title: A mutation of the fission yeast EB1 overcomes negative regulation by phosphorylation and stabilizes microtubules.
abstract: Mal3 is a fission yeast homolog of EB1, a plus-end tracking protein (+TIP). We have generated a mutation (89R) replacing glutamine with arginine in the calponin homology (CH) domain of Mal3. Analysis of the 89R mutant in vitro has revealed that the mutation confers a higher affinity to microtubules and enhances the intrinsic activity to promote the microtubule-assembly. The mutant Mal3 is no longer a +TIP, but binds strongly the microtubule lattice. Live cell imaging has revealed that while the wild type Mal3 proteins dissociate from the tip of the growing microtubules before the onset of shrinkage, the mutant Mal3 proteins persist on microtubules and reduces a rate of shrinkage after a longer pausing period. Consequently, the mutant Mal3 proteins cause abnormal elongation of microtubules composing the spindle and aster. Mal3 is phosphorylated at a cluster of serine/threonine residues in the linker connecting the CH and EB1-like C-terminal motif domains. The phosphorylation occurs in a microtubule-dependent manner and reduces the affinity of Mal3 to microtubules. We propose that because the 89R mutation is resistant to the effect of phosphorylation, it can associate persistently with microtubules and confers a stronger stability of microtubules likely by reinforcing the cylindrical structure.
authors: Graduate School of Biostudies, Kyoto University, Kitashirakawa-Oiwake cho, Sakyo ku, Kyoto 606-8502, Japan.
authors: Makoto
authors: M
authors: Iimori
authors: Kanako
authors: K
authors: Ozaki
authors: Yuji
authors: Y
authors: Chikashige
authors: Toshiyuki
authors: T
authors: Habu
authors: Yasushi
authors: Y
authors: Hiraoka
authors: Takahisa
authors: T
authors: Maki
authors: Ikuko
authors: I
authors: Hayashi
authors: Chikashi
authors: C
authors: Obuse
authors: Tomohiro
authors: T
authors: Matsumoto
mesh_terms: D000595:Amino Acid Sequence; D015536:Down-Regulation; D008869:Microtubule-Associated Proteins / Q000235:genetics* / Q000378:metabolism*; D008870:Microtubules / Q000737:chemistry / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000502:physiology*; D030781:Organisms, Genetically Modified; D010766:Phosphorylation / Q000235:genetics; D011494:Protein Kinases / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D055550:Protein Stability; D012568:Schizosaccharomyces / Q000235:genetics*; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics* / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
chemical_list: C405804:EB1 microtubule binding proteins; C108851:Mal3 protein, S pombe; D008869:Microtubule-Associated Proteins; D029702:Schizosaccharomyces pombe Proteins; D011494:Protein Kinases