Deciphering the role of liquid-liquid phase separation in oncogenic gene expression

Walsh, Peter (2024) Deciphering the role of liquid-liquid phase separation in oncogenic gene expression. PhD thesis, University of Glasgow.

Abstract

Liquid-liquid phase separation is a biophysiochemical process that drives the formation of membraneless organelles, termed biomolecular condensates. Stress granules are a biomolecular condensate that form in the cytoplasm in response to stress. Stress granules contain dense networks of RNA and protein. Although much research into stress granules has been conducted, little about their function in regulating translation is known. Stress granules have been implicated in pathophysiology of multiple human diseases including cancer.
Two physiologically relevant stresses in cancer were used for induction of stress granules. Oxidative stress using sodium arsenite and hypoxia. Morphologically distinct granules formed in response to hypoxia or sodium arsenite. Canonical well defined cytoplasmic foci formed in response to sodium arsenite stress with docking P-bodies on their surface. In response to hypoxia, larger nebulous stress granules deficient in docking P-bodies formed. Distinct signalling cascades regulate stress granule formation in each stress. Sodium arsenite induces stress granule formation through phosphorylation of eIF2α, whereas reduced mTOR activity and hypophosphorylation of 4E-BP1 regulates stress granule assembly in hypoxia. Proteomic analysis of stress granules revealed that the DEAD box helicases eIF4A1 and DDX6 were differentially enriched in hypoxia and sodium arsenite respectively. eIF4A1 is a known regulator of stress granule P-body interactions and inhibition of eIF4A1 reduced stress granule size and rescued P-body docking in hypoxia. Proteomic analysis also revealed an enrichment of large ribosomal subunit proteins in hypoxia induced stress granules.
RNA sequencing of stress granules revealed that mRNA localisation is largely stress dependent as distinct subsets of mRNAs with distinct sequence features are recruited to stress granules in a stress dependent manner. Integration of stress granule sequencing with ribosome profiling data revealed that stress granule mRNAs are translationally repressed in response to sodium arsenite in concordance with previously published data sets. Intriguingly stress granule mRNAs were translationally upregulated in response to hypoxia. This taken with the localisation of components of the large ribosomal subunit proteins suggests that hypoxia induced stress granules are a more translationally active compartment than what has previously been described in response to eIF2α phosphorylation.

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Subjects: R Medicine > RC Internal medicine > RC0254 Neoplasms. Tumors. Oncology (including Cancer)
• Colleges/Schools: College of Medical Veterinary and Life Sciences > School of Cancer Sciences
• Supervisor's Name: Bushell, Professor Martin
• Date of Award: 2024
• Depositing User: Theses Team
• Unique ID: glathesis:2024-84043
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 22 Jan 2024 14:30
• Last Modified: 24 Feb 2026 09:48
• Thesis DOI: 10.5525/gla.thesis.84043
• URI: https://theses.gla.ac.uk/id/eprint/84043