THE REGULATION OF MICROTUBULE DYNAMICS BY +TIPS AND CROSS-LINKING PROTEINS

ABSTRACT Rebecca Cressey Adikes: The regulation of microtubule dynamics by +TIPs and cross-linking proteins (Under the supervision of Kevin C. Slep) The dynamic cytoskeleton is composed of distinct filamentous polymer networks. These polymers create highly dynamic scaffolds for many cellular processes in eukaryotes. The polymers provide tracks for cargo transport, mechanically stabilize the cell, establish cell polarity, and are central components of the force-generating machinery required for cell division. In order to perform these various cellular tasks the polymers undergo spatially and temporally regulated assembly and disassembly. Understanding the spatial and temporal regulation of these cellular polymers is essential to understanding cell function. Here, we investigate the spatial and temporal regulation of two polymer networks in cells, the microtubule (MT) and F-actin networks. We developed and employed methods to alter protein targeting and localization using a novel optogenetic approach and a classical structural/functional approach to understand how the recruitment of effectors of these networks alters network dynamics. We developed and employed an optogenetic tool to gain insight into the msec/sec resolution time regime of MT-F-actin cross-linking. This allowed us to uncover key immediate down stream cellular events of cross-linking including decreased MT comet velocities on the sec time scale and an exclusion of MT from the cell periphery on the min-hr time scale. Additionally, using more classical structure/function based experiments, we were able to alter protein targeting using protein domains and through mutation of the protein sequence by just 1-3Doctor of Philosoph