scholarly journals The keratin-binding protein Albatross regulates polarization of epithelial cells

2008 ◽  
Vol 183 (1) ◽  
pp. 19-28 ◽  
Author(s):  
Masahiko Sugimoto ◽  
Akihito Inoko ◽  
Takashi Shiromizu ◽  
Masanori Nakayama ◽  
Peng Zou ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Binding Protein ◽  
Cell Polarization ◽  
Junctional Complex ◽  
Apical Surface ◽  
Apical Side ◽  
Apical Junctional Complex ◽  
Keratin Filaments ◽  
Keratin Filament ◽  
Filament Network

The keratin intermediate filament network is abundant in epithelial cells, but its function in the establishment and maintenance of cell polarity is unclear. Here, we show that Albatross complexes with Par3 to regulate formation of the apical junctional complex (AJC) and maintain lateral membrane identity. In nonpolarized epithelial cells, Albatross localizes with keratin filaments, whereas in polarized epithelial cells, Albatross is primarily localized in the vicinity of the AJC. Knockdown of Albatross in polarized cells causes a disappearance of key components of the AJC at cell–cell borders and keratin filament reorganization. Lateral proteins E-cadherin and desmoglein 2 were mislocalized even on the apical side. Although Albatross promotes localization of Par3 to the AJC, Par3 and ezrin are still retained at the apical surface in Albatross knockdown cells, which retain intact microvilli. Analysis of keratin-deficient epithelial cells revealed that keratins are required to stabilize the Albatross protein, thus promoting the formation of AJC. We propose that keratins and the keratin-binding protein Albatross are important for epithelial cell polarization.

scholarly journals Differential Roles for Actin Polymerization and a Myosin II Motor in Assembly of the Epithelial Apical Junctional Complex

2005 ◽  
Vol 16 (6) ◽  
pp. 2636-2650 ◽  
Author(s):  
Andrei I. Ivanov ◽  
Dirk Hunt ◽  
Markus Utech ◽  
Asma Nusrat ◽  
Charles A. Parkos
Keyword(s):  
Epithelial Cells ◽  
Actin Polymerization ◽  
Myosin Ii ◽  
Adherens Junction ◽  
Cell Polarization ◽  
Junctional Complex ◽  
Intestinal Epithelial ◽  
Actin Reorganization ◽  
Apical Junctional Complex ◽  
Syndrome Protein

Differentiation and polarization of epithelial cells depends on the formation of the apical junctional complex (AJC), which is composed of the tight junction (TJ) and the adherens junction (AJ). In this study, we investigated mechanisms of actin reorganization that drive the establishment of AJC. Using a calcium switch model, we observed that formation of the AJC in T84 intestinal epithelial cells began with the assembly of adherens-like junctions followed by the formation of TJs. Early adherens-like junctions and TJs readily incorporated exogenous G-actin and were disassembled by latrunculin B, thus indicating dependence on continuous actin polymerization. Both adherens-like junctions and TJs were enriched in actin-related protein 3 and neuronal Wiskott-Aldrich syndrome protein (N-WASP), and their assembly was prevented by the N-WASP inhibitor wiskostatin. In contrast, the formation of TJs, but not adherens-like junctions, was accompanied by recruitment of myosin II and was blocked by inhibition of myosin II with blebbistatin. In addition, blebbistatin inhibited the ability of epithelial cells to establish a columnar phenotype with proper apico-basal polarity. These findings suggest that actin polymerization directly mediates recruitment and maintenance of AJ/TJ proteins at intercellular contacts, whereas myosin II regulates cell polarization and correct positioning of the AJC within the plasma membrane.

Evidence that prostaglandin E2 stimulates chloride secretion in cultured A6 renal epithelial cells

1986 ◽  
Vol 250 (3) ◽  
pp. F511-F515 ◽  
Author(s):  
R. Keeler ◽  
N. L. Wong
Keyword(s):  
Epithelial Cells ◽  
Prostaglandin E2 ◽  
Short Circuit ◽  
Basal Medium ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Apical Surface ◽  
Renal Epithelial Cells ◽  
Apical Side ◽  
Net Flux

The effects of prostaglandin E2 (PGE2) on the transport of sodium and chloride were studied in cultured A6 renal epithelial cells. PGE2 on the basolateral but not the apical surface increased transmonolayer short-circuit current (Isc) and conductance. These changes could not be inhibited with amiloride or furosemide in the apical medium. Flux measurements showed that although Isc and net flux of sodium were equal in unstimulated cells, after addition of PGE2 the current increased with no corresponding changes in bidirectional or net flux of sodium. Immersing the cells in sodium-free or chloride-free media inhibited the effects of PGE2. Measurements of the simultaneous fluxes of sodium and chloride showed that after PGE2 was added there was a net flux of chloride from the basal to the apical side (secretion) that was equal to the change in Isc. The effects of PGE2 were inhibited by furosemide in the basal medium. We conclude that PGE2 stimulates a process of chloride secretion in A6 cells.

