Conditional mutations in γ-tubulin reveal its involvement in chromosome segregation and cytokinesis

gamma -Tubulin is a conserved essential protein required for assembly and function of the mitotic spindle in humans and yeast. For example, human gamma -tubulin can replace the gamma -tubulin gene in Schizosaccharomyces pombe. To understand the structural /functional domains of gamma -tubulin, we performed a systematic alanine-scanning mutagenesis of human gamma -tubulin (TUBG1) and studied phenotypes of each mutant allele in S. pombe. Our screen, both in the presence and absence of the endogenous S. pombe gamma -tubulin, resulted in 11 lethal mutations and 12 cold-sensitive mutations. Based on structural mapping onto a homology model of human gamma -tubulin generated by free energy minimization, all deleterious mutations are found in residues predicted to be located on the surface, some in positions to interact with alpha- and/or beta -tubulins in the microtubule lattice. As expected, one class of tubg1 mutations has either an abnormal assembly or loss of the mitotic spindle. Surprisingly, a subset of mutants with abnormal spindles does not arrest in M phase but proceeds through anaphase followed by abnormal cytokinesis. These studies reveal that in addition to its previously appreciated role in spindle microtubule nucleation, gamma -tubulin is involved in the coordination of postmetaphase events, anaphase, and cytokinesis.