Helical microtubule arrays in a collection of twisting tubulin mutants of Arabidopsis thaliana

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Abstract
Anisotropic expansion of plant cells requires organized arrays of cortical microtubules. Mutations in microtubule-associated proteins and a particular mutation in α-tubulins were reported to cause abnormal microtubule arrays and result in helical growth in Arabidopsis thaliana . However, the way in which these mutations affect the organization of microtubules remains unknown. We here identified 32 Arabidopsis twisting mutants that have either missense or amino acid deletion mutations in α- or β-tubulins. Mutations were mapped to the GTPase-activating region in α-tubulin, intra- and interdimer interfaces of tubulin heterodimers, and lateral contact regions among adjacent protofilaments. These dominant-negative tubulin mutants were incorporated into the microtubule polymer and formed shallow helical arrays of distinct handedness along the long axis of the root epidermal cells. A striking correlation exists between the direction in which cortical helical arrays are skewed and the growth direction of elongating roots. The GTPase-activating-region mutant had left-handed helical arrays composed of highly stabilized microtubules, which could be decorated along the entire microtubule lattices with the otherwise tip-localized End Binding 1 protein. A mutation at the intradimer interface, on the other hand, generated highly dynamic microtubules and right-handed helical arrays. Cortical microtubules in wild type and these two tubulin mutants were composed mainly of 13 protofilaments. This comprehensive analysis of tubulin mutations provides insights into the mechanism by which tubulin structures influence microtubule dynamics and organization.