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Accueil > Départements > Biochimie, Biophysique et Biologie Structurale > Benoit GIGANT & Julie MENETREY : Biochimie Structurale des Microtubules, des Kinésines et de leurs Cargos

Publications de l’équipe


  • T. Q. Nguyen, M. Aumont-Niçaise, J. Andreani, C. Velours, M. Chenon, F. Vilela, C. Geneste, P. F. Varela, P. Llinas, et J. Menetrey, « Characterization of the binding mode of JNK-interacting protein 1 (JIP1) to kinesin-light chain 1 (KLC1) », The Journal of Biological Chemistry, vol. 293, nᵒ 36, p. 13946-13960, juill. 2018.
    Résumé : JIP1 was first identified as scaffold protein for the MAP kinase JNK and is a cargo protein for the kinesin1 molecular motor. JIP1 plays significant and broad roles in neurons, mainly as a regulator of kinesin1-dependent transport, and is associated with human pathologies such as cancer and Alzheimer disease. JIP1 is specifically recruited by the kinesin-light chain 1 (KLC1) of kinesin1, but the details of this interaction are not yet fully elucidated. Here, using calorimetry, we extensively biochemically characterized the interaction between KLC1 and JIP1. Using various truncated fragments of the tetratricopeptide repeat (TPR) domain of KLC1, we narrowed down its JIP1-binding region and identified seven KLC1 residues critical for JIP1 binding. These ITC-based binding data enabled us to footprint the JIP1-binding site on KLC1-TPR. This footprint was used to uncover the structural basis for the marginal inhibition of JIP1 binding by the autoinhibitory LFP-acidic motif of KLC1, as well as for the competition between JIP1 and another cargo protein of kinesin1, the W-acidic motif-containing Alcadein-α. Also, we examined the role of each of these critical residues of KLC1 for JIP1 binding in the light of the previously reported crystal structure of the KLC1-TPR:JIP1 complex. Finally, sequence search in eukaryotic genomes identified several proteins, among which SH2D6 that exhibit similar motif to the KLC1-binding motif of JIP1. Overall, our extensive biochemical characterization of the KLC:JIP1 interaction, as well as identification of potential KLC1-binding partners improve the understanding of how this growing family of cargos is recruited to kinesin1 by KLC1.
    Mots-clés : Alcadein, AMIG, B3S, JNK-interacting protein 1, MIKICA, MST, PF, PIM, SH2D6, TorsinA, TPR domain, Y-acidic motif.

  • T. Q. Nguyen, M. Chenon, F. Vilela, C. Velours, M. Aumont-Nicaise, J. Andreani, P. F. Varela, P. Llinas, et J. Ménétrey, « Correction: Structural plasticity of the N-terminal capping helix of the TPR domain of kinesin light chain », PloS One, vol. 13, nᵒ 5, p. e0197193, 2018.
    Résumé : [This corrects the article DOI: 10.1371/journal.pone.0186354.].
    Mots-clés : AMIG, B3S, MIKICA, PF, PIM.


  • L. Cao, S. Cantos-Fernandes, et B. Gigant, « The structural switch of nucleotide-free kinesin », Scientific Reports, vol. 7, p. 42558, févr. 2017.

