scholarly journals Biochemical and structural analysis of α-catenin in cell–cell contacts

2008 ◽  
Vol 36 (2) ◽  
pp. 141-147 ◽  
Author(s):  
Sabine Pokutta ◽  
Frauke Drees ◽  
Soichiro Yamada ◽  
W. James Nelson ◽  
William I. Weis
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Adherens Junction ◽  
Structural Data ◽  
Cell Contact ◽  
Related Protein ◽  
Imaging Studies ◽  
Actin Networks ◽  
Junction Formation ◽  
Cell Cell

Cadherins are transmembrane adhesion molecules that mediate homotypic cell–cell contact. In adherens junctions, the cytoplasmic domain of cadherins is functionally linked to the actin cytoskeleton through a series of proteins known as catenins. E-cadherin binds to β-catenin, which in turn binds to α-catenin to form a ternary complex. α-Catenin also binds to actin, and it was assumed previously that α-catenin links the cadherin–catenin complex to actin. However, biochemical, structural and live-cell imaging studies of the cadherin–catenin complex and its interaction with actin show that binding of β-catenin to α-catenin prevents the latter from binding to actin. Biochemical and structural data indicate that α-catenin acts as an allosteric protein whose conformation and activity changes depending on whether or not it is bound to β-catenin. Initial contacts between cells occur on dynamic lamellipodia formed by polymerization of branched actin networks, a process controlled by the Arp2/3 (actin-related protein 2/3) complex. α-Catenin can suppress the activity of Arp2/3 by competing for actin filaments. These findings lead to a model for adherens junction formation in which clustering of the cadherin–β-catenin complex recruits high levels of α-catenin that can suppress the Arp2/3 complex, leading to cessation of lamellipodial movement and formation of a stable contact. Thus α-catenin appears to play a central role in cell–cell contact formation.

scholarly journals The formation of ordered nanoclusters controls cadherin anchoring to actin and cell–cell contact fluidity

2015 ◽  
Vol 210 (2) ◽  
pp. 333-346 ◽  
Author(s):  
Pierre-Olivier Strale ◽  
Laurence Duchesne ◽  
Grégoire Peyret ◽  
Lorraine Montel ◽  
Thao Nguyen ◽  
...  
Keyword(s):  
Adherens Junction ◽  
Intercellular Junctions ◽  
Cell Contact ◽  
Collective Cell Migration ◽  
Contact Stability ◽  
Junction Formation ◽  
Mediate Cell ◽  
The Stability ◽  
E Cadherin ◽  
Cell Cell

Oligomerization of cadherins could provide the stability to ensure tissue cohesion. Cadherins mediate cell–cell adhesion by forming trans-interactions. They form cis-interactions whose role could be essential to stabilize intercellular junctions by shifting cadherin clusters from a fluid to an ordered phase. However, no evidence has been provided so far for cadherin oligomerization in cellulo and for its impact on cell–cell contact stability. Visualizing single cadherins within cell membrane at a nanometric resolution, we show that E-cadherins arrange in ordered clusters, providing the first demonstration of the existence of oligomeric cadherins at cell–cell contacts. Studying the consequences of the disruption of the cis-interface, we show that it is not essential for adherens junction formation. Its disruption, however, increased the mobility of junctional E-cadherin. This destabilization strongly affected E-cadherin anchoring to actin and cell–cell rearrangement during collective cell migration, indicating that the formation of oligomeric clusters controls the anchoring of cadherin to actin and cell–cell contact fluidity.

scholarly journals F-pili dynamics by live-cell imaging

2008 ◽  
Vol 105 (46) ◽  
pp. 17978-17981 ◽  
Author(s):  
Margaret Clarke ◽  
Lucinda Maddera ◽  
Robin L. Harris ◽  
Philip M. Silverman
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Recipient Cell ◽  
Cell Communication ◽  
Live Cell ◽  
Cell Contact ◽  
Gram Negative Bacteria ◽  
Donor Cells ◽  
Cell Cell

