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Title: Investigating the role of lysosomes during mammalian cell division
Author: Nugues, Charlotte
ISNI:       0000 0004 7970 5061
Awarding Body: University of Liverpool
Current Institution: University of Liverpool
Date of Award: 2019
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Lysosomes are acidic, intracellular, membrane-bound organelles present in all eukaryotic cells. They are filled with more than 60 different hydrolases responsible for the degradation of a wide range of intracellular targets arising from autophagy and extracellular particles internalised through endocytic and phagocytic pathways. Their luminal acidity is achieved via the action of the Vacuolar-ATPase. This complex, multi-subunit, molecular proton pump generates the low pH essential for optimal catalytic activity of hydrolytic enzymes by actively pumping H+ ions (protons) into the lysosome lumen. Historically, lysosomes have been predominantly considered cellular 'waste bags' involved in the degradation and recycling of cellular and extracellular content, however new findings reveal that they are highly dynamic organelles tasked with a wide spectrum of unique biological functions. Lysosomes are equipped to sense and monitor the nutrient status of the cell and coordinate an appropriate response. Similar to the ER, they contain high levels of calcium (Ca2+), therefore they might participate in calcium homeostasis by acting as Ca2+ stores, and are potentially involved in Ca2+ signalling. Ca2+ is also known to play a pivotal role in lysosome trafficking, recycling and fusion with the plasma membrane (PM). The latter process, known as exocytosis, enables membrane donation during PM wound repair, allows the cell to 'rid itself' of unwanted content and participates in the release of hydrolases that perform important extracellular functions such as bone remodelling and extracellular matrix degradation. More recently, lysosomes have been suggested to play a role in cell division, more particularly, cytokinesis. Cytokinesis is the final stage of cell division. This highly orchestrated process involves the formation of an ingression furrow that progressively constricts forming a transient cytoplasmic intercellular bridge. Eventually, the bridge is severed at abscission allowing separation of daughter cells. Lysosomes were found to cluster at either side of the intercellular bridge that separates nascent daughter cells and disruption of lysosomal positioning through a PI4KIIIβ-dependent mechanism was associated with a significant increase in cytokinesis failure. This suggests a functional requirement for lysosomes in cytokinesis. Understanding the regulation of cytokinesis is of the utmost interest as failure in cell division can lead to aneuploidy, genetic instability and malignant phenotypes. The purpose of this thesis was therefore to identify the specific lysosomal function(s) involved in cytokinesis. Using a combination of biochemical and pharmacological approaches coupled with confocal microscopy and TIRF imaging techniques, exocytosis of these organelles was shown to significantly increase during cell division, most specifically in cytokinesis. Finally, lysosome exocytosis during cytokinesis was linked to PI4KIIIβ activity. These findings demonstrate for the first time a role of lysosomal exocytosis in cytokinesis. Manipulation of cytokinesis through targeting a specific aspect of lysosomal physiology (exocytosis) opens up new avenues of investigation. This knowledge may permit novel future approaches to tackling a number of prevalent human diseases.
Supervisor: Haynes, Lee ; Burgoyne, Robert Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral