Systems approaches to the biology of the spindle pole body

Student thesis: Doctoral ThesisDoctor of Philosophy

Abstract

The yeast centrosome or Spindle Pole Body (SPB) is an organelle situated in the nuclear membrane, where it nucleates spindle microtubules and acts as a signalling hub. The SPB is reproduced conservatively, meaning an existing SPB is required to form a new SPB. Additionally, the Mitotic Exit Network (MEN), a group of proteins controlling exit from mitosis localize to the SPB, in an essential step in MEN activation. These properties are conserved in the MicroTubule Organising Centre’s (MTOC) of higher eukaryotes, which also duplicate conservatively and act as signalling hubs. Various studies have explored the effects of forcing individual proteins to interact with the yeast SPB, however no systematic study has been performed. Furthermore, while the MEN has been studied intensively, a unified understanding of how localization and protein activity function together as a system is lacking. I have used Synthetic Physical Interaction (SPI) screening to detect proteins that inhibit growth when forced to associate with the SPB. I found that the SPB is especially sensitive to relocalization, necessitating a novel data analysis approach for the SPI data. In addition, I found a set of forced associations that perturb SPB duplication, resulting in elevated SPB number and, in some cases, multi-polar spindles. In order to study spatial aspects of MEN regulation in parallel to control of enzymatic activity by post-translational modifications I developed a compartmental, logical model of the MEN that is capable of representing both aspects of regulation. I found that my model is capable of correctly predicting the phenotype of 80% of mutants we tested, including mutants representing mislocalizing proteins. I use this model to uncover new aspects of MEN regulation, study the role of the FEAR (Cdc Fourteen Early Anaphase Release) network in determining anaphase length and investigate stochasticity in the Spindle Position Checkpoint (SPoC). Altogether this project presents a systematic view of which proteins are regulated through interaction with the SPB and how this functions as a mechanism of protein regulation.
Date of Award1 Dec 2020
Original languageEnglish
Awarding Institution
  • King's College London
SupervisorAttila Csikasz-Nagy (Supervisor)

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