1 Ronald O. Perelman Department of Dermatology and Departments of Pharmacology and Urology, Kaplan Comprehensive Cancer Center, New York University School of Medicine, New York, New York 10016; 2 Departments of Medicine and Cell Biology and Physiology, University of Pittsburgh School of Medicine, Pittsburgh, Pennsylvania 15213; and 3 Laboratory of Cellular Biology, National Institute on Deafness and Other Communication Disorders, National Institutes of Health, Bethesda, Maryland 20892
![]() |
ABSTRACT |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Although water, small
nonelectrolytes, and gases are freely permeable through most biological
membranes, apical membranes of certain barrier epithelia exhibit
extremely low permeabilities to these substances. The role of integral
membrane proteins in this barrier function has been unclear. To study
this problem, we have ablated the mouse gene encoding uroplakin III
(UPIII), one of the major protein subunits in urothelial apical
membranes, and measured the permeabilities of these membranes. Ablation
of the UPIII gene greatly diminishes the amounts of uroplakins on the
apical urothelial membrane (Hu P, Deng FM, Liang FX, Hu CM, Auerbach
AB, Shapiro E, Wu XR, Kachar B, and Sun TT. J Cell Biol 151: 961-972, 2000). Our results indicate that normal mouse
urothelium exhibits high transepithelial resistance and low urea and
water permeabilities. The UPIII-deficient urothelium exhibits a normal transepithelial resistance (normal 2,024 ± 122, knockout
2,322 ± 114 · cm2; P > 0.5). However, the UPIII-deficient apical membrane has a significantly
elevated water permeability (normal 0.91 ± 0.06, knockout
1.83 ± 0.14 cm/s × 10
5; P < 0.05). The urea permeability of the UPIII-deficient membrane also
increased, although to a lesser extent (normal 2.22 ± 0.24, knockout 2.93 ± 0.31 cm/s × 10
6;
P = 0.12). These results indicate that reduced
targeting of uroplakins to the apical membrane does not significantly
alter the tight junctional barrier but does double the water
permeability. We provide the first demonstration that integral membrane
proteins contribute to the apical membrane permeability barrier
function of urothelium.
urothelium; uroplakin; permeability; vesicoureteral reflux; knockout
![]() |
INTRODUCTION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
THE MAMMALIAN
BLADDER MAINTAINS for prolonged periods large gradients for
water, small nonelectrolytes, ions, protons, and ammonium between the
urine it stores and blood (4, 5, 16, 36). Thus in humans,
urine osmolality varies widely between 50 and 1,000 mosmol/kgH2O, but blood osmolality remains constant between
280 and 290 mosmol/kgH2O. Similarly, urinary pH varies from
4.5 to 8.0 while blood pH remains stable at around 7.4 (4, 5, 16,
36). The urothelial barrier function is therefore essential for
the kidney to maintain homeostasis. Measurements of barrier function in
vitro indeed showed that the bladder exhibits exceptionally low
permeabilities to ions (transepithelial resistances of
20,000-30,000 · cm2) as well as to water
and urea (2, 15-18, 21, 25). The apical membranes and
the tight junctions between the uppermost urothelial cells (the
so-called umbrella cells), in combination, represent the main site of
the bladder permeability barrier (15, 17, 18, 21, 25).
The apical membranes of umbrella cells are covered with numerous rigid-looking plaques consisting of two-dimensional hexagonal arrays of 16-nm particles (1, 6, 11, 13, 27, 29, 31). Purified urothelial plaques contain four major integral membrane proteins called uroplakins that are synthesized as the major differentiation products of mammalian urothelia (24, 28, 32-35). Although much has been learned about the sequences of these proteins as well as some aspects of their three-dimensional structure as a complex within the membrane, relatively little is known about their precise function. The hypothesized functions of urothelial plaques include the reversible adjustment of urothelial apical surface (5, 18, 19) and the physical strengthening of the apical surface (27). In addition, because the urothelium represents such an effective barrier, it is tempting to speculate that urothelial plaques augment specialized lipid structures and contribute to bladder barrier function (4, 5). To test these hypotheses, we have developed knockout mice lacking a functional uroplakin III (UPIII) gene (9). These mice not only lack UPIII but also have a markedly attenuated expression and apical targeting of the other three uroplakins (UPIa, UPIb, and UPII), resulting in a greatly reduced number and average size of the apical plaques. The animals also develop vesicoureteral reflux and hydronephrosis (9).
