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Title: The spatial and temporal regulation of the monomeric G protein, Ras 1
Author: Weston, Cathryn A.
ISNI:       0000 0004 2749 3335
Awarding Body: University of Warwick
Current Institution: University of Warwick
Date of Award: 2013
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A vast array of cellular processes are controlled by the highly conserved guanine nucleotide-regulated molecular switches, such as Ras. The spatial and temporal regulation of Ras signalling, both positively, through the action of guanine nucleotide exchange factors (GEFs) and negatively, via interaction with GTPase activating proteins (GAPs) is critical to maintain cell viability. Due to the presence of multiple Ras isoforms, regulatory proteins and effectors, the study of Ras signalling in higher eukaryotes is technically challenging. Fission yeast provides a simple system in which to study Ras signalling containing a single Ras homologue (Ras1) that modulates two distinct processes. Ras1 signalling is terminated through GTP-hydrolysis accelerated by the action of a GAP, Gap1. This thesis describes the development of a quantitative FRET assay to visualise the dynamics of Ras1 activity within single cells. Using this reporter, in combination with a variety of other quantitative techniques, the role of the negative regulator during signal transduction was explored. These results highlighted an important difference between the requirement for GTPhydrolysis during signal propagation. Finally, use of the FRET reporter revealed an alternative G protein state, within the Ras1 activation cycle. These data question the accepted models of G protein signal propagation and prompt further investigation into the roles of GTP-hydrolysis in signal transduction. Several positive regulators of Ras1 have previously been described in fission yeast. These proteins have been proposed to direct signalling via the different Ras1 effectors. Using quantitative image analysis and pheromoneresponsive reporter strains, this thesis presents data to suggest that it is not simply a one-activator-one-effector response. Further, the role of the previously uncharacterised, scaffold protein, Ral2 in mediating signal transduction is also investigated. Finally the possibility that the heterotrimeric G protein, Gpa1 can directly activate MAPK signalling to provide a pheromone-sensing mechanism within fission yeast is discussed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available
Keywords: QP Physiology