Ca<sup>2+</sup>Dependency of N-Cadherin Function Probed by Laser Tweezer and Atomic Force Microscopy

Baumgärtner, Werner; Golenhofen, Nikola; Grundhöfer, Niko; Wiegand, Johannes; Drenckhahn, Detlev

doi:10.1523/jneurosci.23-35-11008.2003

Cited by 59 publications

(67 citation statements)

References 58 publications

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“…The order of homophilic adhesion is C͞C Ͼ E͞E Ͼ N͞N, in qualitative agreement with cell adhesion and single bond rupture measurements (11,12,20,(26)(27)(28).…”

Section: Discussionsupporting

confidence: 77%

Similarities between heterophilic and homophilic cadherin adhesion

Prakasam

Maruthamuthu

Leckband³

2006

Proc. Natl. Acad. Sci. U.S.A.

102

107

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The mechanism that drives the segregation of cells into tissuespecific subpopulations during development is largely attributed to differences in intercellular adhesion. This process requires the cadherin family of calcium-dependent glycoproteins. A widely held view is that protein-level discrimination between different cadherins on cell surfaces drives this sorting process. Despite this postulated molecular selectivity, adhesion selectivity has not been quantitatively verified at the protein level. In this work, molecular force measurements and bead aggregation assays tested whether differences in cadherin bond strengths could account for cell sorting in vivo and in vitro. Studies were conducted with chicken N-cadherin, canine E-cadherin, and Xenopus C-cadherin. Both qualitative bead aggregation and quantitative force measurements show that the cadherins cross-react. Furthermore, heterophilic adhesion is not substantially weaker than homophilic adhesion, and the measured differences in adhesion do not correlate with cell sorting behavior. These results suggest that the basis for cell segregation during morphogenesis does not map exclusively to protein-level differences in cadherin adhesion.cell adhesion ͉ force probe ͉ selectivity ͉ surface forces apparatus B oth the formation of distinct tissues during morphogenesis and the maintenance of adult tissue structure are regulated by adhesive contacts between cells. One family of adhesion proteins, cadherins, is critical to several processes associated with the formation and maintenance of tissues. The genetic regulation of the spatiotemporal expression patterns of cadherins in vivo is believed to play a central role in embryogenesis, cell differentiation, and the maintenance of the multicellular structure of an organism (1-5). One prevalent hypothesis is that differences in adhesion between cells direct cell segregation during morphogenesis, analogous to liquid-liquid phase separation (6). A fundamental unresolved question is whether this sorting out is encoded by protein-specific or cell-specific differences in adhesion. A common hypothesis is that preferential binding between identical cadherins (homophilic) relative to heterophilic binding between dissimilar cadherins induces the cell segregation.Both the adhesive and the selectivity functions of classic cadherins map to the extracellular (EC) region (7), which comprises five tandemly arranged EC domains, EC1-5, numbered from the outermost domain (Fig. 1) (8). Classical cadherins also contain a transmembrane segment and a cytoplasmic domain. Cadherins form multiple bonds that require different EC domains (9-12). A common feature of the classical cadherins is that the N-terminal domains bind by inserting the tryptophan 2 (W2) residue from one protein into a hydrophobic pocket on an opposed N-terminal domain (13-16). However, given the weak adhesion between the outer domains (11, 12), it is questionable whether the tryptophan binding accounts for both cadherin selectivity and robust adhesion.The most extensively ci...

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“…The order of homophilic adhesion is C͞C Ͼ E͞E Ͼ N͞N, in qualitative agreement with cell adhesion and single bond rupture measurements (11,12,20,(26)(27)(28).…”

Section: Discussionsupporting

confidence: 77%

Similarities between heterophilic and homophilic cadherin adhesion

Prakasam

Maruthamuthu

Leckband³

2006

Proc. Natl. Acad. Sci. U.S.A.

102

107

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“…Other AFM studies demonstrated that type II VE-cadherin (cadherin-5) promotes stronger adhesion than type I N-cadherin; however, similar differences were reported in the same study between type I N-and E-cadherin (Panorchan et al, 2006b). No major differences were found between N-and VE-cadherin in another work using AFM and laser tweezers (Baumgartner et al, 2003), whereas dual-pipette assays revealed significantly stronger adhesion of cells expressing transfected type I E-and N-cadherin compared with type II cadherin-7 and OB-cadherin (Chu et al, 2006). One common finding of these studies is the absence of heterotypic interactions between type I and II cadherins (Patel et al, 2006).…”

Section: Discussionsupporting

confidence: 60%

Fibrogenic fibroblasts increase intercellular adhesion strength by reinforcing individual OB-cadherin bonds

Pittet

Lee

Kulik

et al. 2008

Journal of Cell Science

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depolymerization reduces single-bond strength to the level of cadherin constructs missing the cytoplasmic domain. Hence, fibroblasts reinforce intercellular contacts by: (1) switching from N-to OB-cadherin expression; (2) increasing the strength of single-molecule bonds in three distinct steps; and (3) actinpromoted intrinsic activation of cadherin extracellular binding. We propose that this plasticity adapts fibroblast adhesions to the changing mechanical microenvironment of tissue under remodeling. Supplementary material available online at

