Chlamydomonas γ-tubulin mutations reveal a critical role for γ-TuRC in maintaining the stability of centriolar microtubules

The centriolar triplet microtubule consists of an A-tubule with 13 protofilaments, and B-and C-tubules, each with 10 protofilaments. Although the formation of the triplets has been shown to require γ-tubulin, its specific role in the formation of each tubule remains elusive. We isolated two novel Chlamydomonas reinhardtii mutants, bld13-1 and bld13-2 , each expressing γ-tubulin with a single amino-acid substitution (T292I or E89D). Similar to known centriole-deficient mutants, both mutants exhibited defects in ciliary assembly, nuclear number, as well as the number and orientation of cytoplasmic microtubules. Genetic analyses of the mutants, along with the expression of the mutant γ-tubulins in the wild-type cells, suggested that both mutants exert dominant-negative effects over wild-type γ-tubulin. Interestingly, although the centrioles in these mutants retained the typical nine triplet structure, their triplets frequently lacked several protofilaments in specific regions of the A- and C-tubules. The protofilament loss occurs more frequently in the proximal region of the centriole. These structural defects suggest a critical role for γ-tubulin in maintaining the stability of the A- and C-tubules of centriolar triplets.