Marxer, F. | Search bioRxiv

bioRxiv2026

An architectural switch in the evolution of the γ-tubulin ring complex

Canonical microtubules contain 13-protofilaments and are templated by the {gamma}-tubulin ring complex ({gamma}-TuRC). However, some eukaryotes assemble non-canonical microtubules, like the 11-protofilament structures found in Caenorhabditis elegans. How {gamma}-TuRCs adapt to template alternative microtubule geometries is unclear. Here, we present the cryo-electron microscopy structure of the C. elegans {gamma}-TuRC ({gamma}-TuRCCe), revealing a cone-shaped assembly consistent with an 11-protofilament template. While the complex incorporates the conserved subunits actin, GCP2 and GCP3, {gamma}-TuRCCe replaces GCP4-6 with a divergent 4-spoked assembly containing additional copies of GCP2 and the nematode-specific proteins GTAP-1 and GTAP-2. Structures of nucleotide-free {gamma}-TuRCCe subcomplexes reveal partial {gamma}-tubulin unfolding, suggesting nucleotide binding stabilizes eukaryotic tubulins. Remarkably, reconstituted 4-spoked assemblies can multimerize into [~]13-fold symmetric microtubule nucleation templates in vitro, contrasting with the native complexs 11-protofilament architecture. Our work defines the structural blueprint of an 11-protofilament microtubule template and shows how divergent {gamma}-tubulin components are repurposed to accommodate non-canonical microtubule lattices.

biochemistry↗

bioRxiv2023

Closure of the gamma-tubulin ring complex by CDK5RAP2 activates microtubule nucleation

Microtubule nucleation in cells is templated by the {gamma}-tubulin ring complex ({gamma}-TuRC), a 2.3 MDa multiprotein assembly concentrated at microtubule organizing centers (MTOCs). Current {gamma}-TuRC structures exhibit an open conformation that deviates from the geometry of /{beta}-tubulin in the microtubule, potentially explaining their low in vitro microtubule-nucleating activity. Several proteins have been proposed to activate the {gamma}-TuRC, but the mechanisms underlying activation are not known. Here, we isolated the porcine {gamma}-TuRC using CDK5RAP2s centrosomin motif 1 (CM1) and determined its structure with cryo-electron microscopy. 3D heterogeneity analysis revealed an unexpected conformation of the {gamma}-TuRC, in which five protein modules containing MZT2, GCP2, and CDK5RAP2 decorate the outer face of the holocomplex. These decorations drive a long-range constriction of the {gamma}-tubulin ring, bringing the GCP2/GCP3-rich core of the complex in close agreement with the architecture of a microtubule. A purified CDK5RAP2 fragment stimulated the microtubule nucleating-activity of the porcine {gamma}-TuRC as well as a reconstituted, CM1-free human complex in single molecule assays. Our results show that CDK5RAP2 activates the {gamma}-TuRC by promoting {gamma}-tubulin ring closure, providing a structural mechanism for the regulation of microtubule nucleation by CM1 motif proteins in mammals and revealing conformational transitions in {gamma}-tubulin that prime it for templating microtubule nucleation at MTOCs.

biochemistry↗