The present studies focus on the effects of ablating the UPIII gene on the barrier function of the urothelium, in terms of the apical membrane permeabilities and of tight junctional function. We show here that the depletion of uroplakins from urothelial apical surface had no effects on transepithelial resistance but led to elevated water permeability. These results provide the first evidence that integral membrane proteins play a role in establishing certain aspects of urothelial barrier function, possibly by influencing the asymmetry and by reducing the fluidity of the bilayer leaflets.
![]() |
METHODS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Materials and solutions. Unless specified otherwise, all chemicals were obtained from Sigma (St. Louis, MO) and were of reagent grade. The composition of the NaCl-Ringer bathing solution was (in mM) 111.2 NaCl, 25 NaHCO3, 5.8 KCl, 2 CaCl2, 1.2 MgSO4, 1.2 KH2PO4, and 11.1 glucose. This solution was oxygenated with 95% O2-5% CO2, pH 7.4, at 37°C.
Mouse strains. The generation and characterization of the UPIII knockout mice were described earlier (9). The knockout vector was designed to delete the first three exons of the UPIII gene. Two 129/SvEv embryonic stem cell (line W4) clones harboring the correct homologous recombination events were amplified, aggregated with eight cell-stage embryos of Swiss Webster (SW) mice, and implanted into pseudopregnant females. Five chimeric mice from these two embryonic stem cell lines were germline transmitting and were bred with SW mice to yield hybrid homozygotes or mated with 129/SvEv mice to yield inbred 129/SvEv UPIII knockout mice (9). The SW mice used as the control and knockout mice were homozygous for UPIII deletion (9). Mice were maintained overnight in metabolic cages for urine collection. All mice had free access to water and a normal mouse chow diet for all experiments.
Measurement of urothelial barrier function. Animal experiments were performed in accordance with the animal use and care committees of both New York University and the University of Pittsburgh. Bladders were excised after lethal anesthesia, washed in oxygenated NaCl Ringer solution, and carefully stretched and mounted on a small ring in the same solution at 37°C. The bladder was then placed between modified Ussing chambers (15, 16, 25). Both compartments of the chambers were under constant stirring and temperature control and allowed electrical measurements and sampling.
The transepithelial potential was measured as described, with Ag-AgCl electrodes placed close to and on opposite sides of the epithelium (15, 16, 25). Transepithelial resistance was determined by passing current through the Ag-AgCl electrodes placed in the rear of each half-chamber across the tissue and measuring the resulting voltage deflection. Both voltage-sensing and current-passing electrodes were connected to an automatic voltage clamp (EC-825, Warner Instruments, Hamden, CT), which was, in turn, connected to a microcomputer with a MacLab interface (16). All permeability measurements were performed at 37°C after stabilization of the transepithelial resistance. Diffusive water, urea, and butanol permeability coefficients were determined by using isotopic fluxes as described (15, 16, 25). Briefly, tritiated water, [14C]urea or [14C]butanol were added to the apical chamber (13.5-ml total volume) and 100-µl samples of both apical and basolateral chambers were taken every 15 min. Sample volumes were replaced quantitatively with warmed NaCl-Ringer. Sample radioactivities were counted with a liquid scintillation counter (model 1500, Packard Tri-Carb), and flux rates and permeabilities were calculated as described (15, 16, 25). To determine the contribution of unstirred layers to the observed flux rates, water and urea fluxes were measured after the exposure of the bladder apical surface to Triton X-100 (7.4 µl/ml). In some experiments, unstirred layers were measured with [14C]butanol (15). Because these results agreed with those obtained with Triton X-100, the values obtained with Triton X-100 were used to correct for unstirred layer effects. Water and urea permeability are reported as apical membrane permeabilities, in which unstirred layers have been taken into account.Electron microscopy. For transmission electron microscopy (EM), mouse bladder was cut into small pieces (<1 mm2), fixed with 2.5% glutaraldehyde in 0.1 M sodium cacodylate buffer, pH 7.4, postfixed with 1% (wt/vol) osmium tetroxide, and embedded in Epon 812 (Polysciences) as described (23). For scanning EM, bladders were bisected, fixed as above, and critical point dried (11).
