Ligands of Inositol Polyphosphate Binding Proteins

Gilmartin, Megan (2022) Ligands of Inositol Polyphosphate Binding Proteins. Doctoral thesis, University of East Anglia.

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Abstract

In plants, cytosolic and organellar inositol polyphosphate (InsP) molecules are involved in multiple stress response pathways, such as drought tolerance and herbivory defence. The turnover and function of InsPs in vivo depend on enzymes including inositol phosphate kinases (IPKs) and phosphatases. InsP receptor proteins co-ordinate cellular responses dependent on the pool of InsPs present. Many IPKs localize to the nucleus, including AtIPK1 which is responsible for the synthesis of phytic acid, the implicated regulatory ligand of an mRNA transporter complex, a phytohormone co-receptor complex and precursor molecule to a subclass of InsPs, inositol pyrophosphates (PP-InsPs). Despite implications of IPKs in nuclear regulatory roles, these have not been described in plants. The primary research aim was to investigate a nuclear role of AtIPK1 through DNA binding assays (Chapter 2) and structure-function studies (Chapter 3).

SPX domains have recently been identified as PP-InsP ligand receptors which mediate plant phosphate homeostasis. A secondary aim of this project was to characterize a subset of Arabidopsis SPX domain proteins (AtSPX1, AtSPX3, AtPHT5;1), with a particular focus on substrate discrimination towards InsPs and PP-InsPs. To this end protein production, ligand-binding assays and structure-function studies were attempted (Chapter 4).

This thesis presents evidence for novel DNA binding of AtIPK1 and AtSPX1, whereby DNA binding activity is regulated by InsP titrations. In the application of in silico analyses, potential DNA binding sites were identified whilst low structural homology between these proteins and known DNA binding proteins were observed. A novel structure of wildtype apo AtIPK1 was solved lending new insights into conformational changes upon ligand binding. Additionally, AtSPX1 ligand binding was studied showing subtle differences of InsP preference in the absence of DNA interaction partner. Thus, demonstrating how InsP-binding proteins may exhibit additional regulatory functions which depend upon specific InsP substrate interactions.

Item Type:	Thesis (Doctoral)
Faculty \ School:	Faculty of Science > School of Biological Sciences
Depositing User:	Chris White
Date Deposited:	21 Mar 2023 08:42
Last Modified:	21 Mar 2023 08:42
URI:	https://ueaeprints.uea.ac.uk/id/eprint/91616
DOI:

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