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 ε-tubulin gene in Tetrahymena thermophila and detected the protein, through fluorescence of a tagged allele, to basal bodies. Immunoelectron microscopy has shown that ε-tubulin localizes primarily to the core microtubule scaffold. A complete genomic knockout of ε-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 ε-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 e-tubulin and identified a novel role for the nucleotide-binding domain in the regulation of basal body assembly and stability.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 3441-3451 |
| Number of pages | 11 |
| Journal | Journal of Cell Science |
| Volume | 126 |
| Issue number | 15 |
| DOIs | |
| State | Published - Sep 9 2013 |
| Externally published | Yes |
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Keywords
- Basal body
- Epsilon tubulin
- Microtubule organizing center
- Tetrahymena thermophila
ASJC Scopus subject areas
- Cell Biology
Cite this
ε-tubulin is essential in Tetrahymena thermophila for the assembly and stability of basal bodies. / Ross, Ian; Clarissa, Christina; Giddings, Thomas H.; Winey, Mark.
In: Journal of Cell Science, Vol. 126, No. 15, 09.09.2013, p. 3441-3451.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - ε-tubulin is essential in Tetrahymena thermophila for the assembly and stability of basal bodies
AU - Ross, Ian
AU - Clarissa, Christina
AU - Giddings, Thomas H.
AU - Winey, Mark
PY - 2013/9/9
Y1 - 2013/9/9
N2 - 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 ε-tubulin gene in Tetrahymena thermophila and detected the protein, through fluorescence of a tagged allele, to basal bodies. Immunoelectron microscopy has shown that ε-tubulin localizes primarily to the core microtubule scaffold. A complete genomic knockout of ε-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 ε-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 e-tubulin and identified a novel role for the nucleotide-binding domain in the regulation of basal body assembly and stability.
AB - 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 ε-tubulin gene in Tetrahymena thermophila and detected the protein, through fluorescence of a tagged allele, to basal bodies. Immunoelectron microscopy has shown that ε-tubulin localizes primarily to the core microtubule scaffold. A complete genomic knockout of ε-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 ε-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 e-tubulin and identified a novel role for the nucleotide-binding domain in the regulation of basal body assembly and stability.
KW - Basal body
KW - Epsilon tubulin
KW - Microtubule organizing center
KW - Tetrahymena thermophila
UR - http://www.scopus.com/inward/record.url?scp=84883375893&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=84883375893&partnerID=8YFLogxK
U2 - 10.1242/jcs.128694
DO - 10.1242/jcs.128694
M3 - Article
C2 - 23704354
AN - SCOPUS:84883375893
VL - 126
SP - 3441
EP - 3451
JO - Journal of Cell Science
JF - Journal of Cell Science
SN - 0021-9533
IS - 15
ER -