Two-sided ubiquitin binding of NF-κB essential modulator (NEMO) zinc finger unveiled by a mutation associated with anhidrotic ectodermal dysplasia with immunodeficiency syndrome
In: Journal of Biological Chemistry Journal of Biological Chemistry, 2013, 288 (47), pp.33722-37. ⟨10.1074/jbc.M113.483305⟩ Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology, 2013, 288 (47), pp.33722-37. ⟨10.1074/jbc.M113.483305⟩; (2013-11-22)
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International audience; Hypomorphic mutations in the X-linked human NEMO gene result in various forms of anhidrotic ectodermal dysplasia with immunodeficiency. NEMO function is mediated by two distal ubiquitin binding domains located in the regulatory C-terminal domain of the protein: the coiled-coil 2-leucine zipper (CC2-LZ) domain and the zinc finger (ZF) domain. Here, we investigated the effect of the D406V mutation found in the NEMO ZF of an ectodermal dysplasia with immunodeficiency patients. This point mutation does not impair the folding of NEMO ZF or mono-ubiquitin binding but is sufficient to alter NEMO function, as NEMO-deficient fibroblasts and Jurkat T lymphocytes reconstituted with full-length D406V NEMO lead to partial and strong defects in NF-κB activation, respectively. To further characterize the ubiquitin binding properties of NEMO ZF, we employed di-ubiquitin (di-Ub) chains composed of several different linkages (Lys-48, Lys-63, and linear (Met-1-linked)). We showed that the pathogenic mutation preferentially impairs the interaction with Lys-63 and Met-1-linked di-Ub, which correlates with its ubiquitin binding defect in vivo. Furthermore, sedimentation velocity and gel filtration showed that NEMO ZF, like other NEMO related-ZFs, binds mono-Ub and di-Ub with distinct stoichiometries, indicating the presence of a new Ub site within the NEMO ZF. Extensive mutagenesis was then performed on NEMO ZF and characterization of mutants allowed the proposal of a structural model of NEMO ZF in interaction with a Lys-63 di-Ub chain.
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Two-sided ubiquitin binding of NF-κB essential modulator (NEMO) zinc finger unveiled by a mutation associated with anhidrotic ectodermal dysplasia with immunodeficiency syndrome
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Autor/in / Beteiligte Person: | Fran ccedilois Traincard ; Fontan, Elisabeth ; Agou, Fabrice ; Raynal, Bertrand ; Ngadjeua, Flora ; Chiaravalli, Jeanne ; Biochimie structurale et cellulaire ; Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS) ; Université Pierre et Marie Curie - Paris 6 - UFR de Médecine Pierre et Marie Curie (UPMC) ; Université Pierre et Marie Curie - Paris 6 (UPMC) ; Biophysique des macromolécules et leurs interactions ; This work was supported, in whole or in part, by Foundation BNP Paribas and the Institut de Recherches SERVIER (Croissy sur Seine, France). ; Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS) |
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Quelle: | Journal of Biological Chemistry Journal of Biological Chemistry, 2013, 288 (47), pp.33722-37. ⟨10.1074/jbc.M113.483305⟩ Journal of Biological Chemistry, American Society for Biochemistry and Molecular Biology, 2013, 288 (47), pp.33722-37. ⟨10.1074/jbc.M113.483305⟩; (2013-11-22) |
Veröffentlichung: | HAL CCSD, 2013 |
Medientyp: | unknown |
ISSN: | 0021-9258 (print) ; 1083-351X (print) |
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