Frontier Research Institute for Interdisciplinary Sciences
Tohoku University

This study aims to elucidate the mechanism by which microtubules, critical components of cellular architecture, are nucleated from the centrosome, a major microtubule-organizing center. While recent structural analyses have begun to resolve the architecture of protein complexes constituting the centrosome, we adopt the perspective that "understanding the structure alone is insufficient without the ability to reconstruct it." Based on this premise, we pursue a synthetic approach to reconstitute the centrosome in vitro.

In cells, microtubule nucleation is facilitated by the γ-tubulin ring complex (γ-TuRC), which forms a ring-shaped scaffold approximately 25 nm in diameter—an architecture thought to determine the intrinsic diameter of microtubules. However, manipulating the size of γ-TuRC in living cells is currently infeasible. To overcome this limitation, we leverage materials engineering techniques to design and fabricate artificial ring-shaped scaffolds with defined diameters that mimic γ-TuRC. These synthetic scaffolds provide a unique experimental platform to directly test the relationship between the γ-TuRC diameter and microtubule architecture.

Using total internal reflection fluorescence (TIRF) microscopy, we will visualize microtubule nucleation from these engineered scaffolds. In addition to simple ring geometries, we will test helical configurations that more closely resemble the native γ-TuRC structure, allowing us to compare their nucleation efficiencies. Furthermore, we will employ electron microscopy to precisely measure the diameters of microtubules formed from each scaffold and assess their correlation with the scaffold diameter.

In the long term, this research aims to lay the foundation for a new field of "synthetic organelle construction," offering novel insights into the design principles of intracellular architecture.