?-tubulin is essential in Tetrahymena thermophila for the assembly and stability of basal bodies.

ABSTRACT: Basal bodies and centrioles are conserved microtubule-based organelles the improper assembly of which leads to a number of diseases, including ciliopathies and cancer. Tubulin family members are conserved components of these structures that are integral to their proper formation and function. We have identified the ?-tubulin gene in Tetrahymena thermophila and detected the protein, through fluorescence of a tagged allele, to basal bodies. Immunoelectron microscopy has shown that ?-tubulin localizes primarily to the core microtubule scaffold. A complete genomic knockout of ?-tubulin has revealed that it is an essential gene required for the assembly and maintenance of the triplet microtubule blades of basal bodies. We have conducted site-directed mutagenesis of the ?-tubulin gene and shown that residues within the nucleotide-binding domain, longitudinal interacting domains, and C-terminal tail are required for proper function. A single amino acid change of Thr150, a conserved residue in the nucleotide-binding domain, to Val is a conditional mutation that results in defects in the spatial and temporal assembly of basal bodies as well as their stability. We have genetically separated functions for the domains of ?-tubulin and identified a novel role for the nucleotide-binding domain in the regulation of basal body assembly and stability.

SUBMITTER: Ross I 

PROVIDER: S-EPMC3730247 | biostudies-literature | 2013 Aug

REPOSITORIES: biostudies-literature