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Effects of UPIII knockout on organ structure and kidney function.
Although the UPIII knockout mice appeared healthy, detailed
measurements of the body and kidney weights as well as the diameters of
the stretched bladder revealed some abnormalities (Table
1). In both males and females, bladders
were larger in diameter in UPIII knockout animals compared with
controls (P < 0.001 for both sexes). In male mice,
body and kidney weights of the knockouts were not statistically
different from the control animals. However, in female mice, the
knockout animals were smaller and their kidneys were larger
(P < 0.05) than the controls. Pathological examination of kidneys from knockout animals of both sexes showed dilatation of the
renal pelvis, as well as clear-cut hydronephrosis, whereas control
animals exhibited normal renal morphology (9). Glomeruli appeared normal in knockout and control animals. Bladders from knockout
animals were far more easily stretched onto the rings of the Ussing
chambers than the bladders from control animals. These results indicate
that the lack of UPIII expression made the bladder as a whole more
compliant than in control animals.
|
|
Barrier function of bladders from control and knockout mice.
To assess the effects of UPIII depletion on the permeability function
of the urothelium, we measured the transepithelial resistance, water
permeabilities, and urea permeabilities of control and UPIII knockout
mice (Fig. 1). These
parameters of normal mouse urothelium were found to be comparable to
those of other species, including rats, rabbits, guinea pigs, and cats,
thus establishing that the bladders of all of these species exhibit
high transepithelial resistances, as well as strikingly low
permeabilities to water and urea (Table
3) (2, 4, 15, 16, 19, 21).
Although all of the water permeabilities are low, the permeability of
the mouse urothelium is four- to fivefold lower than that of the rat or
the cat. Figure 1 shows the permeability data for control and knockout
mouse bladders, with corrections for unstirred layers. Transepithelial
resistances were similar between control and knockout animals,
suggesting that the ablation of UPIII did not alter the function of the
tight junctions between umbrella cells. By contrast, water permeability
was clearly increased in bladders from knockout animals compared with
controls (P < 0.05). Urea permeability tended to be
higher in the knockout animals as well, although this apparent difference did not reach statistical significance (P = 0.12). When data from males and females were pooled, urea permeability was significantly higher in the UPIII knockout mice than in controls (P 0.05). For resistance, water, and urea permeabilities,
values for control or knockout mice did not differ between male and
female mice. In all cases, addition of Triton X-100 increased the
measured fluxes of water and urea by more than fourfold; these
increases were similar in control and knockout animals. These results
indicate that the unstirred layers were similar in magnitude for the
control and knockout bladders and that the observed differences in the permeability of normal and UPIII knockout animals could be attributed to differences in the properties of the apical membrane.
|
|
EM of bladders from control and knockout mice.
To determine the ultrastructural bases for the observed physiological
alterations, we examined the umbrella cells from control and knockout
animals by transmission (Fig. 2) and
scanning EM (Fig. 3), paying special
attention to cell/cell junctions. Umbrella cells from
control animals exhibited numerous fusiform subapical vesicles and an
apical surface that was covered with plaques; in contrast, the
superficial cells from knockout animals had small, round immature
vesicles and few apical surface plaques (Fig. 2 and see also Ref.
9). Umbrella cells from both control and knockout animals
had well-developed tight junctions (Fig. 2). However, in knockout
animals there appeared to be an increase in the number of desmosomes, a
feature that was observed in numerous fields in tissues taken from
multiple bladders from both control and knockout animals (Fig. 2).
Scanning EM of umbrella cells from normal mouse urothelium revealed
large cells covered by ridges (most likely due to the presence of
numerous urothelial plaques) on their apical surfaces (Fig. 3)
(5, 9, 10, 16, 26). In contrast, surface cells in knockout
animals displayed numerous microvilli, and the cells were far smaller
than those of bladders from control animals (Fig. 3 and see also Ref.
9).
