Liu, T.; (2009) The identification of the Tao-1 kinase as a key regulator of microtubule dynamics. Doctoral thesis , UCL (University College London).

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Abstract

In a multicellular animal, the different functions of cells are dependent on their diversity of shapes. Protein phosphorylation and dephosphorylation play an important role in altering the cell shape in response to internal and external signals. In this study I focused on all kinases to explore the gene functions and mechanisms underlying the regulation of cell morphology. Using genomic information I have constructed a Drosophila kinase RNAi library, which was then used to screen six different cell lines from two different tissues of origin for novel genes involved in the generation of cell form. In doing so, I identified a group of common regulators of cell behaviour and morphology together with a set of cell-type specific kinases. Importantly, this analysis also revealed that, when considering the kinome, gene expression signatures are a poor measure of cell type specific differences in gene function, measured by comparing with the microarray data available in our lab. Most significantly, these screens identified a novel role for Mnb/DYRK1A, a kinase associated with Down’s Syndrome, in the regulation of actin-based protrusions in cell lines derived from neuronal lineages. Furthermore, I identified a single STE20 kinase Drosophila Tao-1 in the core set which was required for the normal morphology of all cell lines examined. The RNAi-mediated depletion of Tao-1 or expression of the kinase-dead protein leads to the stabilisation of microtubules and to the formation of long microtubule-based protrusions, whereas the overexpression of Tao-1 destabilizes microtubules. I also showed in this study that Tao-1 acts independently of Par-1 and Tau in this process. Instead, Tao-1 appears to function together with EB1 in the regulation of microtubule plus end instability at the cell cortex. Two microtubule motor proteins, Khc and Khc-73, interact with Tao-1 in a yeast two hybrid screen, which hints at a possible role for the transport of Tao-1 along filaments to microtubule plus ends and the cell cortex. Although evidence for a direct role for Tao-1 in the phosphorylation and/or regulation of EB1 has yet to be established, this is to my knowledge first time a kinase has been found acting at the cell cortex to regulate microtubule plus end stability.

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