Cadherins are holding hands to resist

Cadherins are the major component of adherens junctions, holding cells together by forming 'trans' interactions with cadherin molecules in the plasma membrane of adjacent cells. Crystal structures of cadherin indicate that the protein can also form 'cis' interactions with E-cadherin molecules in the same cell membrane, but whether these interactions help organize cadherin in vivo, and how this might affect the function of adherens junctions, remains unclear.

HFSP Program Grant holders René Marc Mège and Benoit Ladoux and colleagues
authored on Mon, 24 August 2015

In an article published in the Journal of Cell Biology in July, the teams of Benoit Ladoux and René Marc Mège at the Institut Jacques Monod in Paris reveal that interactions between neighboring cadherin molecules help to strengthen their connection to the actin cytoskeleton and stabilize cell-cell contacts.

Together with their collaborators, they visualize for the first time the formation of organized cadherin nanoclusters in the plasma membrane. This oligomerization of cadherins, made possible by cis interactions of their extracellular domain, is responsible for the anchorage of cadherins to the actin and controls the fluidity of the intercellular contact.

Around 35% of proteins are in an oligomeric state. Oligomerization provides several functional advantages such as a mechanism to resist against degradation and more importantly makes higher order long-living subcellular structures such as cytoskeletal filaments and functional nanomachines.

Figure a: Experimental approach. Left: transfected E-cadherin-deficient cells were seeded at low density on Ecad-Fc coated EM grids. After 2 hours, the upper part of the cells was ripped off, leaving plasma membrane sheets corresponding to the bottom part of the cells on the EM grids. Grids were then incubated with 5-7 nm diameter mix-coated gold nanoparticles (in red) conjugated to a single anti-GFP nanobody (in green) and observed by TEM.

Figure b: TEM visualization of anti-GFP NP on plasma membranes sheets of wt Ecad-GFP expressing cells spread on Ecad-Fc. Images (100 x 100 nm, scale bar: 15 nm).

Tissue cohesion is ensured by cell adhesion molecules that establish short lived intercellular protein-protein bonds at the single molecule level. Oligomerization could provide the necessary strength to support intercellular adhesion and resistance to mechanical stress.

This is the case for cadherins too. Cadherins are major cell adhesion molecules in animal cells. Cadherins diffusing at the plasma membrane initiate cell-cell interactions by establishing homophilic intercellular bonds. These trans interactions analyzed by atomic force microscopy or biomembrane force probe have been shown to be short lived, implying that some higher order processes must take place in order that cadherin-mediated adhesion reaches sufficient stability to sustain physiological relevant resistance to mechanical stress.

Recent biochemical and structural data suggest the existence of cis interactions between cadherin carried by the same cell; the combination of cis and trans interactions leading potentially to the formation of organized nanoclusters of cadherin within adherens junctions. Through a combination of molecular and cellular biology, electron microscopy, imaging and biophysics approaches, the team shows that cadherins organize indeed in nanoclusters in the cell membrane with characteristic dimensions predicted by ultrastructural studies (Figures a,b). This oligomerization has major consequences on the relative stabilization of cadherins but leads to a major increase in the functional anchoring of adhesion complexes to actin filaments and ultimately strengthens junctions that limit cell-cell contact fluidity and increases cell cohesion and the collective behavior of migrating cells (Figures c,d).

Figure c: Quantitative analysis of cell layer expansion: Transfected wt Ecad-GFP and cis Ecad-GFP (mutated in 2 amino acids preventing the formation of the cis interface) cells were grown in a dish where part of the culture area was initially masked by an inert PDMS block. When confluence was reached the block was removed freeing new space. After removal of the block, cell monolayer expansion was imaged over time. PIV analysis of wt Ecad and cis Ecad collective cell migration allows velocity fields (not shown) and order parameters to be extracted. Upper panel:  heat maps of the order parameter (-1 means backward, +1 means forward movements); lower panel: order parameters in the velocity field as a function of distance to the front.

Figure d: Schematics of the cis interface-dependent oligomerization of cadherin ectodomain and its effect on F-actin anchoring. The slight stabilization of cell-cell contacts brought by cadherin oligomerization in ordered clusters has strong influence on the anchoring of these clusters to the actomyosin network.

Thus, our results provide direct evidence that cadherin oligomerization indeed stabilizes cell-cell contacts providing the necessary strength to maintain tissue cohesion. Whether this pathway is modulated in vivo during collective cell migration, cell intercalation, wound healing or cancer cell dissemination by factors such as cadherin glycosylation, cadherin and catenin phosphorylation or other post-translational modifications remains to be investigated.

Together with a recent paper describing the molecular mechanism of action of the mechano-sensor responsible for the tension-dependent anchoring of cadherin to actin, α-catenin at the single molecule level (Yao et al., Nat Commun. 2015), the new findings produced by the consortium further advance the mechanisms responsible for mechano-transduction by cell-cell junctions at the nano- and micro-scales, as planned in the initial collaborative grant and provide as well correlation with cell and tissue-scale behavior.

Reference

The formation of ordered nanoclusters controls cadherin anchoring to actin and cell-cell contact fluidity. Strale PO, Duchesne L, Peyret G, Montel L, Nguyen T, Png E, Tampé R, Troyanovsky S, Hénon S, Ladoux B, Mège RM. J Cell Biol. 2015 Jul 20;210(2):333-46. doi: 10.1083/jcb.201410111.

Pubmed link

Other reference

Force-dependent conformational switch of α-catenin controls vinculin binding. Yao M, Qiu W, Liu R, Efremov AK, Cong P, Seddiki R, Payre M, Lim CT, Ladoux B, Mège RM, Yan J. 2014 Jul 31; Nat Commun. 5:4525. doi: 10.1038/ncomms5525.

Pubmed link