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Title: Proteomics of sub-cellular protein distribution in oestrogen and tamoxifen stimulated MCF-7 breast cancer cells
Author: Alkhanjaf, A. A. M.
ISNI:       0000 0004 7230 8258
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2016
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Hormone receptor positive (HR+) breast cancer represent 70% of all breast tumours. Its oncogenesis is multiple step process thought to be driven by the presence of two transcription factors, the oestrogen receptor (ER) and/or the progesterone receptor (PR). Therefore, the endocrine-targeted therapeutic approach has been focused on both receptors as prognostic markers and therapeutic targets, aiming to alter the oestrogen signalling for patients with ERα-positive disease. The benefits gained from the treatment appear to be limited by developing either de novo or acquired resistance following a period of response to tamoxifen. In the present work, global quantitative proteomics was combined with the analysis of fractions enriched in target subcellular locations (nuclear and cytoplasmic fractions), this has allowed measurement of the changes in total abundance and in the compartmental abundance/distribution between the nucleus and cytoplasm for several thousand proteins differentially expressed in MCF-7cells in response to oestrogen and tamoxifen stimulation in MCF-7 cells. In the first part of the thesis, we have used a proteomics subcellular spatial razor approach to look at changes in total protein abundance and in protein distribution between the nucleus and cytoplasm following exposure of MCF7 breast cancer cells to oestradiol. The dominant response of MCF7 cells to oestrogen stimulation involves dynamic changes in protein subcellular spatial distribution rather than changes in total protein abundance. Of the 3604 quantitatively monitored proteins, only about 2% show substantial changes in total abundance (>2-fold), whereas about 20% of the proteins show substantial changes in local abundance and/or redistribution of their subcellular location, with up to 16-fold changes in their local concentration in the nucleus or the cytoplasm. The second part of the thesis focuses on the spatial distribution of proteins in the three sub-proteome fractions (total lysate, nuclear and cytoplasmic fractions) in 4-OHT stimulated MCF-7 cells. Of the 3493 quantitatively monitored proteins, 97 of quantified proteins were detected as a core data set with differential abundance (DA) that was significantly changed in total abundance and/or subcellular location (P < 0.05). More than 50% of the proteins show significant changes in local abundance and/or redistribution of their subcellular location, only 19 proteins show substantial changes in the total lysate. Following the rigorous downstream analysis of the pathways significantly overrepresented and GO terms significantly enriched, this analysis showed that protein changes in abundance with 4-OHT across multiple sample types (CNT) were involved in pathways related to metabolism, signal transduction, growth and proliferation, and development. The nuclear response involved upregulation of the proteins participating in the GPCR downstream signalling pathway leading to accumulated response in the cancer pathway by moderately modulating the cell survival (PI3K/AKT) pathway in a circadian rhythm. Furthermore, this was accompanied by substantial changes in the local abundance and /or the redistribution of the metastasis mediating proteins and apoptotic cleavages of cell adhesion proteins. We propose that dynamic redistribution of the subcellular location of multiple proteins in response to stimuli is a fundamental characteristic of cells and suggest that perturbation of cellular spatial control may be an important feature of cancer.
Supervisor: Godovac-Zimmermann, J. ; Walker, A. Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available