The microtubule polymerase Stu2 promotes oligomerization of the γ-TuSC for cytoplasmic microtubule nucleation

Stu2/XMAP215/ZYG-9/Dis1/Alp14/Msps/ch-TOG family members in association with with gamma-tubulin complexes nucleate microtubules, but we know little about the interplay of these nucleation factors. Here, we show that the budding yeast Stu2 in complex with the gamma-tubulin receptor Spc72 nucleates microtubules in vitro without the small gamma-tubulin complex (gamma-TuSC). Upon gamma-TuSC addition, Stu2 facilitates Spc72-gamma-TuSC interaction by binding to Spc72 and gamma-TuSC. Stu2 together with Spc721 gamma-TuSC increases microtubule nucleation in a process that is dependent on the TOG domains of Stu2. Importantly, these activities are also important for microtubule nucleation in vivo. Stu2 stabilizes Spc72-gamma-TuSC at the minus end of cytoplasmic microtubules (cMTs) and an in vivo assay indicates that cMT nucleation requires the TOG domains of Stu2. Upon gamma-tubulin depletion, we observed efficient cMT nucleation away from the spindle pole body (SPB), which was dependent on Stu2. Thus, gamma-TuSC restricts cMT assembly to the SPB whereas Stu2 nucleates cMTs together with gamma-TuSC and stabilizes gamma-TuSC at the cMT minus end.