Bacteria have evolved numerous mechanisms for cell–cell communication, many of which have important consequences for human health. Among these is conjugation, the direct transfer of DNA from one cell to another. For Gram-negative bacteria, conjugation requires thin, flexible filaments (conjugative pili) that are elaborated by DNA donor cells. The structure, function, and especially the dynamics of conjugative pili are poorly understood. Here, we have applied live-cell imaging to characterize the dynamics of F-pili (conjugative pili encoded by the F plasmid of Escherichia coli). We establish that F-pili normally undergo cycles of extension and retraction in the absence of any obvious triggering event, such as contact with a recipient cell. When made, such contacts are able to survive the shear forces felt by bacteria in liquid media. Our data emphasize the role of F-pilus flexibility both in efficiently sampling a large volume surrounding donor cells in liquid culture and in establishing and maintaining cell–cell contact. Additionally and unexpectedly, we infer that extension and retraction are accompanied by rotation about the long axis of the filament.

scholarly journals Dynamics of adherens junctions in epithelial establishment, maintenance, and remodeling

2011 ◽  
Vol 192 (6) ◽  
pp. 907-917 ◽  
Author(s):  
Buzz Baum ◽  
Marios Georgiou
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Adherens Junction ◽  
Dynamic Structure ◽  
Rho Family Gtpases ◽  
Junction Formation ◽  
Rho Family ◽  
And Function ◽  
Adhesive Contacts ◽  
E Cadherin

The epithelial cadherin (E-cadherin)–catenin complex binds to cytoskeletal components and regulatory and signaling molecules to form a mature adherens junction (AJ). This dynamic structure physically connects neighboring epithelial cells, couples intercellular adhesive contacts to the cytoskeleton, and helps define each cell’s apical–basal axis. Together these activities coordinate the form, polarity, and function of all cells in an epithelium. Several molecules regulate AJ formation and integrity, including Rho family GTPases and Par polarity proteins. However, only recently, with the development of live-cell imaging, has the extent to which E-cadherin is actively turned over at junctions begun to be appreciated. This turnover contributes to junction formation and to the maintenance of epithelial integrity during tissue homeostasis and remodeling.

scholarly journals PLAC-24 Is a Cytoplasmic Dynein-Binding Protein That Is Recruited to Sites of Cell-Cell Contact

2002 ◽  
Vol 13 (5) ◽  
pp. 1722-1734 ◽  
Author(s):  
Sher Karki ◽  
Lee A. Ligon ◽  
Jamison DeSantis ◽  
Mariko Tokito ◽  
Erika L. F. Holzbaur
Keyword(s):  
Epithelial Cells ◽  
Recent Observation ◽  
Polyclonal Antibodies ◽  
Adherens Junction ◽  
Cytoplasmic Dynein ◽  
Cell Contact ◽  
Dynein Intermediate Chain ◽  
Cellular Cytoskeleton ◽  
Cell Cell ◽  
Intermediate Chain

We screened for polypeptides that interact specifically with dynein and identified a novel 24-kDa protein (PLAC-24) that binds directly to dynein intermediate chain (DIC). PLAC-24 is not a dynactin subunit, and the binding of PLAC-24 to the dynein intermediate chain is independent of the association between dynein and dynactin. Immunocytochemistry using PLAC-24–specific polyclonal antibodies revealed a punctate perinuclear distribution of the polypeptide in fibroblasts and isolated epithelial cells. However, as epithelial cells in culture make contact with adjacent cells, PLAC-24 is specifically recruited to the cortex at sites of contact, where the protein colocalizes with components of the adherens junction. Disruption of the cellular cytoskeleton with latrunculin or nocodazole indicates that the localization of PLAC-24 to the cortex is dependent on intact actin filaments but not on microtubules. Overexpression of β-catenin also leads to a loss of PLAC-24 from sites of cell-cell contact. On the basis of these data and the recent observation that cytoplasmic dynein is also localized to sites of cell-cell contact in epithelial cells, we propose that PLAC-24 is part of a multiprotein complex localized to sites of intercellular contact that may function to tether microtubule plus ends to the actin-rich cellular cortex.

