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Quantitative proteomic and phospho-proteomic analysis of human DLD1 cells differing in ploidy and chromosome stability

Viganò, Cristina. Quantitative proteomic and phospho-proteomic analysis of human DLD1 cells differing in ploidy and chromosome stability. 2017, Doctoral Thesis, University of Basel, Faculty of Science.

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Official URL: http://edoc.unibas.ch/diss/DissB_12973

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Abstract

Aneuploidy is a state in which cells harbor a chromosome number that is not a whole multiple of the haploid chromosome set. This condition is poorly tolerated during embryogenesis and it the cause of developmental disorders such as Down syndrome (trisomy 21). Beside, aneuploidy is often associated with whole chromosomal instability (CIN), a constant chromosome mis-segregation ongoing from one cell division to the next. Aneuploidy and CIN are a common hallmark of many cancers, even if to date, the cellular processes involved in aneuploidization and tumorigenesis are poorly understood. This raises the questions of how CIN originates, how it is tolerated at the cellular level, and which cellular pathways are involved in this tolerance. In order to try to solve these questions, I performed a comprehensive proteomic analysis of cancer cell lines with different karyotypic and chromosome stability states. I have compared stable isogenic diploid and tetraploid colon cancer cell lines with descendant unstable aneuploid post-tetraploid (PTAs) and engineered trisomic clones. By applying quantitative mass-spectrometric approaches, I was able to identify the relative abundance of around 7’500 and 6’000 proteins across PTAs and trisomic clones, respectively. Analysis of proteomic data allowed me to conclude that most changes of protein abundance and phosphorylation, present in aneuploid clones, already occur after chromosome mass increase, i.e. the transition to the tetraploid state, rather than the presence of CIN. In particular I observed the deregulation of pathways involved in protein folding, proteolysis and response to oxidative stress. Additionally, in order to identify possible modifications in protein activity, I performed phospho-enrichment analysis in the generated cell lines, and this resulted in the identification of 13’500 and 9’000 phospho-peptides in PTAs and trisomic clones, respectively. Importantly, while a large number of proteins previously associated with CIN and cell cycle remained largely unaltered in their expression levels (compared with the parental diploid line), their phosphorylation levels showed substantial difference. Most interestingly, I observed a higher phosphorylation state at specific activation sites of key mitotic protein kinases, notably Aurora A and Plk1. Consequentially, tetraploid and post-tetraploid clones showed similar sensitivity profiles in a chemotherapeutic drug screen, notably increased sensitivity to several Plk1 and Aurora A inhibitors.
These results suggest that in transformed cancer cells, a gain in chromosome number, rather than an increased chromosome mis-segregation rate, triggers a clonal stress response at the protein level. Moreover, these results indicate that chromosome gains lead to activation or deactivation of pathways involved in cell division and mitosis primarily through hyper- or hypo-phosphorylation, rather than massive changes in protein expression. Being able to identify deregulated pathways in response to chromosome mass increase or instability may provide new opportunities to specifically targets cancer cells and block disease progression. Results from our drug screening approach, although preliminary, support this notion. They suggest that a common sensitivity profile may exist across aneuploid and polyploid cells, raising the prospect of new treatment strategies for tumors harboring a large excess of chromosomes.
Advisors:Nigg, Erich A. and Hall, Michael N.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Former Organization Units Biozentrum > Cell Biology (Nigg)
UniBasel Contributors:Vigano, Cristina and Nigg, Erich A. and Hall, Michael N.
Item Type:Thesis
Thesis Subtype:Doctoral Thesis
Thesis no:12973
Thesis status:Complete
Bibsysno:Link to catalogue
Number of Pages:1 Online-Ressource (126 Seiten)
Language:English
Identification Number:
Last Modified:15 Nov 2019 14:02
Deposited On:26 Mar 2019 13:51

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