Noncore Components of the Fission Yeast γ-Tubulin Complex

Abstract

Relatively little is known about the in vivo function of individual components of the eukaryotic γ-tubulin complex (γ-TuC). We identified three genes, gfh1+, mod21+, and mod22+, in a screen for fission yeast mutants affecting microtubule organization. gfh1+ is a previously characterized γ-TuC protein weakly similar to human γ-TuC subunit GCP4, whereas mod21+ is novel and shows weak similarity to human γ-TuC subunit GCP5. We show that mod21p is a bona fide γ-TuC protein and that, like gfh1Δ mutants, mod21Δ mutants are viable. We find that gfh1Δ and mod21Δ mutants have qualitatively normal microtubule nucleation from all types of microtubule-organizing centers (MTOCs) in vivo but quantitatively reduced nucleation from interphase MTOCs, and this is exacerbated by mutations in mod22+. Simultaneous deletion of gfh1p, mod21p, and alp16p, a third nonessential γ-TuC protein, does not lead to additive defects, suggesting that all three proteins contribute to a single function. Coimmunoprecipitation experiments suggest that gfh1p and alp16p are codependent for association with a small “core” γ-TuC, whereas mod21p is more peripherally associated, and that gfh1p and mod21p may form a subcomplex independently of the small γ-TuC. Interestingly, sucrose gradient analysis suggests that the major form of the γ-TuC in fission yeast may be a small complex. We propose that gfh1p, mod21p, and alp16 act as facultative “noncore” components of the fission yeast γ-TuC and enhance its microtubule-nucleating ability.