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The role of MOR1 in the control of microtubule organization and dynamics in Arabidopsis thaliana Donnelly, Caitlin Charlotte Anne

Abstract

The plant cortical microtubule array plays a role in the control of directional cell expansion, and the organization and dynamics of the array are subject to control by a variety of microtubule-associated proteins, many of which coordinate organization of the cortical array in response to environmental stimuli. Point mutations affecting MOR1, a microtubule polymerase/depolymerase, result in disruption to the organization and dynamic properties of microtubules under specific conditions: mutations in the N-terminal TOG1 (tubulin-binding) domain have temperature-conditional phenotypes, while the phenotype of a mutation in the C-terminal region is induced by treatment with the microtubule-destabilizing drug propyzamide. In this thesis, I used mor1 mutants with conditional phenotypes to characterize genetic interactions between different domains of the MOR1 protein, microtubules, and components of a microtubule-targeted environmental stress signalling pathway. Analysis of microtubule organization and dynamics in mor1-tubulin double mutants demonstrated that the handedness of helical growth phenotypes does not always correlate with microtubule growth and shrinkage rates, and showed that a mutation in β-tubulin promoted recovery of microtubule dynamics in the temperature-sensitive mor1-1 mutant. I used live-cell imaging to observe interactions between fluorescently tagged MOR1 and microtubules, demonstrating that addition of a fluorescent tag to the MOR1 C-terminus alters MOR1 function and results in phosphorylation of α-tubulin, which is normally a response to environmental stress. Despite this effect, differences in microtubule binding affinity were observed for MOR1 variants with mutations in the TOG1 and C-terminal regions. I determined that mutation of the C-terminal region of MOR1 (mor1-11) results in activation of the tubulin kinase PHS1, though this did not appear to be mediated by MPK18, a previously characterized PHS1-interacting MAP kinase. In order to identify other possible components of this signalling pathway, I carried out a modifier mutant screen in the mor1-11 genetic background, identifying one enhancer and six suppressors of mor1-11.

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