scholarly journals Direct observation of aggregate-triggered selective autophagy

2021 ◽  
Author(s):  
Anne FJ Janssen ◽  
Giel Korsten ◽  
Wilco Nijenhuis ◽  
Eugene Katrukha ◽  
Lukas Kapitein
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Relative Timing ◽  
Imaging Studies ◽  
Selective Autophagy ◽  
The Core ◽  
Spatiotemporal Regulation ◽  
Core Components ◽  
Inducible Protein

Degradation of aggregates by selective autophagy is important as damaged proteins may impose a threat to cellular homeostasis. Although the core components of the autophagy machinery are well-characterized, the spatiotemporal regulation of many selective autophagy processes, including aggrephagy, remains largely unexplored. Furthermore, because most live-cell imaging studies have so far focused on starvation-induced autophagy, little is known about the dynamics of aggrephagy. Here, we describe the development and application of the mKeima-PIM assay, which enables live-cell observation of autophagic turnover and degradation of inducible protein aggregates in conjunction with key autophagy players. This allowed us to quantify the relative timing and duration of different steps of aggrephagy and revealed the short-lived nature of the autophagosome. The assay furthermore showed the spatial distribution of omegasome formation, highlighting that autophagy initiation is directly instructed by the cargo. Moreover, we found that nascent autophagosomes mostly remain immobile until acidification occurs. Thus, our assay provides new insights into the spatiotemporal regulation and dynamics of aggrephagy.

scholarly journals A lateral protrusion latticework connects neuroepithelial cells and is regulated during neurogenesis

2021 ◽  
Author(s):  
Ioannis Kasioulis ◽  
Alwyn Dady ◽  
John James ◽  
Alan R Prescott ◽  
Pamela A Halley ◽  
...  
Keyword(s):  
Cell Polarity ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Cell Signalling ◽  
Actin Binding ◽  
Cell Contact ◽  
Tissue Integrity ◽  
Neuroepithelial Cells ◽  
Membrane Contacts ◽  
New Form

Dynamic contacts between cells within the developing neuroepithelium are poorly understood but play important roles in cell and tissue morphology and cell signalling. Here, using live-cell imaging and electron microscopy we reveal multiple distinct protrusive structures in chicken neuroepithelial apical endfeet, including sub-apical protrusions that extend laterally within the tissue, and observe similar structures in human neuroepithelium. We characterise the dynamics, shape, and cytoskeleton of these lateral protrusions and distinguish these structures from cytonemes/filopodia and tunnelling nanotubes. We demonstrate that lateral protrusions form a latticework of membrane contacts between non-adjacent cells, depend on actin but not microtubule dynamics and provide a lamellipodial-like platform for further extending fine actin-dependent filipodia. We find that lateral protrusions depend on the actin-binding protein WAVE1: mutant-WAVE1 misexpression attenuated protrusion and generated a round-ended apical endfoot morphology. However, this did not alter apico-basal cell polarity nor reduce tissue integrity. During normal neuronal delamination sub-apical protrusions were withdrawn, but mutant-WAVE1-induced precocious protrusion loss was insufficient to trigger neurogenesis. This study uncovers a new form of cell-cell contact within the developing neuroepithelium regulation of which prefigures neuronal delamination.

scholarly journals Adherens Junction Distribution Mechanisms during Cell-Cell Contact Elongation in Drosophila

PLoS ONE ◽  
2013 ◽  
Vol 8 (11) ◽  
pp. e79613 ◽  
Author(s):  
Gabrielle Goldenberg ◽  
Tony J. C. Harris
Keyword(s):  
Adherens Junction ◽  
Cell Contact ◽  
Cell Cell

scholarly journals Protein zero, a nervous system adhesion molecule, triggers epithelial reversion in host carcinoma cells.