  • T. Q. Nguyen, M. Chenon, F. Vilela, C. Velours, M. Aumont-Nicaise, J. Andreani, P. F. Varela, P. Llinas, et J. Ménétrey, « Structural plasticity of the N-terminal capping helix of the TPR domain of kinesin light chain », PloS One, vol. 12, nᵒ 10, p. e0186354, 2017.
    Résumé : Kinesin1 plays a major role in neuronal transport by recruiting many different cargos through its kinesin light chain (KLC). Various structurally unrelated cargos interact with the conserved tetratricopeptide repeat (TPR) domain of KLC. The N-terminal capping helix of the TPR domain exhibits an atypical sequence and structural features that may contribute to the versatility of the TPR domain to bind different cargos. We determined crystal structures of the TPR domain of both KLC1 and KLC2 encompassing the N-terminal capping helix and show that this helix exhibits two distinct and defined orientations relative to the rest of the TPR domain. Such a difference in orientation gives rise, at the N-terminal part of the groove, to the formation of one hydrophobic pocket, as well as to electrostatic variations at the groove surface. We present a comprehensive structural analysis of available KLC1/2-TPR domain structures that highlights that ligand binding into the groove can be specific of one or the other N-terminal capping helix orientations. Further, structural analysis reveals that the N-terminal capping helix is always involved in crystal packing contacts, especially in a TPR1:TPR1' contact which highlights its propensity to be a protein-protein interaction site. Together, these results underline that the structural plasticity of the N-terminal capping helix might represent a structural determinant for TPR domain structural versatility in cargo binding.
    Mots-clés : AMIG, Amino Acid Motifs, Amino Acid Sequence, Animals, B3S, Conserved Sequence, Humans, Ligands, Mice, Microtubule-Associated Proteins, MIKICA, Models, Molecular, PF, PIM, Protein Conformation, alpha-Helical, Protein Domains.

  • W. Wang, S. Cantos-Fernandes, Y. Lv, H. Kuerban, S. Ahmad, C. Wang, et B. Gigant, « Insight into microtubule disassembly by kinesin-13s from the structure of Kif2C bound to tubulin », Nature Communications, vol. 8, nᵒ 1, p. 70, juill. 2017.
    Résumé : Kinesin-13s are critical microtubule regulators which induce microtubule disassembly in an ATP dependent manner. To clarify their mechanism, we report here the crystal structure of a functional construct of the kinesin-13 Kif2C/MCAK in an ATP-like state and bound to the αβ-tubulin heterodimer, a complex mimicking the species that dissociates from microtubule ends during catalytic disassembly. Our results picture how Kif2C stabilizes a curved tubulin conformation. The Kif2C α4-L12-α5 region undergoes a remarkable 25° rotation upon tubulin binding to target the αβ-tubulin hinge. This movement leads the β5a-β5b motif to interact with the distal end of β-tubulin, whereas the neck and the KVD motif, two specific elements of kinesin-13s, target the α-tubulin distal end. Taken together with the study of Kif2C mutants, our data suggest that stabilization of a curved tubulin is an important contribution to the Kif2C mechanism.Kinesin-13s are microtubule depolymerizing enzymes. Here the authors present the crystal structure of a DARPin fused construct comprising the short neck region and motor domain of kinesin-13 in complex with an αβ-tubulin heterodimer, which shows that kinesin-13 functions by stabilizing a curved tubulin conformation.
    Mots-clés : B3S, MIKICA.

  • Y. Wang, Y. Yu, G. - B. Li, S. - A. Li, C. Wu, B. Gigant, W. Qin, H. Chen, Y. Wu, Q. Chen, et J. Yang, « Mechanism of microtubule stabilization by taccalonolide AJ », Nature Communications, vol. 8, p. 15787, juin 2017.
    Résumé : As a major component of the cytoskeleton, microtubules consist of αβ-tubulin heterodimers and have been recognized as attractive targets for cancer chemotherapy. Microtubule-stabilizing agents (MSAs) promote polymerization of tubulin and stabilize the polymer, preventing depolymerization. The molecular mechanisms by which MSAs stabilize microtubules remain elusive. Here we report a 2.05 Å crystal structure of tubulin complexed with taccalonolide AJ, a newly identified taxane-site MSA. Taccalonolide AJ covalently binds to β-tubulin D226. On AJ binding, the M-loop undergoes a conformational shift to facilitate tubulin polymerization. In this tubulin-AJ complex, the E-site of tubulin is occupied by GTP rather than GDP. Biochemical analyses confirm that AJ inhibits the hydrolysis of the E-site GTP. Thus, we propose that the β-tubulin E-site is locked into a GTP-preferred status by AJ binding. Our results provide experimental evidence for the connection between MSA binding and tubulin nucleotide state, and will help design new MSAs to overcome taxane resistance.
    Mots-clés : B3S, MIKICA.