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Uroplakin knockout increases the water permeability of urothelial membrane. We showed here that the ablation of UPIII doubled the water permeability of urothelial membrane. Because the absolute value for water permeability of the bladders from knockout animals was still quite low, it is important to rule out the possibility of an "unstirred layer" effect, whereby the unstirred layer adjacent to the urothelial apical surface limits the extent to which an increased rate of water flux could be detected in our apparatus. However, addition of Triton X-100 to the apical surface of the urothelium from both control and knockout mice led to similar striking increases in water permeability. This proves that had water permeability in knockout animals been higher, we would have been able to detect it, and the loss of UPIII results in only a partial reduction in the water barrier function (see below).
The strikingly low water permeabilities of both control and UPIII knockout urothelia suggest that the lipid component of the mouse urothelial apical membrane is particularly resistant to water permeation and that this resistance persists even when the levels of uroplakins in the apical membrane are markedly reduced. Future studies will address the lipid structures of the apical membranes in control and uroplakin knockout mice. An additional issue concerns the surface area of apical membrane in control and knockout animals. Although the surface area of epithelium was identical in the two preparations, we cannot quantify the levels of surface area of membrane in situ in the two animal types because we cannot measure capacitance in the intact bladder.Complementary roles of lipids and integral membrane proteins in barrier function. In prior studies of barrier apical membranes, we and others have determined the unique lipid compositions of urothelial plaques (12, 30) and defined certain aspects of lipid structure that play a critical role in reducing the membrane permeabilities to small molecules (7, 8, 14, 25). Biological membranes have asymmetric lipid bilayers, with phosphatidyl choline, glycosphingolipids (such as cerebrosides), sphingomyelin, and cholesterol segregated in the outer leaflet, and phosphatidyl serine and ethanolamine segregated in the inner or cytoplasmic leaflet (7, 8, 14, 25). Because the outer and inner leaflets of the bilayer act as independent resistors of permeation (8), barrier membranes tend to use phospholipids whose fatty acids have few double bonds and to have large quantities of sphingomyelin and cholesterol (which rigidify the acyl chains of the fatty acids) in their outer leaflets (7, 8, 14, 25). It is entirely possible that the special composition and structural packing of lipids that are characteristic of the urothelial apical membrane are largely responsible for the extraordinarily effective urothelial barrier function. However, how the lipids accomplish this task and whether membrane proteins also play a role in this process remain unclear.
A striking feature of mammalian urothelial apical membrane is that >90% of this membrane is covered by two-dimensional crystalline arrays of 16-nm protein particles (6, 13, 27, 29). These plaques are composed of four major uroplakins, i.e., UPIa (27 kDa), UPIb (28 kDa), UPII (15 kDa), and UPIII (47 kDa) (28). The fact that most of the hydrophilic domains of uroplakins are extracellular can account for the fact that the outer leaflet of apical urothelial plaques is twice as thick as the inner leaflet (24, 28, 33, 34). These crystalline arrays of uroplakins form rigid-appearing plaques within the apical membrane; areas between the plaques are referred to as hinge regions (22). We showed recently that the ablation of UPIII gene results in a marked diminution of the plaque surface area in favor of hinge regions and that UPIII knockout led to vesicoureteral reflux (9). UPIII knockout mice thus provide an excellent model for defining the role of uroplakins in urothelial physiology and have allowed us to establish in this study a role of uroplakin proteins in water barrier function (Fig. 1).Possible mechanism of uroplakin influence on barrier function. There are several possible mechanisms by which uroplakins affect the water permeability of urothelium. Thus uroplakins may function by organizing and rigidifying the lipids in the outer leaflet of the bilayer so that water cannot penetrate into the aliphatic chains of the fatty acids (7, 8). Alternatively, uroplakins may stabilize a particularly rigid set of lipids within the outer leaflet. These mechanisms seem reasonable given our earlier finding that the 16-nm urothelial particle is a mushroom-shaped structure with a 16-nm head domain that is exposed luminally and is anchored into the lipid bilayer via a thinner 11-nm transmembrane domain (11). The transmembrane domains of uroplakins are most likely also regularly packed forming two-dimensional crystalline structures that can exert major constraints on the arrangement of lipid molecules. In addition, the uroplakin domains that are close to the outer and inner leaflet of the bilayer are clearly asymmetrical (34); this protein asymmetry may exert strong constraints on the allowable lipid composition of the outer and inner leaflets. The uroplakin proteins may therefore play a major role in determining both the asymmetrical lipid composition of the two leaflets and the rigidity of the (interparticle) lipid components, thereby influencing the overall permeability of the apical urothelial membrane. The fact that the apical urothelial surface of UPIII knockout mice is still partially covered by urothelial plaques, albeit smaller (9), may explain why the water barrier function was only partially compromised.
