title: Microtubule-associated proteins control the kinetics of microtubule nucleation.
abstract: Microtubules are born and reborn continuously, even during quiescence. These polymers are nucleated from templates, namely γ-tubulin ring complexes (γ-TuRCs) and severed microtubule ends. Using single-molecule biophysics, we show that nucleation from γ-TuRCs, axonemes and seed microtubules requires tubulin concentrations that lie well above the critical concentration. We measured considerable time lags between the arrival of tubulin and the onset of steady-state elongation. Microtubule-associated proteins (MAPs) alter these time lags. Catastrophe factors (MCAK and EB1) inhibited nucleation, whereas a polymerase (XMAP215) and an anti-catastrophe factor (TPX2) promoted nucleation. We observed similar phenomena in cells. We conclude that GTP hydrolysis inhibits microtubule nucleation by destabilizing the nascent plus ends required for persistent elongation. Our results explain how MAPs establish the spatial and temporal profile of microtubule nucleation.
authors.affiliation: McGill University, Department of Biology, 1205 ave Docteur Penfield Montréal, Québec H3A 1B1, Canada.
authors.forename: Michal
authors.initials: M
authors.lastname: Wieczorek
authors.affiliation: McGill University, Department of Biology, 1205 ave Docteur Penfield Montréal, Québec H3A 1B1, Canada.
authors.forename: Susanne
authors.initials: S
authors.lastname: Bechstedt
authors.affiliation: McGill University, Department of Biology, 1205 ave Docteur Penfield Montréal, Québec H3A 1B1, Canada.
authors.forename: Sami
authors.initials: S
authors.lastname: Chaaban
authors.affiliation: McGill University, Department of Biology, 1205 ave Docteur Penfield Montréal, Québec H3A 1B1, Canada.
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Brouhard
mesh_terms: D000818:Animals; D054468:Axoneme / Q000378:metabolism; D016466:CHO Cells; D045744:Cell Line, Tumor; D018385:Centrosome / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D015151:Immunoblotting; D007700:Kinetics; D018374:LLC-PK1 Cells; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence / Q000379:methods; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000378:metabolism*; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D015739:Nocodazole / Q000494:pharmacology; D058105:Polymerization / Q000187:drug effects; D013552:Swine; D014404:Tubulin / Q000378:metabolism*; D050257:Tubulin Modulators / Q000494:pharmacology
chemical_list: D008869:Microtubule-Associated Proteins; D014404:Tubulin; D050257:Tubulin Modulators; D049452:Green Fluorescent Proteins; C004805:5'-guanylylmethylenebisphosphonate; D006160:Guanosine Triphosphate; D015739:Nocodazole