Proteolytic E-cadherin activation followed by solution NMR and X-ray crystallography

Häußinger, Daniel; Ahrens, Tom; Aberle, Thomas; Engel, Jürgen; Stetefeld, Jörg; Grzesiek, Stephan

doi:10.1038/sj.emboj.7600192

Cited by 139 publications

(196 citation statements)

References 32 publications

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“…Despite uncertainties as to the physical basis for the role of Trp2, this residue provides a clear sequence marker for EC1 domains. 21 have found that the dimerization of E-cadherin EC1-EC2 domain constructs is enhanced by Ca 2+ binding, suggesting a possible role in the swapping process; however, the molecular mechanism through which this might occur is still obscure.…”

Section: Discussionmentioning

confidence: 99%

“…The three-dimensional structures of a number of type I and type II cadherin ectodomain adhesive regions have been determined by both X-ray crystallography and NMR, and the structure for the full ectodomain of Xenopus laevis C-cadherin has also been determined [13][14][15][16][17][18][19][20][21][22][23] . Table 1 contains a list of all available cadherin structures 13-25 .…”

Section: Introductionmentioning

confidence: 99%

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Sequence and Structural Determinants of Strand Swapping in Cadherin Domains: Do All Cadherins Bind Through the Same Adhesive Interface?

Posy

Shapiro

Honig

2008

Journal of Molecular Biology

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SUMMARYCadherins are cell surface adhesion proteins important for tissue development and integrity. Type I and type II, or "classical", cadherins form adhesive dimers via an interface formed through the exchange, or "swapping", of the N-terminal β-strands from their membrane-distal EC1 domains. Here we ask which sequence and structural features in EC1 domains are responsible for β-strand swapping and whether members of other cadherin families also form similar strand-swapped binding interfaces. We first create a comprehensive database consisting of multiple alignments of each type of cadherin domain. We use the known three-dimensional structures of classical cadherins to identify conserved positions in multiple sequence alignments that appear to be crucial determinants of the cadherin domain structure. We further identify features that are unique to EC1 domains. On the basis of our analysis we conclude that all cadherin domains have very similar overall folds but, with the exception of classical and desmosomal cadherin EC1 domains, most of them do not appear to bind through a strand swapping mechanism. Thus, non-classical cadherins that function in adhesion are likely to use different protein-protein interaction interfaces. Our results have implications for the evolution of molecular mechanisms of cadherin-mediated adhesion in vertebrates.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Sequence and Structural Determinants of Strand Swapping in Cadherin Domains: Do All Cadherins Bind Through the Same Adhesive Interface?

Posy

Shapiro

Honig

2008

Journal of Molecular Biology

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“…Electron microscopy (12,13,24,31), X-ray crystallography (10,11,15,22), NMR (26,32), mutational experiments (22), and domain swapping experiments (33) have implicated the EC1 domain in cadherin trans interactions. However, direct force measurements (16)(17)(18) supported by cell attachment and bead binding assays (19) showed 3 distinct adhesive alignments interpreted as the overlap of opposing EC1-5, EC1-3, and EC1 domains.…”

mentioning

confidence: 99%

“…Models of cadherin cis dimerization are based on indirect evidence from the packing interactions in cadherin crystal structures (11,15,22,23), electron tomographs of desmosomes (12, 13), electron micrographs of E-cadherin fusion constructs (24), and chemical cross-linking and gel filtration of cadherin extracted from cells (25). Although the cadherin cis interaction site was initially localized to the EC1 domains (10, 24), it was later proposed that EC1 and EC2 domains of neighboring cadherin molecules are involved in these interactions (11,26). However, other studies using protein cross-linking and coimmunoprecipitation demonstrated that cis cadherin dimers were formed only in cells grown artificially at low calcium concentrations, and that cis and trans interactions shared the same adhesive interface (27-29).…”

mentioning

confidence: 99%

Resolving cadherin interactions and binding cooperativity at the single-molecule level

