Seminar Title: Structural and biochemical studies reveal principles of microtubule nucleation
Tarun Kapoor, Ph.D.
Pels Family Professor
The Rockefeller University
Seminar abstract: Assembly of the cell division apparatus depends on the 𝛾-tubulin ring complex (𝛾-TuRC), an essential regulator of centrosomal and non-centrosomal microtubule formation. Our structural studies of this ~2.3 MDa complex reveal how >31 proteins, including 𝛾-tubulin and GCP2-6, as well as MZT1 and an actin-like protein in a “lumenal bridge” (LB) are organized into an asymmetric cone-shape architecture. We have also biochemically reconstituted the human 𝛾-TuRC (𝛾-TuRC-GFP), a ~35S complex that nucleates microtubules in vitro. In addition, we have characterized a subcomplex, 𝛾-TuRCΔLB-GFP, which lacks MZT1 and actin. Remarkably, we find that 𝛾-TuRCΔLB-GFP nucleates microtubules in a guanine nucleotide-dependent manner and with similar kinetics as the holocomplex. Electron microscopy reveals that 𝛾-TuRC-GFP resembles the native 𝛾-TuRC architecture, while 𝛾-TuRCΔLB-GFP adopts a partial cone shape presenting only 8-10 𝛾-tubulin subunits and lacks a well-ordered lumenal bridge. Our structure-function analysis suggests that the lumenal bridge facilitates the self-assembly of regulatory interfaces around a microtubule-nucleating “core” within the 𝛾-TuRC. Together, recombinant forms of human 𝛾-TuRC, a/b-tubulin and the hetero-octameric augmin complex will help define the critical components and uncover the basic principles of microtubule formation during cell division.
The department of Physiology & Biophysics acknowledges the Coast Salish peoples of this land, the land which touches the shared waters of all tribes and bands within the Suquamish, Tulalip and Muckleshoot nations. It is in this land where we work, teach, and learn.