Abstract
Centrioles and basal bodies are essential for a variety of cellular processes that include the recruitment of proteins to these structures for both centrosomal and ciliary function. This recruitment is compromised when centriole/basal body assembly is defective. Mutations that cause basal body assembly defects confer supersensitivity to Taxol. These include bld2, bld10, bld12, uni3, vfl1, vfl2, and vfl3. Flagellar motility mutants do not confer sensitivity with the exception of mutations in the p60 (pf19) and p80 (pf15) subunits of the microtubule severing protein katanin. We have identified additional pf15 and bld2 (ε-tubulin) alleles in screens for Taxol sensitivity. Null pf15 and bld2 alleles are viable and are not essential genes in Chlamydomonas. Analysis of double mutant strains with the pf15-3 and bld2-6 null alleles suggests that basal bodies in Chlamydomonas may recruit additional proteins beyond katanin that affect spindle microtubule stability. The bld2-5 allele is a hypomorphic allele and its phenotype is modulated by nutritional cues. Basal bodies in bld2-5 cells are missing proximal ends. The basal body mutants show aberrant localization of an epitope-tagged p80 subunit of katanin. Unlike IFT proteins, katanin p80 does not localize to the transition fibers of the basal bodies based on an analysis of the uni1 mutant as well as the lack of colocalization of katanin p80 with IFT74. We suggest that the triplet microtubules are likely to play a key role in katanin p80 recruitment to the basal body of Chlamydomonas rather than the transition fibers that are needed for IFT localization.
Highlights
Taxol is a natural product that binds to b-tubulin and stabilizes microtubules in eukaryotic cells
In the unicellular green alga, Chlamydomonas reinhardtii, Schibler and Huang showed that a mutation in b-tubulin confers resistance to colchicine and supersensitivity to Taxol
We screened other mutants with defects in axonemal substructures needed for motility [55,57,58]and only the two katanin mutants confer Taxol supersensitivity
Summary
Taxol is a natural product that binds to b-tubulin and stabilizes microtubules in eukaryotic cells. In the unicellular green alga, Chlamydomonas reinhardtii, Schibler and Huang showed that a mutation in b-tubulin (tub2-1) confers resistance to colchicine and supersensitivity to Taxol. They concluded that the microtubules in tub cells are hyperstabilized, which causes the Taxol supersensitive phenotype [2]. Katanin is a microtubule severing protein [3,4] It is composed of two subunits; the p60 subunit is a catalytic AAA ATPase and the p80 subunit targets the heterodimer to the centrosome in metazoans. Katanin influences microtubule dynamics through its ability to sever microtubules. This property is observed in Drosophila S2 mitotic spindles [5] and C. elegans meiotic spindles [6]. There appear to be several pathways in cells that when mutated confer Taxol sensitivity
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