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Proteomic profiling of Breast cell lines exhibiting epithelial or mesenchymal morphology

ABSTRACT: We performed quantitative TMT-based proteomic analysis of eight human-derived breast cell lines growing in 2D. The dataset includes the total and phospho-proteomes of seven tumour cell lines: T47D (Epithelial, Luminal, ER+/PR+/HER2-), BT474 (Epithelial, Luminal, ER+/PR+/HER2+), SKBR3 (Epithelial, Luminal, ER-/PR-/HER2+), MDA-MB-468 (Epithelial, Basal A, ER-/PR-/HER2-), MDA-MB-231 (Mesenchymal, Basal B, ER-/PR-/HER2-), MDA-MB-231-LM2 (Mesenchymal, Basal B, ER-/PR-/HER2-, a highly metastatic subpopulation 4175 from MDA-MB-231), and SUM159 (Mesenchymal, Basal B, ER-/PR-/HER2-) and one non-tumour cell line: MCF10A (Epithelial, Basal B, ER-/PR-/HER2-). We achieved a depth of over 8,000 proteins and a total of over 20,000 phosphopeptides with MS2 acquisition covering a wide range of biological processes. These eight lines encompass a wide range of genetic subtypes and have diverse morphologies in either 2D (epithelial vs mesenchymal), and/or in 3D environments (Mass, Grape-like, Stellate, Round) (Neve RM, 2006) (Kenny PA, 2007). The study also explored the effects of TGF-β1 during 10 days on the non-tumorigenic MCF10A cells, and their posterior washout and recovery for 40 days. TGF-β1 is a cytokine known to induce epithelial-to-mesenchymal transition (EMT), a process associated with cancer progression (Xu J, 2009) (Nieto, 2016). The data can be analysed in isolation or in combination with other orthogonal datasets such as transcriptomes or image-based data (OMICS), for the purpose of predicting tumour outcome, identifying markers, drug response or mechanisms of resistance to therapeutics. These data can provide insights for therapeutic strategies and better understanding of the diverse molecular landscapes of breast cancer cells.

INSTRUMENT(S): Orbitrap Fusion Lumos

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Graeme Benstead-Hume

LAB HEAD: Jyoti Choudhary

PROVIDER: PXD047164 | Pride | 2024-09-16

REPOSITORIES: Pride

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			Action	DRS
	Req102-MB-Bakal_18plex_TP_01.raw	Raw
	Req102-MB-Bakal_18plex_TP_02.raw	Raw
	Req102-MB-Bakal_18plex_TP_03.raw	Raw
	Req102-MB-Bakal_18plex_TP_04.raw	Raw
	Req102-MB-Bakal_18plex_TP_05.raw	Raw

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Project description:Breast cancer is a heterogeneous disease comprised of at least five major subtypes. Luminal subtype tumors confer a more favourable patient prognosis, which is in part, attributed to the Estrogen Receptor-alpha (ER) positivity and anti-hormone responsiveness of these tumors. Expression of the forkhead box transcription factor, FOXA1, also correlates with the luminal subtype and patient survival, but is present in a subset of ER-negative tumors. Similarly, FOXA1 is consistently expressed in luminal breast cancer cell lines even in the absence of ER. In contrast, basal breast cancer cell lines do not express FOXA1, and loss of FOXA1 in luminal cells increases migration and invasion, characteristics of the basal subtype. To delineate an ER-independent role for FOXA1 in maintaining the luminal phenotype, and hence a more favourable prognosis, we performed cDNA microarray analyses on luminal FOXA1-positive, ER-positive (MCF7, T47D) and FOXA1-positive, ER-negative (MDA-MB-453, SKBR3) cell lines in the presence or absence of transient FOXA1 silencing. This resulted in three FOXA1 transcriptomes: (1) a luminal-signature (consistent across cell lines), (2) an ER-positive signature (restricted to MCF7 and T47D) and (3) an ER-negative signature (restricted to MDA-MB-453 and SKBR3). Use of Gene Set Enrichment Analyses (GSEA) as a phenotyping tool revealed that FOXA1 silencing resulted in a transcriptome shift from luminal to basal gene expression signatures. FOXA1 binds to both luminal and basal genes within luminal breast cancer cells, suggesting that it not only transactivates luminal genes, but also represses basal-associated genes. From these results we conclude that FOXA1 controls plasticity between basal and luminal cells, playing a dominant role in repressing the basal phenotype, and thus tumor aggressiveness, in luminal breast cancer cells. Although it has been proposed that FOXA1-targeting agents may be useful for treating luminal tumors, these data suggest that this approach may promote transitions toward a more aggressive cancer. FOXA1 siRNA treated breast cell lines compared directly to nonspecific siRNA treated cell lines using Agilent 4X44 microarrays.

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Proteomic profiling of Breast cell lines exhibiting epithelial or mesenchymal morphology

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