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Title: Developing Dictyostelium discoideum as a model to study the synthesis and metabolism of inositol pyrophosphates and inorganic polyphosphate
Author: Livermore, T. M.
ISNI:       0000 0004 7230 3633
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2016
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Inositol pyrophosphates and inorganic polyphosphate (polyP) represent two of the most phosphate-rich molecules in the cell. However, the synthesis, metabolism and function of these “high-energy” molecules are not well understood. This work develops the amoeba, Dictyostelium discoideum, as a genetic model to study both classes of molecule. By applying improved polyacrylamide gel electrophoresis (PAGE) techniques, unlabelled inositol pyrophosphates and polyP extracted from D. discoideum have been visualised for the first time. Deletion of a range of inositol phosphate kinases reveals a complex synthetic pathway of the inositol pyrophosphates in the amoeba. Surprisingly, strains lacking the majority of inositol pyrophosphates do not display a dramatic phenotype. However, these cells retain low levels of inositol pyrophosphates. In fact, this work identified at least four enzymes capable of synthesising inositol pyrophosphates in the amoeba, including the novel inositol phosphate kinases IPKA and IPKB. In vitro, both IPKA and IPKB act as specific IP 7 kinases. However, in vivo both enzymes are catalytically flexible and are able to phosphorylate other substrates. In addition, PAGE technology allows the detection of polyP extracted from D. discoideum, revealing the unprecedented increase of polyP during development. Deletion of the Polyphosphate kinase (PPK)1 gene confirms this to be the only PPK in the amoeba. Deletion of PPK has a negative effect on the fitness of the amoeba, a phenotype underpinned by the ability of polyP to regulate primary metabolism. The ppk1-null strain fails to accumulate polyP during development and is defective in spore germination. Loss of polyP during the development of ppk1 is partially compensated by an increase in inositol pyrophosphates. These observations support a model in which there is a functional interplay between inositol pyrophosphates, polyP and primary metabolism.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available