Functional dissection and hierarchy of tubulin-folding cofactor homologues in fission yeast.

We describe the isolation of fission yeast homologues of tubulin-folding cofactors B (Alp11) and E (Alp21), which are essential for cell viability and the maintenance of microtubules. Alp11(B) contains the glycine-rich motif (the CLIP-170 domain) involved in microtubular functions, whereas, unlike mammalian cofactor E, Alp21(E) does not. Both mammalian and ...
yeast cofactor E, however, do contain leucine-rich repeats. Immunoprecipitation analysis shows that Alp11(B) interacts with both alpha-tubulin and Alp21(E), but not with the cofactor D homologue Alp1, whereas Alp21(E) also interacts with Alp1(D). The cellular amount of alpha-tubulin is decreased in both alp1 and alp11 mutants. Overproduction of Alp11(B) results in cell lethality and the disappearance of microtubules, which is rescued by co-overproduction of alpha-tubulin. Both full-length Alp11(B) and the C-terminal third containing the CLIP-170 domain localize in the cytoplasm, and this domain is required for efficient binding to alpha-tubulin. Deletion of alp11 is suppressed by multicopy plasmids containing either alp21(+) or alp1(+), whereas alp21 deletion is rescued by overexpression of alp1(+) but not alp11(+). Finally, the alp1 mutant is not complemented by either alp11(+) or alp21(+). The results suggest that cofactors operate in a linear pathway (Alp11(B)-Alp21(E)-Alp1(D)), each with distinct roles.
Mesh Terms:
Amino Acid Sequence, Cell Polarity, Cytoplasm, Fungal Proteins, Gene Dosage, Genes, Essential, Genes, Fungal, Genetic Complementation Test, Glycine, Microtubule-Associated Proteins, Microtubules, Molecular Sequence Data, Molecular Weight, Mutation, Neoplasm Proteins, Protein Binding, Schizosaccharomyces, Schizosaccharomyces pombe Proteins, Sequence Homology, Amino Acid, Suppression, Genetic, Temperature, Tubulin
Mol. Biol. Cell
Date: Sep. 01, 1999
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