Other functional roles of uroplakins. The ablation of the UPIII gene had several striking effects on cell structure (9). Umbrella cells from knockout animals lacked apical membrane plaques on transmission EM (Fig. 2) and exhibited microvilli rather than ridges on scanning EM (Fig. 3). Their subapical vesicles were discoidal rather than fusiform (Fig. 2). In addition, the cells were smaller than in control animals (Fig. 3), and it appeared that the junctions between the cells had more desmosomes in knockout animals (Fig. 2). These results indicate that uroplakins are responsible for the unique shape of the apical membranes and of the subapical vesicles of umbrella cells.
It is unclear why cells lacking uroplakin do not grow to the large size of normal umbrella cells. It is possible that the absence of uroplakin renders the apical membrane unstable as it expands so that umbrella cells that become too large disintegrate and are sloughed off. This interpretation is consistent with our laboratory's recent observation that there seem to be more apoptotic cells than control cells detaching from the UPIII-deficient urothelium (9). In unpublished studies in which we have induced a selective injury to the umbrella cells by using protamine, we have shown that regenerating urothelial cells reach the surface and develop normal ridges, normal plaques, and normal barrier function before they begin to enlarge (Lavelle JP, Meyers S, Doty D, Apodaca G, and Zeidel ML, unpublished observations).Uroplakin knockout does not affect the tight junctional function.
An unexpected finding was that UPIII knockout mice had no apparent
effects on transepithelial resistance, and EM revealed that knockout
animals exhibited intact tight junctions. These results
indicate that uroplakins play no role in the formation of tight
junctions and that the lack of uroplakins does not appear to alter
trafficking of essential tight junctional proteins such as claudins to
the correct location (Fig. 2). Because the surface urothelial cells are
smaller in the knockout animals, tight junctions actually occupy a much
higher proportion of the apical surface area in these animals than in
controls. Because these studies were performed in intact bladders
rather than dissected urothelium, our measured transepithelial
resistances for bladders from both control and knockout mice were well
below the 20,000 · cm2 that is observed with
dissected urothelium. This prevented us from determining whether the
tight junctions in the knockout animals functioned quantitatively as
well as those of the controls. It is also unclear why there appear to
be more desmosomes in the knockout animals than in the controls.
![]() |
ACKNOWLEDGEMENTS |
---|
We thank Alexander Joyner for advice on the generation of the knockout mice and Edith S. Robbins and David Sabatini for use of their electron microscopes.
![]() |
FOOTNOTES |
---|
This work was supported by National Institute of Diabetes and Digestive and Kidney Diseases Grants DK-43955 and DK-48217 (to M. L. Zeidel) and DK-52206, DK-57269, and DK-3975 (to T.-T. Sun).
Address for reprint requests and other correspondence: M. L. Zeidel, Dept. of Medicine, Univ. of Pittsburgh School of Medicine, Rm. 1218 Scaife Hall, 3550 Terrace St., Pittsburgh, PA 15213 (E-mail: Zeidel{at}msx.dept-med.pitt.edu); or T-T Sun, Depts. of Pharmacology and Urology, New York Univ. Medical School, 550 First Ave., New York, NY 10016 (E-mail: sunt01{at}mcrcr.med.nyu.edu).
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.
July 30, 2002;10.1152/ajprenal.00043.2002
Received 31 January 2002; accepted in final form 11 July 2002.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
1.
Brisson, A,
and
Wade RH.
Three-dimensional structure of luminal plasma membrane protein from urinary bladder.
J Mol Biol
166:
21-36,
1983[ISI][Medline].
2.
Chang, A,
Hammond TG,
Sun TT,
and
Zeidel ML.
Permeability properties of the mammalian bladder apical membrane.
Am J Physiol Cell Physiol
267:
C1483-C1492,
1994
3.
Curhan, GC,
McDougal WS,
and
Zeidel ML.
Urinary tract obstruction.
In: The Kidney, edited by Brenner BM.. Philadelphia: Saunders, 2000.
