EGF-induced FoxO3a nuclear-cytoplasmic pulsing dynamics in mammary epithelial cell lines following treatment with increasing doses of a MEK inhibitor. Dataset 3 of 3: mean reporter pulsing metrics.
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ABSTRACT:
SUBMITTER: Somponnat Sampattavanich, Bernhard Steiert
Project description:Cylindromatosis tumor suppressor protein (CYLD) is deubiquitinase, best known as an essential negative regulator of the NFkB pathway. Previous studies have suggested an involvement of CYLD in epidermal growth factor (EGF)-dependent signal transduction as well, as it was found enriched within the tyrosine-phosphorylated complexes in cells stimulated with the growth factor. EGF receptor (EGFR) signaling participates in central cellular processes and its tight regulation, partly through ubiquitination cascades, is decisive for a balanced cellular homeostasis. Here, using a combination of mass spectrometry-based quantitative proteomic approaches with biochemical and immunofluorescence strategies, we demonstrate the involvement of CYLD in the regulation of the ubiquitination events triggered by EGF. Our data show that CYLD regulates the magnitude of ubiquitination of several major effectors of the EGFR pathway by assisting the recruitment of the ubiquitin ligase Cbl-b to the activated EGFR complex. Notably, the deficiency of CYLD impairs the interaction of EGFR with Cbl-b, which leads to a reduced ubiquitination of the receptor followed by its decreased degradation. This represents a previously uncharacterized strategy exerted by this deubiquitinase and tumors pressor for the negative regulation of a tumorigenic signaling pathway.
Project description:Phosphoproteomic analysis of EGF stimulated MCF7 cells. This dataset is the basis for the development of modeling of signaling networks. A fundamental challenge in biology is to delineate the signaling pathways that govern cellular responses to genetic and environmental cues. Phosphoproteomics is an emerging technology that provides key data on activity levels of proteins under conditions of interest. However, the interpretation of these data is hampered by the lack of methods that can translate site-specific information into global maps of active proteins and signaling networks. To meet this challenge, we propose PHOTON, a method for integrating phosphorylation data with protein-protein interaction networks to identify active proteins and pathways and pinpoint functional phosphosites. We demonstrate the utility of PHOTON by applying it to interpret existing and novel phosphoproteomic datasets related to EGF and insulin responses. PHOTON substantially outperforms the widely-used cutoff approach, providing highly reproducible predictions that are more in line with current biological knowledge
Project description:In the present work we propose a new therapy for NRAS mutant melanoma. Simultaneous inhibition of MEK and ROCK caused induction of BimEL , PARP, and Puma, and hence apoptosis. In vivo, MEK and ROCK inhibition suppressed growth of established tumors. Our findings warrant clinical investigation of the effectiveness of combinatorial targeting of MAPK/ERK and ROCK in NRAS mutant melanoma.
Project description:Mutations in BRAF are common to many cancers, including CRC. The MEK inhibitors are being investigated in BRAF-mutant CRC. In this study, we aimed to investigate how MEK inhibitor suppresses growth of BRAF-mutated CRC cells as well as its potential mechanisms. Our findings indicated that MEK inhibitor promote PUMA expression via ERK/FoxO3a signaling pathway. In addition, PUMA induction is essential for MEK inhibitor-induced apoptosis. Moreover, PUMA induction is required for MEK inhibitors to induced apoptosis in combination with cisplatin, dabrafenib, or Gefitinib. Knockdown of PUMA suppressed the anticancer effect of the MEK inhibitor in vivo. Our findings indicate a novel role for PUMA as a regulator of the antitumor effects of MEK inhibitor, suggesting that PUMA induction may modulate MEK inhibitor sensitivity.
Project description:ELK1 is a well-known target of the ERK branch (EGF-responsive) of the MAP kinase pathway. This transcription profiling experiment studied the effects of ELK1 depletion by siRNA and subsequent EGF stimulation.