The Variation of STRIPAK Components in Regulating Katanin Microtubule-severing in the Caenorhabditis elegans Embryo
Date
2021-06-29
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Abstract
Microtubule-severing plays important roles in organism development, including for cell structure and cell division. Using C. elegans, I studied the regulation of katanin, a conserved microtubule-severing complex. Katanin is required for oocyte meiotic spindle formation; however, it must be inactivated for mitosis to proceed as continued katanin expression is lethal. In addition to two ubiquitin-based protein degradation pathways, another ubiquitin ligase, HECTD1 ubiquitin ligase homologue (HECD-1), regulates katanin activity without affecting katanin levels. Most regulators of katanin act as either a katanin activator or inhibitor; however, hecd-1 switches from being an activator in meiosis to an inhibitor in mitosis. My project set out to characterize the mechanism of hecd-1 mediated regulation of katanin. I used a CRISPR FLAG-tagged HECD-1 to compare the levels and localization of HECD-1 in meiosis and mitosis. HECD-1 showed ubiquitous expression in the cytoplasm throughout meiosis and early development and its levels remained unchanged. In other organisms, HECD-1 is a component of the striatin-interacting kinase phosphatase (STRIPAK) complex. This complex has been known to affect blood vessel formation, apoptosis, cell proliferation and a variety of signalling pathways. Using double mutants and genetic knockdowns, I found the STRIPAK complex components affected katanin activity differently. STRIPAK core components (farl-11, cash-1, let-92, gck-1), except for ccm-3, were katanin inhibitors in mitosis and activators in meiosis, much like hecd-1. In contrast, variable components (M4.1, otub-2) functioned as activators of katanin activity in meiosis and mitosis, while other components (mob-4, C49H3.6) were not involved in katanin microtubule-severing. These experiments outline separate functions for the STRIPAK complex and suggest a paradigm in which the addition of variable components alters the function of the complex. Additional experiments indicate that katanin may be involved with the central spindlin or a tubulin chaperone pathway. My research elucidated the interactions of nearly all the STRIPAK complex components in a single in vivo system and revealed their roles in katanin microtubule severing and cell division.
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Keywords
C. elegans, katanin, meiosis, STRIPAK, development, mitosis, microtubules, genetics, HECD-1
Citation
Lu, T. (2021). The Variation of STRIPAK Components in Regulating Katanin Microtubule-severing in the Caenorhabditis elegans Embryo (Master's thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca.