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Title: The cytoskeleton and polarity in the C. elegans embryo: Understanding microtubule dependent signalling in the generation of cellular asymmetries.
Authors: Martin, Jack
Issue Date: 2022
Publisher: Newcastle University
Abstract: Asymmetry within cells relies on the uneven distribution of molecules and organelles required for cell function and viability. For example, epithelial cells form an apical-basolateral polarity which confers different protein and phospholipid compositions at the membrane of each domain. Controlled enrichment of integrins on the basolateral domain ensures that an epithelial cell remains anchored to the basement membrane, required to prevent cell plasticity and epithelial to mesenchymal transition. Migrating cells require the assembly of the cytoskeleton at their leading edge and disassembly at the rear to move with direction, whilst asymmetrically dividing cells rely on different concentrations of proteins and RNAs across the mitotic axis to program the distinct fates of each daughter cell after division. The polarity effector proteins, which control polarity establishment and maintenance in cells are broadly conserved, however respond to different cues and have variable contributions dependent on cell type, organism, and stage, be it inherited or de novo polarity. Failure to generate and maintain correct asymmetry in cells can lead to disease states including cancer. In our lab, we study the contribution of the cytoskeleton in the establishment of cell polarity. For this, we use the C. elegans zygote as a model due to the conserved molecular machinery and its well characterised embryonic development. Asymmetry first initiates in the single cell zygote, post-fertilisation, upon which the globally dynamic actomyosin network underlying the cell membrane undergoes localised relaxation at the site in closest proximity to the matured sperm-derived centrosome pair. The resultant gradient of actomyosin contraction generates a cortical cytoplasmic flow towards the future anterior pole and carries with it, the cortical anterior polarity effector PAR proteins. The newly unoccupied cortex is then inhabited by a cytoplasmic pool of posterior PAR proteins. The cue for this change in cortical dynamics is derived from a signal of sequestered AIR-1 which diffuses from the sperm donated centrosome. A second, lesser studied, pathway to establish polarity in the zygote is believed to occur due to the enhanced stabilisation of posterior PARs at the cortex, aided via centrosomal microtubules which prevent phosphorylation from anterior PARs that initially inhibit posterior PAR membrane recruitment. Through mutual antagonism the anterior and posterior PARs can define their own boundaries and maintain polarity. As both pathways rely on the centrosome, studies to characterise these processes individually have been difficult. Disrupting the centrosome compromises both pathways while interrupting downstream components of either pathway results in masked phenotypes due to effective cell polarisation via the other pathway. To overcome the functional redundancy that makes studying the microtubule-dependent pathway difficult, I have utilised a functionally null mutant of the gene, nop-1, which lacks early actomyosin flows to then screen for regulators of the microtubule-dependent pathway of polarity establishment, now required for embryo viability. Through this, and follow up immunofluorescent stains of polarity markers, I have identified novel proteins required for efficient cellular asymmetry. One such protein was the chromokinesin, KLP-19, previously known to aid chromosomal alignment and segregation during metaphase/ anaphase of IV meiosis and mitosis. I have shown that KLP-19 is required to keep the centrosome restricted to the posterior pole during polarity establishment, necessary to facilitate a robust symmetry breaking signal and ensure centrosome separation occurs in a timely manner for the setup of the mitotic spindle. It is likely that KLP-19 performs this role by localising to, and crosslinking, centrosomal and cortical microtubules that meet in an antiparallel manner.
Description: Ph. D. Thesis
Appears in Collections:Biosciences Institute

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