4.
Hicks, RM.
The permeability of rat transitional epithelium. Keratinization and the barrier to water.
J Cell Biol
28:
21-31,
1966
5.
Hicks, RM.
The mammalian urinary bladder: an accommodating organ.
Biol Rev Camb Philos Soc
50:
215-246,
1975[ISI][Medline].
6.
Hicks, RM,
and
Ketterer B.
Hexagonal lattice of subunits in the thick luminal membrane of the rat urinary bladder.
Nature
224:
1304-1305,
1969[ISI][Medline].
7.
Hill, WG,
Rivers RL,
and
Zeidel ML.
Role of leaflet asymmetry in the permeability of model biological membranes to protons, solutes, and gases.
J Gen Physiol
114:
405-414,
1999
8.
Hill, WG,
and
Zeidel ML.
Reconstituting the barrier properties of a water-tight epithelial membrane by design of leaflet-specific liposomes.
J Biol Chem
275:
30176-30185,
2000
9.
Hu, P,
Deng FM,
Liang FX,
Hu CM,
Auerbach AB,
Shapiro E,
Wu XR,
Kachar B,
and
Sun TT.
Ablation of uroplakin III gene results in small urothelial plaques, urothelial leakage, and vesicoureteral reflux.
J Cell Biol
151:
961-972,
2000
10.
Jacobs, JB,
Arai M,
Cohen SM,
and
Friedell GH.
Early lesions in experimental bladder cancer: scanning electron microscopy of cell surface markers.
Cancer Res
36:
2512-2517,
1976[Abstract].
11.
Kachar, B,
Liang F,
Lins U,
Ding M,
Wu XR,
Stoffler D,
Aebi U,
and
Sun TT.
Three-dimensional analysis of the 16 nm urothelial plaque particle: luminal surface exposure, preferential head-to-head interaction, and hinge formation.
J Mol Biol
285:
595-608,
1999[ISI][Medline].
12.
Ketterer, B,
Hicks RM,
Christodoulides L,
and
Beale D.
Studies of the chemistry of the luminal plasma membrane of rat bladder epithelial cells.
Biochim Biophys Acta
311:
180-190,
1973[ISI][Medline].
13.
Koss, LG.
The asymmetric unit membranes of the epithelium of the urinary bladder of the rat. An electron microscopic study of a mechanism of epithelial maturation and function.
Lab Invest
21:
154-168,
1969[ISI][Medline].
14.
Lande, MB,
Donovan JM,
and
Zeidel ML.
The relationship between membrane fluidity and permeabilities to water, solutes, ammonia, and protons.
J Gen Physiol
106:
67-84,
1995[Abstract].
15.
Lavelle, JP,
Apodaca G,
Meyers SA,
Ruiz WG,
and
Zeidel ML.
Disruption of guinea pig urinary bladder permeability barrier in noninfectious cystitis.
Am J Physiol Renal Physiol
274:
F205-F214,
1998
15a.
Lavelle, J,
Meyers S,
Ramage R,
Bastacky S,
Doty D,
Apodaca G,
and
Zeidel ML.
Bladder permeability barrier: recovery from selective injury of surface epithelial cells.
Am J Physiol Renal Physiol
283:
F242-F253,
2002
16.
Lavelle, JP,
Meyers SA,
Ruiz WG,
Buffington CA,
Zeidel ML,
and
Apodaca G.
Urothelial pathophysiological changes in feline interstitial cystitis: a human model.
Am J Physiol Renal Physiol
278:
F540-F553,
2000
17.
Lewis, SA,
Berg JR,
and
Kleine TJ.
Modulation of epithelial permeability by extracellular macromolecules.
Physiol Rev
75:
561-589,
1995
18.
Lewis, SA,
and
de Moura JL.
Incorporation of cytoplasmic vesicles into apical membrane of mammalian urinary bladder epithelium.
Nature
297:
685-688,
1982[ISI][Medline].
19.
Lewis, SA,
and
de Moura JL.
Apical membrane area of rabbit urinary bladder increases by fusion of intracellular vesicles: an electrophysiological study.
J Membr Biol
82:
123-136,
1984[ISI][Medline].
20.
Lewis, SA,
and
Diamond JM.
