Elucidating the Functions and Impact of Inhibition of Mitotic Kinesins in Candida Albicans
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Authors
Shoukat, Irsa
Date
Type
thesis
Language
eng
Keyword
Candida Albicans , Cell Biology , Fungal Proteins , Kinesin-5 , Mitosis , Antifungal Agents
Alternative Title
Abstract
Kinesin-5 family members are classically known for generating outward forces that push spindle poles apart to drive bipolar spindle assembly and elongation. Kinesin-14 members contribute inward forces that pull spindle poles together and control its elongation prior to anaphase. In many eukaryotes, loss of kinesin-5 activity leads to failure of bipolar spindle formation and cell death. However, simultaneous loss of the counteracting kinesin-14 rectifies this problem and cells survive. This phenomenon led to the widely accepted “force-balance” model explaining the actions of these mitotic motor proteins in orchestrating spindle assembly. In this thesis, the cell biological studies of kinesin-5 (Kip1) and kinesin-14 (Kar3Cik1) in Candida albicans reveal distinct mechanisms of spindle assembly and stabilization. Although defective in normal progression through mitosis, kip1Δ/Δ cells are viable and can form bipolar spindles. Also unique is that simultaneous loss of Kip1 and Kar3Cik1 is lethal. Therefore, spindle force-balance is either not restored upon dual depletion of these motors, or their combined loss cannot be compensated for by other spindle-associated factors. Using time-lapse microscopy of kip1Δ/Δ cells, it was observed that many budding cells contained two bipolar spindles that often segregated independently to the mother and bud. In kip1Δ/Δ cells that had experienced cell-cycle arrest, essentially all exhibited this duplicated spindle morphology. These phenotypes imply that C. albicans predominantly employs kinesin-5 as a spindle microtubule crosslinking protein, rather than as a microtubule sliding motor needed for outward pushing force. Examination of Kip1 localization by fluorescence microscopy indicated that these putative crosslinking activities are focused at the spindle poles and at the spindle midzone. By following spindle dynamics, the spindle duplication event was established to occur at the onset of anaphase by bipolar spindle breakage at the midzone. Additionally, it appeared kip1Δ/Δ cells contained multiple clusters of chromatin. These findings imply a role of Kip1 in maintaining spindle integrity, presumably by interpolar microtubule cross-bridging. Finally, similar to kar3cik1Δ/Δ cells, kip1Δ/Δ cells are defective at mature hyphal growth. Combined with the compromised spindle function of kip1Δ/Δ cells, the lack of identifiable septa in immature hyphal compartments suggests that improper distribution of nuclear material underlies this defect.
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ProQuest PhD and Master's Theses International Dissemination Agreement
Intellectual Property Guidelines at Queen's University
Copying and Preserving Your Thesis
This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner.