Several
critical proteins including nephrin, Neph1, podocin, and CD2AP have
been identified and were localized to the slit diaphragm (8, 30-33).
However, until now it is not clear how these different components are
organized at the filtration slit. Our results suggest that the PDZ
domain-containing multiadapter protein ZO-1 may help to cluster the
transmembrane protein Neph1. This conclusion is based on two
observations. First, ZO-1 specifically interacts with the
carboxyl-terminal cytoplasmic tail of Neph1. This interaction, mediated
by the first PDZ domain of ZO-1, was present in vivo and may
be dynamically regulated by phosphorylation. Second, direct binding of
ZO-1 to the carboxyl terminus of Neph1 dramatically alters the
phosphorylation state of Neph1 and the ability to induce signal
transduction. It is well established that PDZ domain proteins act as
scaffolds for signaling complexes and serve to recruit signal
transduction components (14, 34). ZO-1 has been shown to regulate
signaling involved in the control of cell polarity, the tightness of
the paracellular seal, and transcriptional responses (35, 36). Our
findings extend the function of ZO-1 and suggest that ZO-1 is involved
in the regulation of signaling by slit diaphragm proteins. It has been
suggested that ZO-1 in addition to its signaling function interacts
with the actin cytoskeleton and components of the paracellular seal (37). Although the functional implications of ZO-1/actin association have not yet been established in vivo ZO-1 could link Neph1
and its associated binding proteins to the actin cytoskeleton and contribute to the organization of the filtration slit. Given the importance of the submembranous actin cytoskeleton for the maintenance of the podocyte architecture (1), it will be interesting to evaluate
the exact role of ZO-1 for the organization of the podocyte actin
cytoskeleton in animal models that selectively target the ZO-1-Neph1 interaction.
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ACKNOWLEDGEMENTS |
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We thank Christina Engel, Stefanie Keller, and Birgit Schilling for excellent technical assistance; members of the Walz laboratory for helpful suggestions; and Dr. A. Blaukat for helpful advice with in vivo labeling and phosphoamino acid analysis.
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FOOTNOTES |
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* This study was supported by Deutsche Forschungsgemeinschaft Grants Be2212 and Wa597 and the Deutsche Nierenstiftung (to T. B. H.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
¶ To whom correspondence should be addressed. Tel.: 49-761-270-3250; Fax: 49-761-270-3245; E-mail: walz@med1.ukl.uni-freiburg.de.
Published, JBC Papers in Press, February 10, 2003, DOI 10.1074/jbc.C200678200
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ABBREVIATIONS |
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The abbreviations used are: NPHS1, congenital nephrotic syndrome of the Finnish type; NPHS2, steroid-resistant nephrotic syndrome; CD2AP, CD2-associated protein; PDZ, PSD95/Dlg/ZO-1; ZO, zonula occludens; SH3, Src homology 3; HA, hemagglutinin; CHAPS, 3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonic acid; HEK, human embryonic kidney; GST, glutathione S-transferase; AP-1, activator protein-1.
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REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() |
---|
1. |
Kerjaschki, D.
(2001)
J. Clin. Investig.
108,
1583-1587 |
2. | Tryggvason, K., and Wartiovaara, J. (2001) Curr. Opin. Nephrol. Hypertens. 10, 543-549[CrossRef][Medline] [Order article via Infotrieve] |
3. |
Tryggvason, K.
(1999)
J. Am. Soc. Nephrol.
10,
2440-2445 |
4. |
Huber, T. B.,
Kottgen, M.,
Schilling, B.,
Walz, G.,
and Benzing, T.
(2001)
J. Biol. Chem.
276,
41543-41546 |
5. | Fuchshuber, A., Jean, G., Gribouval, O., Gubler, M. C., Broyer, M., Beckmann, J. S., Niaudet, P., and Antignac, C. (1995) Hum. Mol. Genet. 4, 2155-2158[Abstract] |
6. | Boute, N., Gribouval, O., Roselli, S., Benessy, F., Lee, H., Fuchshuber, A., Dahan, K., Gubler, M. C., Niaudet, P., and Antignac, C. (2000) Nat. Genet. 24, 349-354[CrossRef][Medline] [Order article via Infotrieve] |
7. |
Shih, N. Y.,
Li, J.,
Karpitskii, V.,
Nguyen, A.,
Dustin, M. L.,
Kanagawa, O.,
Miner, J. H.,
and Shaw, A. S.
(1999)
Science
286,
312-315 |
8. |
Shih, N. Y.,
Li, J.,
Cotran, R.,
Mundel, P.,
Miner, J. H.,
and Shaw, A. S.
(2001)
Am. J. Pathol.
159,
2303-2308 |
9. | Inoue, T., Yaoita, E., Kurihara, H., Shimizu, F., Sakai, T., Kobayashi, T., Ohshiro, K., Kawachi, H., Okada, H., Suzuki, H., Kihara, I., and Yamamoto, T. (2001) Kidney Int. 59, 1003-1012[CrossRef][Medline] [Order article via Infotrieve] |
10. | Kaplan, J. M., Kim, S. H., North, K. N., Rennke, H., Correia, L. A., Tong, H. Q., Mathis, B. J., Rodriguez-Perez, J. C., Allen, P. G., Beggs, A. H., and Pollak, M. R. (2000) Nat. Genet. 24, 251-256[CrossRef][Medline] [Order article via Infotrieve] |
11. |
Donoviel, 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.,
and Powell, D. R.
(2001)
Mol. Cell. Biol.
21,
4829-4836 |
12. | Sellin, L., Huber, T. B., Gerke, P., Quack, I., Pavenstadt, H., and Walz, G. (2002) FASEB J. 17, 115-117[Medline] [Order article via Infotrieve] |
13. | Brett, T. J., Traub, L. M., and Fremont, D. H. (2002) Structure (Camb.) 10, 797-809[Medline] [Order article via Infotrieve] |
14. |
Hung, A. Y.,
and Sheng, M.
(2002)
J. Biol. Chem.
277,
5699-5702 |
15. |
Songyang, Z.,
Fanning, A. S.,
Fu, C.,
Xu, J.,
Marfatia, S. M.,
Chishti, A. H.,
Crompton, A.,
Chan, A. C.,
Anderson, J. M.,
and Cantley, L. C.
(1997)
Science
275,
73-77 |
16. | Mitic, L. L., Van Itallie, C. M., and Anderson, J. M. (2000) Am. J. Physiol. 279, G250-G254 |
17. |
Fanning, A. S.,
Mitic, L. L.,
and Anderson, J. M.
(1999)
J. Am. Soc. Nephrol.
10,
1337-1345 |
18. | Schnabel, E., Anderson, J. M., and Farquhar, M. G. (1990) J. Cell Biol. 111, 1255-1263[Abstract] |
19. |
Kawachi, H.,
Kurihara, H.,
Topham, P. S.,
Brown, D.,
Shia, M. A.,
Orikasa, M.,
Shimizu, F.,
and Salant, D. J.
(1997)
Am. J. Physiol.
273,
F984-F993 |
20. | Kurihara, H., Anderson, J. M., Kerjaschki, D., and Farquhar, M. G. (1992) Am. J. Pathol. 141, 805-816[Abstract] |
21. |
Tsiokas, L.,
Kim, E.,
Arnould, T.,
Sukhatme, V. P.,
and Walz, G.
(1997)
Proc. Natl. Acad. Sci. U. S. A.
94,
6965-6970 |
22. |
Benzing, T.,
Yaffe, M. B.,
Arnould, T.,
Sellin, L.,
Schermer, B.,
Schilling, B.,
Schreiber, R.,
Kunzelmann, K.,
Leparc, G. G.,
Kim, E.,
and Walz, G.
(2000)
J. Biol. Chem.
275,
28167-28172 |
23. |
Antignac, C.
(2002)
J. Clin. Investig.
109,
447-449 |
24. | Yaffe, M. B., Leparc, G. G., Lai, J., Obata, T., Volinia, S., and Cantley, L. C. (2001) Nat. Biotechnol. 19, 348-353[CrossRef][Medline] [Order article via Infotrieve] |
25. | Cao, T. T., Deacon, H. W., Reczek, D., Bretscher, A., and von Zastrow, M. (1999) Nature 401, 286-290[CrossRef][Medline] [Order article via Infotrieve] |
26. | Cohen, N. A., Brenman, J. E., Snyder, S. H., and Bredt, D. S. (1996) Neuron 17, 759-767[Medline] [Order article via Infotrieve] |
27. |
Chetkovich, D. M.,
Chen, L.,
Stocker, T. J.,
Nicoll, R. A.,
and Bredt, D. S.
(2002)
J. Neurosci.
22,
5791-5796 |
28. |
Schwarz, K.,
Simons, M.,
Reiser, J.,
Saleem, M. A.,
Faul, C.,
Kriz, W.,
Shaw, A. S.,
Holzman, L. B.,
and Mundel, P.
(2001)
J. Clin. Investig.
108,
1621-1629 |
29. |
Simons, M.,
Schwarz, K.,
Kriz, W.,
Miettinen, A.,
Reiser, J.,
Mundel, P.,
and Holthofer, H.
(2001)
Am. J. Pathol.
159,
1069-1077 |
30. |
Roselli, S.,
Gribouval, O.,
Boute, N.,
Sich, M.,
Benessy, F.,
Attie, T.,
Gubler, M. C.,
and Antignac, C.
(2002)
Am. J. Pathol.
160,
131-139 |
31. |
Holthofer, H.,
Ahola, H.,
Solin, M. L.,
Wang, S.,
Palmen, T.,
Luimula, P.,
Miettinen, A.,
and Kerjaschki, D.
(1999)
Am. J. Pathol.
155,
1681-1687 |
32. | Holzman, L. B., St. John, P. L., Kovari, I. A., Verma, R., Holthofer, H., and Abrahamson, D. R. (1999) Kidney Int. 56, 1481-1491[CrossRef][Medline] [Order article via Infotrieve] |
33. |
Ruotsalainen, V.,
Ljungberg, P.,
Wartiovaara, J.,
Lenkkeri, U.,
Kestila, M.,
Jalanko, H.,
Holmberg, C.,
and Tryggvason, K.
(1999)
Proc. Natl. Acad. Sci. U. S. A.
96,
7962-7967 |
34. | Harris, B. Z., and Lim, W. A. (2001) J. Cell Sci. 114, 3219-3231[Medline] [Order article via Infotrieve] |
35. | Balda, M. S., and Matter, K. (2000) Semin. Cell Dev. Biol. 11, 281-289[CrossRef][Medline] [Order article via Infotrieve] |
36. |
Balda, M. S.,
and Matter, K.
(2000)
EMBO J.
19,
2024-2033 |
37. |
Fanning, A. S.,
Ma, T. Y.,
and Anderson, J. M.
(2002)
FASEB J.
16,
1835-1837 |