Erlemann et al. count the number of γ-tubulin complexes that assemble together to nucleate microtubules in S. cerevisiae.
Budding yeast microtubules are nucleated from spindle pole bodies (SPBs) by γ-tubulin small complexes (γ-TuSCs), which consist of two γ-tubulin subunits and one molecule each of Spc97 and Spc98. In vitro, γ-TuSCs form a spiral with 13 γ-tubulin molecules per turn, which could serve as a template for the 13 tubulin protofilaments of a microtubule. But whether similar numbers of γ-TuSCs nucleate microtubules in vivo was unknown.
By comparing the fluorescence of GFP-tagged γ-TuSC proteins to known fluorescent standards, Erlemann et al. counted the number of γ-TuSC components at yeast SPBs and at the minus ends of individual microtubules, which had been released into the cytoplasm by a genetic trick. The quantification revealed that approximately seven γ-TuSCs nucleate each microtubule in vivo, with an additional three γ-tubulin and two Spc98 molecules also incorporating into each nucleation site.
Budding yeast probably only nucleate microtubules during G1, but γ-TuSC components remained tightly associated with SPBs throughout the cell cycle and changed very little in abundance, suggesting that similar numbers of γ-TuSCs nucleate microtubules in G1 and anchor them to SPBs during the rest of the cell cycle.
Erlemann et al.’s results support the idea that γ-TuSCs nucleate microtubules in vivo by forming a spiral-shaped template with 13 γ-tubulin monomers per turn. The authors think that the extra γ-tubulin and Spc98 molecules may stabilize the spiral, fulfilling roles played in higher eukaryotes by specialized proteins that are absent in budding yeast.