scholarly journals Sorbin and SH3 domain-containing protein 2 (SORBS2) is a component of the acto-myosin ring at the apical junctional complex in epithelial cells

PLoS ONE ◽  
2017 ◽  
Vol 12 (9) ◽  
pp. e0185448 ◽  
Author(s):  
Karin Fredriksson-Lidman ◽  
Christina M. Van Itallie ◽  
Amber J. Tietgens ◽  
James M. Anderson
Keyword(s):  
Epithelial Cells ◽  
Sh3 Domain ◽  
Junctional Complex ◽  
Apical Junctional Complex

Dengue virus enters and exits epithelial cells through both apical and basolateral surfaces and perturbs the apical junctional complex

2018 ◽  
Vol 258 ◽  
pp. 39-49
Author(s):  
Liliana Ramirez ◽  
Abigail Betanzos ◽  
Arturo Raya-Sandino ◽  
Lorenza González-Mariscal ◽  
Rosa M. del Angel
Keyword(s):  
Epithelial Cells ◽  
Dengue Virus ◽  
Junctional Complex ◽  
Apical Junctional Complex

scholarly journals Rearrangement of the keratin cytoskeleton after combined treatment with microtubule and microfilament inhibitors.

1983 ◽  
Vol 97 (6) ◽  
pp. 1788-1794 ◽  
Author(s):  
L W Knapp ◽  
W M O'Guin ◽  
R H Sawyer
Keyword(s):  
Epithelial Cells ◽  
Structural Organization ◽  
Immunofluorescence Microscopy ◽  
Combined Treatment ◽  
Cytoskeletal Organization ◽  
Functional Roles ◽  
Indirect Immunofluorescence Microscopy ◽  
Keratin Filaments ◽  
Keratin Filament ◽  
Keratin Fibers

In addition to containing microtubule and microfilament systems, vertebrate epithelial cells contain an elaborate keratin intermediate-filament cytoskeleton. Little is known about its structural organization or function. Using indirect immunofluorescence microscopy with an antikeratin antiserum probe, we found that destabilization of microtubules and microfilaments with cytostatic drugs induces significant alterations in the cytoskeletal organization of keratin filaments in HeLa and fetal mouse epidermal cells. Keratin filament organization was observed to undergo a rapid (1-2 h) transition from a uniform distribution to an open lattice of keratin fibers stabilized by membrane-associated focal centers. Since addition of any one drug alone did not elicit significant organizational change in the keratin cytoskeleton, we suggest that microfilaments and microtubules have a combined role in maintaining the arrangement of keratin in these cells. Vimentin filaments, the only other intermediate-sized filaments found in HeLa cells, did not co-distribute with keratin in untreated or drug-treated cells. These findings offer a new way to approach the study of the dynamics and functional roles of the keratin cytoskeleton in epithelial cells.

Novel Electron Tomographic Methods to Study the Morphology of Keratin Filament Networks

2010 ◽  
Vol 16 (4) ◽  
pp. 462-471 ◽  
Author(s):  
Michaela Sailer ◽  
Katharina Höhn ◽  
Sebastian Lück ◽  
Volker Schmidt ◽  
Michael Beil ◽  
...  
Keyword(s):  
High Pressure ◽  
Low Voltage ◽  
Signal To Noise Ratio ◽  
3D Structure ◽  
Signal To Noise ◽  
Freeze Dried ◽  
Keratin Filaments ◽  
Keratin Filament ◽  
Noise Ratio ◽  
Filament Network

AbstractThe three-dimensional (3D) keratin filament network of pancreatic carcinoma cells was investigated with different electron microscopical approaches. Semithin sections of high-pressure frozen and freeze substituted cells were analyzed with scanning transmission electron microscope (STEM) tomography. Preservation of subcellular structures was excellent, and keratin filaments could be observed; however, it was impossible to three-dimensionally track the individual filaments. To obtain a better signal-to-noise ratio in transmission mode, we observed ultrathin sections of high-pressure frozen and freeze substituted samples with low-voltage (30 kV) STEM. Contrast was improved compared to 300 kV, and individual filaments could be observed. The filament network of samples prepared by detergent extraction was imaged by high-resolution scanning electron microscopy (SEM) with very good signal-to-noise ratio using the secondary electron signal and the 3D structure could be elucidated by SEM tomography. In freeze-dried samples it was possible to discern between keratin filaments and actin filaments because the helical arrangement of actin subunits in the F-actin could be resolved. When comparing the network structures of the differently prepared samples, we found no obvious differences in filament length and branching, indicating that the intermediate filament network is less susceptible to preparation artifacts than the actin network.

scholarly journals Rho/Rho-associated Kinase-II Signaling Mediates Disassembly of Epithelial Apical Junctions

