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Role of Phosphorylation in Moesin Interactions with PIP 2 -Containing Biomimetic Membranes

Abstract : Moesin, a protein of the ezrin, radixin, and moesin family, which links the plasma membrane to the cytoskeleton, is involved in multiple physiological and pathological processes, including viral budding and infection. Its interaction with the plasma membrane occurs via a key phosphoinositide, the phosphatidyl(4,5)inositol-bisphosphate (PIP2), and phosphorylation of residue T558, which has been shown to contribute, in cellulo, to a conformationally open protein. We study the impact of a double phosphomimetic mutation of moesin (T235D, T558D), which mimics the phosphorylation state of the protein, on protein/PIP2/microtubule interactions. Analytical ultracentrifugation in the micromolar range showed moesin in the monomer and dimer forms, with wild-type (WT) moesin containing a slightly larger fraction (∼30%) of dimers than DD moesin (10-20%). Only DD moesin was responsive to PIP2 in its micellar form. Quantitative cosedimentation assays using large unilamellar vesicles and quartz crystal microbalance on supported lipid bilayers containing PIP2 reveal a specific cooperative interaction for DD moesin with an ability to bind two PIP2 molecules simultaneously, whereas WT moesin was able to bind only one. In addition, DD moesin could subsequently interact with microtubules, whereas WT moesin was unable to do so. Altogether, our results point to an important role of these two phosphorylation sites in the opening of moesin: since DD moesin is intrinsically in a more open conformation than WT moesin, this intermolecular interaction is reinforced by its binding to PIP2. We also highlight important differences between moesin and ezrin, which appear to be finely regulated and to exhibit distinct molecular behaviors.
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https://hal.univ-grenoble-alpes.fr/hal-01682169
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Soumis le : vendredi 12 janvier 2018 - 09:38:41
Dernière modification le : mardi 6 octobre 2020 - 16:12:09

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Quentin Lubart, Helene Vitet, Fabien Dalonneau, Aline Le Roy, Mathieu Kowalski, et al.. Role of Phosphorylation in Moesin Interactions with PIP 2 -Containing Biomimetic Membranes. Biophysical Journal, Biophysical Society, 2018, 114 (1), pp.98 - 112. ⟨10.1016/j.bpj.2017.10.041⟩. ⟨hal-01682169⟩

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