γ-Tubulin is a minus end-specific microtubule binding protein

Abstract

The role of microtubules in mediating chromosome segregation during mitosis is well-recognized. In addition, interphase cells depend upon a radial and uniform orientation of microtubules, which are intrinsically asymmetric polymers, for the directional transport of many cytoplasmic components and for the maintenance of the structural integrity of certain organelles. The slow growing minus ends of microtubules are linked to the centrosome ensuring extension of the fast growing plus ends toward the cell periphery. However, the molecular mechanism of this linkage is not clear. One hypothesis is that γ-tubulin, located at the centrosome, binds to the minus ends of microtubules. To test this model, we synthesized radiolabeled γ- tubulin in vitro. We demonstrate here biochemically a specific, saturable, and tight (K(d) = 10-10 M) interaction of γ-tubulin and microtubule ends with a stoichiometry of 12.6 ± 4.9 molecules of γ-tubulin per microtubule. In addition, we designed an in vitro assay to visualize γ-tubulin at the minus ends of axonemal microtubules. These data show that γ-tubulin represents the first protein to bind microtubule minus ends and might be responsible for mediating the link between microtubules and the centrosome.