1995 ◽  
Vol 131 (2) ◽  
pp. 465-482 ◽  
Author(s):  
J P Doyle ◽  
J G Stempak ◽  
P Cowin ◽  
D R Colman ◽  
D D'Urso
Keyword(s):  
Plasma Membrane ◽  
Cell Adhesion ◽  
Adhesion Molecule ◽  
Hela Cells ◽  
Adherens Junction ◽  
Cell Contact ◽  
Immunoglobulin Gene ◽  
Epithelial Junctions ◽  
Protein Zero ◽  
Cell Cell

Protein zero (P(o)) is the immunoglobulin gene superfamily glycoprotein that mediates the self-adhesion of the Schwann cell plasma membrane that yields compact myelin. HeLa is a poorly differentiated carcinoma cell line that has lost characteristic morphological features of the cervical epithelium from which it originated. Normally, HeLa cells are not self-adherent. However, when P(o) is artificially expressed in this line, cells rapidly aggregate, and P(o) concentrates specifically at cell-cell contact sites. Rows of desmosomes are generated at these interfaces, the plasma membrane localization of cingulin and ZO-1, proteins that have been shown to be associated with tight junctions, is substantially increased, and cytokeratins coalesce into a cohesive intracellular network. Immunofluorescence patterns for the adherens junction proteins N-cadherin, alpha-catenin, and vinculin, and the desmosomal polypeptides desmoplakin, desmocollin, and desmoglein, are also markedly enhanced at the cell surface. Our data demonstrate that obligatory cell-cell adhesion, which in this case is initially brought about by the homophilic association of P(o) molecules across the intercellular cleft, triggers pronounced augmentation of the normally sluggish or sub-basal cell adhesion program in HeLa cells, culminating in suppression of the transformed state and reversion of the monolayer to an epithelioid phenotype. Furthermore, this response is apparently accompanied by an increase in mRNA and protein levels for desmoplakin and N-cadherin which are normally associated with epithelial junctions. Our conclusions are supported by analyses of ten proteins we examined immunochemically (P(o), cingulin, ZO-1, desmoplakin, desmoglein, desmocollin, N-cadherin, alpha-catenin, vinculin, and cytokeratin-18), and by quantitative polymerase chain reactions to measure relative amounts of desmoplakin and N-cadherin mRNAs. P(o) has no known signaling properties; the dramatic phenotypic changes we observed are highly likely to have developed in direct response to P(o)-induced cell adhesion. More generally, the ability of this "foreign" membrane adhesion protein to stimulate desmosome and adherens junction formation by augmenting well-studied cadherin-based adhesion mechanisms raises the possibility that perhaps any bona fide cell adhesion molecule, when functionally expressed, can engage common intracellular pathways and trigger reversion of a carcinoma to an epithelial-like phenotype.

scholarly journals Integrated control of transporter endocytosis and recycling by the arrestin-related protein Rod1 and the ubiquitin ligase Rsp5

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Michel Becuwe ◽  
Sébastien Léon
Keyword(s):  
Ubiquitin Ligase ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Integrated Control ◽  
Monocarboxylate Transporter ◽  
Related Protein ◽  
Endocytic Sorting ◽  
Lysosomal Targeting ◽  
Golgi Network ◽  
External Signals

After endocytosis, membrane proteins can recycle to the cell membrane or be degraded in lysosomes. Cargo ubiquitylation favors their lysosomal targeting and can be regulated by external signals, but the mechanism is ill-defined. Here, we studied the post-endocytic trafficking of Jen1, a yeast monocarboxylate transporter, using microfluidics-assisted live-cell imaging. We show that the ubiquitin ligase Rsp5 and the glucose-regulated arrestin-related trafficking adaptors (ART) protein Rod1, involved in the glucose-induced internalization of Jen1, are also required for the post-endocytic sorting of Jen1 to the yeast lysosome. This new step takes place at the trans-Golgi network (TGN), where Rod1 localizes dynamically upon triggering endocytosis. Indeed, transporter trafficking to the TGN after internalization is required for their degradation. Glucose removal promotes Rod1 relocalization to the cytosol and Jen1 deubiquitylation, allowing transporter recycling when the signal is only transient. Therefore, nutrient availability regulates transporter fate through the localization of the ART/Rsp5 ubiquitylation complex at the TGN.

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