2007 ◽  
Vol 18 (9) ◽  
pp. 3429-3439 ◽  
Author(s):  
Stanislav N. Samarin ◽  
Andrei I. Ivanov ◽  
Gilles Flatau ◽  
Charles A. Parkos ◽  
Asma Nusrat
Keyword(s):  
Epithelial Cells ◽  
Rho Gtpase ◽  
Critical Role ◽  
Junctional Complex ◽  
Nucleotide Exchange ◽  
Calcium Depletion ◽  
Pharmacological Inhibition ◽  
Apical Junctional Complex ◽  
Apical Junctions

Apical junctional complex (AJC) plays a vital role in regulation of epithelial barrier function. Disassembly of the AJC is observed in diverse physiological and pathological states; however, mechanisms governing this process are not well understood. We previously reported that the AJC disassembly is driven by the formation of apical contractile acto-myosin rings. In the present study, we analyzed the signaling pathways regulating acto-myosin–dependent disruption of AJC by using a model of extracellular calcium depletion. Pharmacological inhibition analysis revealed a critical role of Rho-associated kinase (ROCK) in AJC disassembly in calcium-depleted epithelial cells. Furthermore, small interfering RNA (siRNA)-mediated knockdown of ROCK-II, but not ROCK-I, attenuated the disruption of the AJC. Interestingly, AJC disassembly was not dependent on myosin light chain kinase and myosin phosphatase. Calcium depletion resulted in activation of Rho GTPase and transient colocalization of Rho with internalized AJC proteins. Pharmacological inhibition of Rho prevented AJC disassembly. Additionally, Rho guanine nucleotide exchange factor (GEF)-H1 translocated to contractile F-actin rings after calcium depletion, and siRNA-mediated depletion of GEF-H1 inhibited AJC disassembly. Thus, our findings demonstrate a central role of the GEF-H1/Rho/ROCK-II signaling pathway in the disassembly of AJC in epithelial cells.

scholarly journals Defining interactions and distributions of cadherin and catenin complexes in polarized epithelial cells.

1994 ◽  
Vol 125 (6) ◽  
pp. 1341-1352 ◽  
Author(s):  
I S Näthke ◽  
L Hinck ◽  
J R Swedlow ◽  
J Papkoff ◽  
W J Nelson
Keyword(s):  
Epithelial Cells ◽  
Functional Organization ◽  
Insoluble Fraction ◽  
Junctional Complex ◽  
Beta Catenin ◽  
Lateral Membrane ◽  
Apical Junctional Complex ◽  
Polarized Epithelial Cells ◽  
Triton X ◽  
E Cadherin

The cadherin/catenin complex plays important roles in cell adhesion, signal transduction, as well as the initiation and maintenance of structural and functional organization of cells and tissues. In the preceding study, we showed that the assembly of the cadherin/catenin complex is temporally regulated, and that novel combinations of catenin and cadherin complexes are formed in both Triton X-100-soluble and -insoluble fractions; we proposed a model in which pools of catenins are important in regulating assembly of E-cadherin/catenin and catenin complexes. Here, we sought to determine the spatial distributions of E-cadherin, alpha-catenin, beta-catenin, and plakoglobin, and whether different complexes of these proteins accumulate at steady state in polarized Madin-Darby canine kidney cells. Protein distributions were visualized by wide field, optical sectioning, and double immunofluorescence microscopy, followed by reconstruction of three-dimensional images. In cells that were extracted with Triton X-100 and then fixed (Triton X-100-insoluble fraction), more E-cadherin was concentrated at the apical junction relative to other areas of the lateral membrane. alpha-Catenin and beta-catenin colocalize with E-cadherin at the apical junctional complex. There is some overlap in the distribution of these proteins in the lateral membrane, but there are also areas where the distributions are distinct. Plakoglobin is excluded from the apical junctional complex, and its distribution in the lateral membrane is different from that of E-cadherin. Cells were also fixed and then permeabilized to reveal the total cellular pool of each protein (Triton X-100-soluble and -insoluble fractions). This analysis showed lateral membrane localization of alpha-catenin, beta-catenin, and plakoglobin, and it also revealed that they are distributed throughout the cell. Chemical cross-linking of proteins and analysis with specific antibodies confirmed the presence at steady state of E-cadherin/catenin complexes containing either beta-catenin or plakoglobin, and catenin complexes devoid of E-cadherin. Complexes containing E-cadherin/beta-catenin and E-cadherin/alpha-catenin are present in both the Triton X-100-soluble and -insoluble fractions, but E-cadherin/plakoglobin complexes are not detected in the Triton X-100-insoluble fraction. Taken together, these results show that different complexes of cadherin and catenins accumulate in fully polarized epithelial cells, and that they distribute to different sites. We suggest that cadherin/catenin and catenin complexes at different sites have specialized roles in establishing and maintaining the structural and functional organization of polarized epithelial cells.

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