Zhang¹,

Sivasankar

Nelson

et al. 2009

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

176

195

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The cadherin family of Ca 2؉ -dependent cell adhesion proteins are critical for the morphogenesis and functional organization of tissues in multicellular organisms, but the molecular interactions between cadherins that are at the core of cell-cell adhesion are a matter of considerable debate. A widely-accepted model is that cadherins adhere in 3 stages. First, the functional unit of cadherin adhesion is a cis dimer formed by the binding of the extracellular regions of 2 cadherins on the same cell surface. Second, formation of low-affinity trans interactions between cadherin cis dimers on opposing cell surfaces initiates cell-cell adhesion. Third, lateral clustering of cadherins cooperatively strengthens intercellular adhesion. Evidence of these cadherin binding states during adhesion is, however, contradictory, and evidence for cooperativity is lacking. We used single-molecule structural (fluorescence resonance energy transfer) and functional (atomic force microscopy) assays to demonstrate directly that cadherin monomers interact via their N-terminal EC1 domain to form trans adhesive complexes. We could not detect the formation of cadherin cis dimers, but found that increasing the density of cadherin monomers cooperatively increased the probability of trans adhesive binding.atomic force microscope ͉ cell adhesion ͉ cis dimer ͉ fluorescence resonance energy transfer ͉ trans binding C adherins are Ca 2ϩ -dependent cell-cell adhesion proteins that play fundamental roles in the functional organization of cells and in maintaining the structural integrity of solid tissues (1, 2). Cadherin adhesion is modified during normal embryonic development, and disruption of adhesion is common in metastatic cancers (2-4). Despite detailed studies of cadherinmediated adhesion in multicellular organisms, a molecular understanding of the adhesive states of cadherin is less clear (5). Structural studies have shown that cadherin-cadherin contacts are mediated by the extracellular domain comprised of tandem repeats of 5 cadherin (EC) domains. A widely accepted model as summarized in textbooks (6) and recent review articles (1, 7-9) is that the functional unit of cadherin adhesion is a cis dimer formed by binding of the extracellular domains of 2 cadherins on the same cell surface. Cell-cell adhesion is initiated by the formation of trans adhesive complexes between the EC1 domains of cadherin cis dimers on opposing cell surfaces (1, 6-15). Strong cadherin adhesion may also require trans binding along additional EC domains (16)(17)(18)(19). Trans-cadherin binding is a lowaffinity interaction, but cell-cell adhesion is believed to be enhanced cooperatively by the lateral clustering of cadherins (20,21).Evidence for these trans-and cis-cadherin binding states, however, is controversial, and evidence for cooperativity in promoting adhesion is lacking. Models of cadherin cis dimerization are based on indirect evidence from the packing interactions in cadherin crystal structures (11,15,22,23), electron tomographs of desmosomes (12, 13), elect...

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“…The first conformation, called an X-dimer, is formed by extensive surface interactions via the two outer domains (EC1-2) of the cadherin EC region 6,9 . The second conformation, called a strand-swap dimer, is formed when the side chain of a conserved Tryptophan at position 2 (W2) is inserted into a pocket on the adhesive partner 7,11 .…”

mentioning

confidence: 99%

Resolving the molecular mechanism of cadherin catch bond formation

et al. 2014

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Classical cadherin Ca 2 þ -dependent cell-cell adhesion proteins play key roles in embryogenesis and in maintaining tissue integrity. Cadherins mediate robust adhesion by binding in multiple conformations. One of these adhesive states, called an X-dimer, forms catch bonds that strengthen and become longer lived in the presence of mechanical force. Here we use single-molecule force-clamp spectroscopy with an atomic force microscope along with molecular dynamics and steered molecular dynamics simulations to resolve the molecular mechanisms underlying catch bond formation and the role of Ca 2 þ ions in this process. Our data suggest that tensile force bends the cadherin extracellular region such that they form long-lived, force-induced hydrogen bonds that lock X-dimers into tighter contact. When Ca 2 þ concentration is decreased, fewer de novo hydrogen bonds are formed and catch bond formation is eliminated.

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Proteolytic E-cadherin activation followed by solution NMR and X-ray crystallography

Cited by 139 publications

References 32 publications

Sequence and Structural Determinants of Strand Swapping in Cadherin Domains: Do All Cadherins Bind Through the Same Adhesive Interface?

Sequence and Structural Determinants of Strand Swapping in Cadherin Domains: Do All Cadherins Bind Through the Same Adhesive Interface?

Resolving cadherin interactions and binding cooperativity at the single-molecule level

Resolving the molecular mechanism of cadherin catch bond formation

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