  • S. Ahmad, L. Pecqueur, B. Dreier, D. Hamdane, M. Aumont-Nicaise, A. Plückthun, M. Knossow, et B. Gigant, « Destabilizing an interacting motif strengthens the association of a designed ankyrin repeat protein with tubulin », Scientific Reports, vol. 6, p. 28922, juill. 2016.
    Mots-clés : B3S, MIKICA, PF, PIM.

  • S. Fetics, A. Thureau, V. Campanacci, M. Aumont-Nicaise, I. Dang, A. Gautreau, J. Pérez, et J. Cherfils, « Hybrid Structural Analysis of the Arp2/3 Regulator Arpin Identifies Its Acidic Tail as a Primary Binding Epitope », Structure (London, England: 1993), vol. 24, nᵒ 2, p. 252-260, févr. 2016.
    Résumé : Arpin is a newly discovered regulator of actin polymerization at the cell leading edge, which steers cell migration by exerting a negative control on the Arp2/3 complex. Arpin proteins have an acidic tail homologous to the acidic motif of the VCA domain of nucleation-promoting factors (NPFs). This tail is predicted to compete with the VCA of NPFs for binding to the Arp2/3 complex, thereby mitigating activation and/or tethering of the complex to sites of actin branching. Here, we investigated the structure of full-length Arpin using synchrotron small-angle X-ray scattering, and of its acidic tail in complex with an ankyrin repeats domain using X-ray crystallography. The data were combined in a hybrid model in which the acidic tail extends from the globular core as a linear peptide and forms a primary epitope that is readily accessible in unbound Arpin and suffices to tether Arpin to interacting proteins with high affinity.
    Mots-clés : Actin-Related Protein 2-3 Complex, Animals, B3S, Binding Sites, Carrier Proteins, Crystallography, X-Ray, Epitopes, Fish Proteins, Fishes, Humans, MIKICA, Models, Molecular, PF, PIM, Protein Binding, Protein Conformation, Scattering, Small Angle, X-Ray Diffraction.

  • G. Lippens, I. Landrieu, C. Smet, I. Huvent, N. Gandhi, B. Gigant, C. Despres, H. Qi, et J. Lopez, « NMR Meets Tau: Insights into Its Function and Pathology », Biomolecules, vol. 6, nᵒ 2, p. 28, juin 2016.

  • P. Llinas, M. Chenon, T. Q. Nguyen, C. Moreira, A. de Régibus, A. Coquard, M. J. Ramos, R. Guérois, P. A. Fernandes, et J. Ménétrey, « Structure of a truncated form of leucine zipper II of JIP3 reveals an unexpected antiparallel coiled-coil arrangement », Acta Crystallographica Section F Structural Biology Communications, vol. 72, nᵒ 3, p. 198-206, mars 2016.

  • Y. Wang, H. Zhang, B. Gigant, Y. Yu, Y. Wu, X. Chen, Q. Lai, Z. Yang, Q. Chen, et J. Yang, « Structures of a diverse set of colchicine binding site inhibitors in complex with tubulin provide a rationale for drug discovery », FEBS Journal, vol. 283, nᵒ 1, p. 102-111, 2016.


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Publications Majeures 2009-2014


- Cao, L. et al. (2014) The structure of apo-kinesin bound to tubulin links the nucleotide cycle to movement. Nature Communications 5, 5364, doi : 10.1038/ncomms6364.

- Gigant, B. et al. (2013) Structure of a kinesin-tubulin complex and implications for kinesin motility. Nature structural & molecular biology 20, 1001-1007, doi:10.1038/nsmb.2624.

- Khan A. and Ménétrey J. (2013) Structural biology of Arf and Rab GTPases’ effector recruitment and specificity. Structure, 21(8):1284-97.

