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Mechanisms of microtubule dynamics regulation by the MICROTUBULE ORGANIZATION 1 protein Kawamura, Eiko

Abstract

The Arabidopsis thaliana MOR1 (MICROTUBULE ORGANIZATION1) protein belongs to the MAP215/Dis1 family of microtubule-associated proteins. The temperature-sensitive mor1 mutants have N-terminal amino acid substitutions, which lead to cortical microtubule disorganization (Whittington et al., 2001). Here I demonstrate by use of live cell imaging and immunolabelling that MOR1 is important for function and organization of all microtubule arrays during cell division and keeps microtubules highly dynamic. Although disruption of mitotic and cytokinetic microtubule arrays is not detected in all dividing mor1-1 cells, quantitative analysis identified distinct defects in preprophase bands, spindles and phragmoplasts. In nearly half of dividing mor1-1 cells, preprophase bands are not detected, and those that do form are often disrupted. mor1-1 spindles and phragmoplasts are short and abnormally organized and persist for longer times than in wild-type, leading to aberrant chromosome arrangements, misaligned cell plates and multinucleate cells. Immunofluorescence indicates that the mutant mor1-1[sup L174F] protein remains associated along the full length of all microtubule arrays, in spite of their disorganization. This suggests the N-terminal region altered by the mor1-1 mutation does not regulate the binding of MOR1 to microtubules, but that it instead plays a role in microtubule dynamics. Microtubule dynamics were therefore measured in living leaf cells expressing three microtubule reporter proteins, GFP-TUA, CMV35S ::GFP-EB1 and Pro [sub EB1] ::EB1-GFP. Dynamics analysis indicates that MOR1 promotes constant and rapid growth and shrinkage and prevents pausing of microtubules. Integrating this new information with previous observations showing that MOR1 and its tobacco homologue MAP200 can bind tubulin oligomers (Twell et al., 2002; Hamada et al., 2004), and that XMAP215 speeds up microtubule growth and shrinkage in 40-60nm increments (Kerssemakers et al., 2006), I postulate that MOR1 might promote microtubule growth and shrinkage by adding and removing tubulin oligomers. Consistent with this idea, the N-terminal region of MOR1 consists of 5 TOG domains, which each span the approximate length of one tubulin dimer within a protofilament chain. I define experiments and present preliminary data to test the hypothesis that each MOR1 protein can add or remove up to 5 tubulin dimers at a time.

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