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UBC Theses and Dissertations
CLASP : a microtubule-based integrator of the hormone-mediated transitions from cell division to elongation in Arabidopsis thaliana Ruan, Yuan
Microtubules have long been known to play a vital role in plant growth and development, which is a complex process and needs to be regulated by both environmental and endogenous hormonal signals. CLASP, an important microtubuleassociated protein, has been shown to be involved in both cell division and expansion. The major goal of my thesis was to explore the function of CLASP in new pathways as well as to identify novel factors that are responsible for its function and distribution. Yeast 2-hybird analysis done independently by two of our collaborators indicated that CLASP interacts strongly with the endocytic membrane-associated protein SNX1. SNX1 had been implicated in the intracellular trafficking of the auxin efflux carrier PIN2. We proved the direct interaction between CLASP and SNX1 by colocalization and BiFC using live cell imaging and we found that clasp-1 mutants have an altered distribution pattern of SNX1, reduced abundance of PIN2 and a series of auxin-related phenotypes such as dwarfism, enhanced lateral branching and aberrant auxin distribution. Druginduced microtubule disruption caused clasp-1-like defects on PIN2 stability. This study illustrated the role of CLASP and microtubules in polar auxin transport and auxin signalling pathway. Previous studies revealed a cross-talk between auxin and brassinosteroids. I found that the two major transcription factors in the brassinosteroid signalling pathway directly target the CLASP promoter to repress its transcription. Reduced CLASP expression is correlated with a transverse orientation of cortical microtubules in root meristematic cells, switching them from division to differentiation. Also CLASP and microtubules stabilize the BR receptor BRI1 by preventing its degradation, a similar mechanism as for PIN2. This research highlights the importance of CLASP in BR-modulated meristem cell identity and the potential role of CLASP as a node between auxin and BR pathways. I carried out an immunoprecipitation experiment to identify putative CLASP interactors and obtained TRM19. Further analysis failed to confirm a direct interaction but suggested that TRM19 is a microtubule-associated protein. Its high expression in dividing cells is consistent with prior report that it functions in PPB formation during the cell cycle.
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