- Ménétrey J., Isabet, T. Ropars V., Mukherjea M., Pylypenko O., Liu X., Perez J., Vachette P., Sweeney H.L. and Houdusse A.M. (2012) Processive steps in the reverse direction require uncoupling of the lead head lever arm of myosin VI. Molecular Cell, 48(1) :75-86.

- Pecqueur, L. et al. (2012) A designed ankyrin repeat protein selected to bind to tubulin caps the microtubule plus end. Proc Natl Acad Sci U S A 109, 12011-12016, doi:10.1073/pnas.1204129109 .

- Reymond P., Coquard A., Chenon M., Zeghouf M, El Marjou A, Thompson A, Ménétrey J. (2012) Structure of the GDP-bound G domain of the RGK protein Rem2. Acta Crystallogr F, 68(Pt 6):626-31.

- Nawrotek, A. et al. (2011) The determinants that govern microtubule assembly from the atomic structure of GTP-tubulin. Journal of molecular biology 412, 35-42, doi:10.1016/j.jmb.2011.07.029.

- Chavrier P. and Ménétrey J. (2010) Toward a structural understanding of arf family:effector specificity. Structure 18(12):1552-8.

- Dorleans, A. et al. (2009) Variations in the colchicine-binding domain provide insight into the structural switch of tubulin. Proc Natl Acad Sci U S A 106, 13775-13779, doi:10.1073/pnas.0904223106.

- Isabet T., Montagnac G., Regazzoni K., Raynal B., El Khadali F., England P., Franco M. Chavrier P. Houdusse A., and Ménétrey J.(2009) The structural basis of Arf effector specificity : the crystal structure of ARF6 in a complex with JIP4. EMBO journal 28(18) : 2835-45.

- Llinas P., Mukherjea M., Kim H., Travaglia M., Safer D., Zong A.B., Ménétrey J., Franzini-Armstrong C., Selvin P. R., Houdusse A. and Sweeney H. L. (2009) Myosin VI dimerization triggers an unfolding of a 3-helix bundle in order to extend its reach. Molecular Cell 35(3) : 305-15.


Publications Majeures avant 2009


- Ménétrey J., Llinas P., Cicolari J., Squires G., Liu X., Li A., Sweeney H.L. and Houdusse A. (2008) The post-rigor structure of myosin VI and implications for the recovery stroke. EMBO J. 27(1):244-52.

- Ménétrey J., Llinas P., Mukherjea M., Sweeney H.L., and Houdusse A. (2007) The structural basis for the large powerstroke of myosin VI. Cell 131(2) :300-308.

- Ménétrey J., Perderiset M., Cicolari J., Dubois T., Elkhatib N., El Khadali F., Franco M., Chavrier P. et Houdusse A. (2007) Structural basis for ARF1-mediated recruitment of ARHGAP21 to Golgi membranes. EMBO Journal, 26(7), 1953-1962, 2007.

- Splingard A., Ménétrey J., Perderiset M, Cicolari J., Regazoni K., Hamoudi F., Cabanié L., El Marjou A., Wells AL., Houdusse A. and de Gunzburg J. (2007) Biochemical and Structural characterization of the Gem GTPase. JBC, 282(3), 1905-15.

- Gigant, B. et al. (2005) Structural basis for the regulation of tubulin by vinblastine. Nature 435, 519-522, doi:10.1038/nature03566.

- Ménétrey J., Bahloul A., Wells AL, Sweeney HL and Houdusse A. (2005) The structure of myosin VI reveals the mechanism of directionality reversal for the myosin motors. Nature, 435(7043), 779-785

- Ravelli, R. B. et al. (2004) Insight into tubulin regulation from a complex with colchicine and a stathmin-like domain. Nature 428, 198-202.

- Gigant, B. et al. (2000) The 4 Å X-ray structure of a tubulin:stathmin-like domain complex. Cell 102, 809-816.


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