Na+ transport by rabbit urinary bladder, a tight epithelium.
J Membr Biol
28:
1-40,
1976[ISI][Medline].
21.
Lewis, SA,
and
Hanrahan JW.
Physiological approaches for studying mammalian urinary bladder epithelium.
Methods Enzymol
192:
632-650,
1990[Medline].
22.
Liang, F,
Kachar B,
Ding M,
Zhai Z,
Wu XR,
and
Sun TT.
Urothelial hinge as a highly specialized membrane: detergent-insolubility, urohingin association, and in vitro formation.
Differentiation
65:
59-69,
1999[ISI][Medline].
23.
Liang, FX,
Riedel I,
Deng FM,
Zhou G,
Xu C,
Wu XR,
Kong XP,
Moll R,
and
Sun TT.
Organization of uroplakin subunits: transmembrane topology, pair formation and plaque composition.
Biochem J
355:
13-18,
2001[ISI][Medline].
24.
Lin, JH,
Wu XR,
Kreibich G,
and
Sun TT.
Precursor sequence, processing, and urothelium-specific expression of a major 15-kDa protein subunit of asymmetric unit membrane.
J Biol Chem
269:
1775-1784,
1994
25.
Negrete, HO,
Lavelle JP,
Berg J,
Lewis SA,
and
Zeidel ML.
Permeability properties of the intact mammalian bladder epithelium.
Am J Physiol Renal Fluid Electrolyte Physiol
271:
F886-F894,
1996
26.
Pauli, BU,
Alroy J,
and
Weinstein RS.
The ultrastructure and pathobiology of urinary bladder cancer.
In: The Pathology of Bladder Cancer, edited by Bryan GT,
and Cohen SM.. Boca Raton, FL: CRC, 1983, p. 42-140.
27.
Staehelin, LA,
Chlapowski FJ,
and
Bonneville MA.
Lumenal plasma membrane of the urinary bladder. I. Three-dimensional reconstruction from freeze-etch images.
J Cell Biol
53:
73-91,
1972
28.
Sun, TT,
Zhao H,
Provet J,
Aebi U,
and
Wu XR.
Formation of asymmetric unit membrane during urothelial differentiation.
Mol Biol Rep
23:
3-11,
1996[ISI][Medline].
29.
Vergara, J,
Longley W,
and
Robertson JD.
A hexagonal arrangement of subunits in membrane of mouse urinary bladder.
J Mol Biol
46:
593-596,
1969[ISI][Medline].
30.
Vergara, J,
Zambrano F,
Robertson JD,
and
Elrod H.
Isolation and characterization of luminal membranes from urinary bladder.
J Cell Biol
61:
83-94,
1974
31.
Walz, T,
Haner M,
Wu XR,
Henn C,
Engel A,
Sun TT,
and
Aebi U.
Towards the molecular architecture of the asymmetric unit membrane of the mammalian urinary bladder epithelium: a closed "twisted ribbon" structure.
J Mol Biol
248:
887-900,
1995[ISI][Medline].
32.
Wu, XR,
Manabe M,
Yu J,
and
Sun TT.
Large scale purification and immunolocalization of bovine uroplakins I, II, and III. Molecular markers of urothelial differentiation.
J Biol Chem
265:
19170-19179,
1990
33.
Wu, XR,
and
Sun TT.
Molecular cloning of a 47 kDa tissue-specific and differentiation-dependent urothelial cell surface glycoprotein.
J Cell Sci
106:
31-43,
1993
34.
Yu, J,
Lin JH,
Wu XR,
and
Sun TT.
Uroplakins Ia and Ib, two major differentiation products of bladder epithelium, belong to a family of four transmembrane domain (4TM) proteins.
J Cell Biol
125:
171-182,
1994[Abstract].
35.
Yu, J,
Manabe M,
Wu XR,
Xu C,
Surya B,
and
Sun TT.
Uroplakin I: a 27-kD protein associated with the asymmetric unit membrane of mammalian urothelium.
J Cell Biol
111:
1207-1216,
1990[Abstract].
36.
Zeidel, ML.
Low permeabilities of apical membranes of barrier epithelia: what makes watertight membranes watertight?
Am J Physiol Renal Fluid Electrolyte Physiol
271:
F243-F245,
1996