Negative regulation of EB1 turnover at microtubule plus ends by interaction with microtubule-associated protein ATIP3

// Lauriane Velot 1,2,3,* , Angie Molina 1,2,3,* , Sylvie Rodrigues-Ferreira 1,2,3 , Anne Nehlig 1,2 , Benjamin Pierre Bouchet 4 , Marina Morel 3 , Ludovic Leconte 5 , Laurence Serre 6 , Isabelle Arnal 6 , Diane Braguer 7,8 , Ariel Savina 9 , Stephane Honore 7,8 and Clara Nahmias 1,2,3 1 Inserm U981, Institut Gustave Roussy Department of Molecular Medicine, Villejuif, France 2 Universite Paris-Saclay, Villejuif, France 3 CNRS UMR8104, Institut Cochin, Paris, France 4 Cell Biology, Faculty of Science, Utrecht University, Padualaan, CH Utrecht, The Netherlands 5 Cell and Tissue Imaging Core Facilty, PICT-IBiSA, CNRS UMR144 Institut Curie, Centre de Recherche, Paris, France 6 Inserm U836, Grenoble Institut des Neurosciences, Grenoble, France 7 Aix Marseille Universite, Inserm, CRO2 UMR_S 911, Marseille, France 8 APHM, Hopital Timone, Marseille, France 9 Scientific Partnerships Roche SAS, Boulogne Billancourt, France * These authors have contributed equally to this work Correspondence to: Clara Nahmias, email: // Keywords : EB1, MTUS1, protein interaction, +TIP, microtubule dynamics Received : May 07, 2015 Accepted : October 14, 2015 Published : October 20, 2015 Abstract The regulation of microtubule dynamics is critical to ensure essential cell functions. End binding protein 1 (EB1) is a master regulator of microtubule dynamics that autonomously binds an extended GTP/GDP-Pi structure at growing microtubule ends and recruits regulatory proteins at this location. However, negative regulation of EB1 association with growing microtubule ends remains poorly understood. We show here that microtubule-associated tumor suppressor ATIP3 interacts with EB1 through direct binding of a non-canonical proline-rich motif. Results indicate that ATIP3 does not localize at growing microtubule ends and that in situ ATIP3-EB1 molecular complexes are mostly detected in the cytosol. We present evidence that a minimal EB1-interacting sequence of ATIP3 is both necessary and sufficient to prevent EB1 accumulation at growing microtubule ends in living cells and that EB1-interaction is involved in reducing cell polarity. By fluorescence recovery of EB1-GFP after photobleaching, we show that ATIP3 silencing accelerates EB1 turnover at microtubule ends with no modification of EB1 diffusion in the cytosol. We propose a novel mechanism by which ATIP3-EB1 interaction indirectly reduces the kinetics of EB1 exchange on its recognition site, thereby accounting for negative regulation of microtubule dynamic instability. Our findings provide a unique example of decreased EB1 turnover at growing microtubule ends by cytosolic interaction with a tumor suppressor.