Centrosome

Ultimate tensile strength

Optics

Physics

Total internal reflection fluorescence microscope

Biophysics

Cell

Fluorescence microscope

Chemistry

Metallurgy

Materials science

Microtubule

Cell cycle

Elongation

Membrane

Biochemistry

Nucleation

Crystallography

Tubulin

Biology

Fluorescence

Microtubule nucleation

Cell biology

Thermodynamics

Developmental Cell

The γ-tubulin ring complex (γTuRC) is the major microtubule nucleator in cells. The mechanism of its regulation is not understood. We purified human γTuRC and measured its nucleation properties in a total internal reflection fluorescence (TIRF) microscopy-based real-time nucleation assay. We find that γTuRC stably caps the minus ends of microtubules that it nucleates stochastically. Nucleation is inefficient compared with microtubule elongation. The 4 Å resolution cryoelectron microscopy (cryo-EM) structure of γTuRC, combined with crosslinking mass spectrometry analysis, reveals an asymmetric conformation with only part of the complex in a “closed” conformation matching the microtubule geometry. Actin in the core of the complex, and MZT2 at the outer perimeter of the closed part of γTuRC appear to stabilize the closed conformation. The opposite side of γTuRC is in an “open,” nucleation-incompetent conformation, leading to a structural asymmetry explaining the low nucleation efficiency of purified human γTuRC. Our data suggest possible regulatory mechanisms for microtubule nucleation by γTuRC closure.

Microtubule Nucleation Properties of Single Human γTuRCs Explained by Their Cryo-EM Structure