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“…In addition, experimental measurements in the synaptic cleft during high-frequency stimulation have shown that Ca 2+ concentration may drop to as low as 0.3-0.8 mM (41,42), and Ca 2+ -dependent inhibition of cadherin function has been observed during synaptic plasticity (43). Biophysical measurements of purified cadherin extracellular domains, using laser tweezers, showed that an extracellular drop in Ca 2+ from 1.5 to 0.8 mM is predicted to decrease binding of N-cadherin by ∼40%, whereas a further drop to 0.3 mM would result in a reduction of binding by 85% (44). Thus, the Ca 2+ dependence of cadherin interactions seems to be appropriately tuned to monitor physiological fluctuations of Ca 2+ concentrations.…”

Section: Resultsmentioning

confidence: 99%

Calcium-dependent dynamics of cadherin interactions at cell–cell junctions

Kim

Tai

Mok

et al. 2011

Proc. Natl. Acad. Sci. U.S.A.

145

116

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Cadherins play a key role in the dynamics of cell-cell contact formation and remodeling of junctions and tissues. Cadherincadherin interactions are gated by extracellular Ca 2+ , which serves to rigidify the cadherin extracellular domains and promote trans junctional interactions. Here we describe the direct visualization and quantification of spatiotemporal dynamics of N-cadherin interactions across intercellular junctions in living cells using a genetically encodable FRET reporter system. Direct measurements of transjunctional cadherin interactions revealed a sudden, but partial, loss of homophilic interactions (τ = 1.17 ± 0.06 s ), suggesting two types of native cadherin interactions-one that is rapidly modulated by changes in extracellular Ca 2+ and another with relatively stable adhesive activity that is Ca 2+ independent. The Ca 2+-sensitive dynamics of cadherin interactions were transmitted to the cell interior where β-catenin translocated to N-cadherin at the junction in both cells. These data indicate that cadherins can rapidly convey dynamic information about the extracellular environment to both cells that comprise a junction.cell adhesion | fluorescence resonance energy transfer | trans binding T he junctions between cells are populated with a variety of cell adhesion molecules that drive the recognition, assembly, and dynamics of cell-cell interactions. Parsing the distinct functions of different adhesion molecules has been challenging, in part due to a paucity of truly revealing in vivo assays. Among cell adhesion molecule families, the classic cadherins exhibit a unique dependence on extracellular Ca 2+ to rigidify the extracellular domains and enable trans junctional homophilic interactions. Three Ca 2+ ions bind with different affinities to each of the pockets between cadherin extracellular domains (1, 2). The affinity of the various Ca 2+ binding sites is in the micromolar to millimolar range (3-8), suggesting the possibility that cadherins can respond dynamically to changes in junctional Ca 2+ levels, and by virtue of physical interactions with cytoplasmic molecules, signal information about the junctional status to the cell interior.3D reconstructions of desmosomes by cryoelectron tomography reveal cadherins interact across intercellular interfaces (9, 10), and crystallographic data of several cadherin domains (4, 11, 12) implicate the first two extracellular repeats (ECs 1 and 2) as critical for homophilic interactions. Previous studies have indicated the importance of a highly conserved tryptophan residue (Trp2), present in the first EC domain, for cadherin-dependent adhesion (2, 12). This tryptophan inserts into the hydrophobic pocket of the partner N-cadherin (Ncad) molecule to form a strand-swapped dimeric structure (11, 12). Mutation of Trp2 to an alanine residue (W2A) prevents strand swapping (13) and results in a loss of adhesive function (2, 14, 15). Although extensive work has elucidated the structure (4, 9-14), biochemistry (2, 16) and single-molecule characterization (15, 17) of ...

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Ca²⁺Dependency of N-Cadherin Function Probed by Laser Tweezer and Atomic Force Microscopy

Cited by 59 publications

References 58 publications

Similarities between heterophilic and homophilic cadherin adhesion

Similarities between heterophilic and homophilic cadherin adhesion

Fibrogenic fibroblasts increase intercellular adhesion strength by reinforcing individual OB-cadherin bonds

Calcium-dependent dynamics of cadherin interactions at cell–cell junctions

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Ca2+Dependency of N-Cadherin Function Probed by Laser Tweezer and Atomic Force Microscopy

Cited by 59 publications

References 58 publications

Similarities between heterophilic and homophilic cadherin adhesion

Similarities between heterophilic and homophilic cadherin adhesion

Fibrogenic fibroblasts increase intercellular adhesion strength by reinforcing individual OB-cadherin bonds

Calcium-dependent dynamics of cadherin interactions at cell–cell junctions

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Product

Resources

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Ca²⁺Dependency of N-Cadherin Function Probed by Laser Tweezer and Atomic Force Microscopy