Identification of KSR1 as a novel target and decoding tyrosine kinase proteome in breast cancer
File(s)
Author(s)
Zhang, Hua
Type
Thesis
Abstract
Kinase suppressor of Ras-1 (KSR1), originally identified as a novel protein kinase in the Ras-Raf cascade, plays a role in activation of mitogen-activated protein kinases (MAPKs). Although efforts have been devoted to study the role of KSR1 in certain tumour types, its biological functions in breast cancer have remained largely undefined.
A SILAC-based proteomic analysis was conducted to identify the KSR1-regulated phosphoproteins in breast cancer. Our results revealed that KSR1 overexpression decreases deleted in breast cancer 1 (DBC1) phosphorylation. We then demonstrated that KSR1 decreases transcriptional activity of p53 by reducing phosphorylation of DBC1, which leads to a reduced interaction of DBC1 with sirtuin 1 (SIRT1); this in turn enables SIRT1 to deacetylate p53. We further examined the correlation between KSR1 expression and clinical outcome in breast cancer. Our results showed that patients with breast cancer with high KSR1 in our cohort (n>1000) had better disease free- and overall survival. Moreover, in KSR1-transfected stable cells, fewer and smaller size colonies were formed in comparison to parental cells, while an in vivo study demonstrated that the growth of xenograft tumours overexpressing KSR1 was inhibited. Mechanistically, the tumour suppressive action of KSR1 is BRCA1 dependent,
which was shown by in vitro 3D matrigel and soft agar assays. KSR1 regulates BRCA1 ubiquitination through elevated BRCA1-associated RING domain 1 (BARD1) expression and increased BRCA1-BARD1 interaction.
Deregulation of tyrosine kinases (TKs) signalling can contribute to tumourigenesis. A combined approach of RNAi and SILAC-based quantitative proteomics was employed to decode the TKs-regulated proteomes upon silencing individually each 65 validated TKs in MCF7 breast cancer cells. Bioinformatics analysis identified 10 new distinctive clusters based on similarity in the TKs-regulated proteomes. The biological relevance of our proteomic study in interpreting the TKs-regulated proteomes supports the essential role of TKs in regulating
all aspects of cellular activities.
A SILAC-based proteomic analysis was conducted to identify the KSR1-regulated phosphoproteins in breast cancer. Our results revealed that KSR1 overexpression decreases deleted in breast cancer 1 (DBC1) phosphorylation. We then demonstrated that KSR1 decreases transcriptional activity of p53 by reducing phosphorylation of DBC1, which leads to a reduced interaction of DBC1 with sirtuin 1 (SIRT1); this in turn enables SIRT1 to deacetylate p53. We further examined the correlation between KSR1 expression and clinical outcome in breast cancer. Our results showed that patients with breast cancer with high KSR1 in our cohort (n>1000) had better disease free- and overall survival. Moreover, in KSR1-transfected stable cells, fewer and smaller size colonies were formed in comparison to parental cells, while an in vivo study demonstrated that the growth of xenograft tumours overexpressing KSR1 was inhibited. Mechanistically, the tumour suppressive action of KSR1 is BRCA1 dependent,
which was shown by in vitro 3D matrigel and soft agar assays. KSR1 regulates BRCA1 ubiquitination through elevated BRCA1-associated RING domain 1 (BARD1) expression and increased BRCA1-BARD1 interaction.
Deregulation of tyrosine kinases (TKs) signalling can contribute to tumourigenesis. A combined approach of RNAi and SILAC-based quantitative proteomics was employed to decode the TKs-regulated proteomes upon silencing individually each 65 validated TKs in MCF7 breast cancer cells. Bioinformatics analysis identified 10 new distinctive clusters based on similarity in the TKs-regulated proteomes. The biological relevance of our proteomic study in interpreting the TKs-regulated proteomes supports the essential role of TKs in regulating
all aspects of cellular activities.
Version
Open Access
Date Issued
2014-04
Date Awarded
2014-07
Copyright Statement
Attribution NoDerivatives 4.0 International Licence (CC BY-ND)
Advisor
Stebbing, Justin
Giamas, Georgios
Publisher Department
Department of Surgery & Cancer
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)