Exploring integrin and growth factor receptor crosstalk mechanisms



Thomas, J
(2018) Exploring integrin and growth factor receptor crosstalk mechanisms. PhD thesis, University of Liverpool.

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Abstract

Abstract Joanna R. Thomas; Exploring integrin and growth factor receptor crosstalk mechanisms. The pro-invasive integrin αvβ6 is upregulated in a range of carcinomas, from normally very low levels, and is associated with poor prognosis. Therapeutic targeting of αvβ6 with inhibitory antibodies has produced promising results in breast cancer models in vivo. Preclinical evidence has suggested that αvβ6 and EGFR expression correlate in disease, and that a co-operative relationship between αvβ6 and EGFR may exist in vivo. This study aimed to determine if αvβ6 and EGFR are functionally integrated, and to investigate wider mechanisms of integrin and growth factor receptor crosstalk. Immunofluorescent imaging, under steady-state conditions, demonstrated co-localisation between αvβ6 and EGFR in MDA-MB-468 cells in protrusive structures and endosomes. In addition, imaging and integrin-associated complex (IAC) enrichment approaches revealed that EGFR was recruited to sites of αvβ6 engagement on the αvβ6-selective ligand, latency-associated peptide (LAP). Importantly, reciprocal EGF or LAP stimulation increased EGFR and αvβ6 co-localisation with early endosomes, indicative of co-internalisation. Together, these data suggest a co-regulatory relationship between αvβ6 integrin and EGFR that impacts receptor trafficking mechanisms. Stimulation with LAP caused the phosphorylation of ERK, indicating αvβ6 positively regulates MAPK pathway signalling, and that signalling crosstalk may also exist between αvβ6 and EGFR. Bioinformatic interrogation of αvβ6-IACs revealed αvβ6-medated adhesions are conducive sites of EGFR signalling. The identification of proteins associated with endocytosis and receptor trafficking highlighted liprin α1 as an important potential mediator of αvβ6 and EGFR crosstalk. The over-representation of Hippo pathway regulators at αvβ6-IACs led to on-going work that has identified αvβ6 as a regulator of YAP nuclear localisation and suggests that αvβ6-EGFR crosstalk is a determinant of this function. Proteomic analysis of EGF regulated IAC components led to the identification of the adaptor protein Eps8 as a novel mediator of integrin and EGFR crosstalk. Eps8 functions to constrain endocytosis of α5β1 and EGFR and is required for adhesion organisation. These functions have direct consequences for adhesion and cytoskeletal dynamics. On-going analysis of candidates identified from αvβ6-IAC proteomics will elucidate how αvβ6 and EGFR signalling networks and trafficking are integrated. In the future we aim to determine how these mechanisms contribute to αvβ6-dependent breast cancer cell invasion, and how αvβ6-EGFR receptor crosstalk understanding may contribute to the success of αvβ6-targeted therapeutics.

Item Type: Thesis (PhD)
Divisions: Faculty of Health and Life Sciences > Faculty of Health and Life Sciences
Depositing User: Symplectic Admin
Date Deposited: 21 Nov 2018 15:52
Last Modified: 02 Apr 2021 07:18
DOI: 10.17638/03025717
Supervisors:
URI: https://livrepository.liverpool.ac.uk/id/eprint/3025717