Dataset Information

Integrative analysis of proteome and ubiquitylome reveals unique features of protein ubiquitylation and mitochondria in gefitinib resistant non-small cell lung cancer cells

ABSTRACT: Even though gefitinib, a tyrosine kinase inhibitor, widely used in treating non-small cell lung cancer (NSCLC) patients carrying EGFR activating mutations, most patients will develop gefitinib resistance. Protein ubiquitylation is one of major posttranslational modifications affecting the stability or function of the protein. However, the role of protein ubiquitylation in gefitinib resistance is poorly understood. To systematically identify the global change in protein ubiquitylation during gefitinib resistance, we carried out quantitative global proteome and ubiquitylome study by using cells with stable isotopic labeling with amino acid in cell culture (SILAC) followed by affinity enrichment of ubiquitylated peptides and subsequent liquid chromatography–tandem mass spectrometry (LC-MS/MS) analysis. Our study quantified are differentially regulated in ubiquitylation between gefitinib resistant and sensitive cells. Among them, 799 lysine sites were up-regulated in ubiquitylation in resistant cells, which are enriched in pathways, such as SNARE interaction in vesicle transport, endocytosis, phagosome, and lysosome, etc. In addition, we also found cells. The proteins carry these sites are involved in pathways including metabolic pathways, gap junction, and biosynthesis of amino acids, etc, This data indicate that gefitinib resistance dramatically alters the landscape of ubiquitylation in the cells. Furthermore, by integrating the genome-wide transcriptome data, we discovered that complexes I and IV in the electron transport chain of the mitochondria are up-regulated, while complexes II and III are down-regulated in resistant cells. Overall, these results reveal the unique features of ubiquitylation and mitochondria during gefitinib resistance, which will help to a better understanding of the role(s) of ubiquitylation, and to identify new therapeutic targets in overcoming gefitinib resistance.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture, Smooth Muscle Cell Of Small Intestine

DISEASE(S): Lung Cancer

SUBMITTER: zhu taotao

LAB HEAD: Zhang Fan

PROVIDER: PXD004941 | Pride | 2018-07-05

REPOSITORIES: Pride

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Integrative Analysis of Proteome and Ubiquitylome Reveals Unique Features of Lysosomal and Endocytic Pathways in Gefitinib-Resistant Non-Small Cell Lung Cancer Cells.

Li Wang W Wang Heyong H Yang Yan Y Zhao Tian T Zhang Zhixiong Z Tian Ye Y Shi Zhaomie Z Peng Xiaojun X Li Fei F Feng Yonghong Y Zhang Lei L Jiang Gening G Zhang Fan F

Proteomics 20180708 15

Non-small cell lung cancer (NSCLC) patients carrying EGFR activating mutations treated with gefitinib, a tyrosine kinase inhibitor, will develop drug resistance. Ubiquitylation is one of major posttranslational modifications of proteins affecting the stability or function of proteins. However, the role of protein ubiquitylation in gefitinib resistance is poorly understood. To systematically identify the global change in protein expression and ubiquitylation during gefitinib resistance, a quantit ...[more]

PMID: 29901268

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Project description:Despite initial and often dramatic responses of epidermal growth factor receptor (EGFR)-addicted lung tumors to the EGFR-specific tyrosine kinase inhibitors (TKIs), gefitinib and erlotinib, nearly all develop resistance and relapse. To explore novel mechanisms mediating acquired resistance, we employed non-small-cell lung cancer (NSCLC) cell lines bearing activating mutations in EGFR and rendered them resistant to EGFR-specific TKIs through chronic adaptation in tissue culture. In addition to previously observed resistance mechanisms including EGFR-T790M 'gate-keeper' mutations and MET amplification, a subset of the seven chronically adapted NSCLC cell lines including HCC4006, HCC2279 and H1650 cells exhibited marked induction of fibroblast growth factor (FGF) 2 and FGF receptor 1 (FGFR1) mRNA and protein. Also, adaptation to EGFR-specific TKIs was accompanied by an epithelial to mesenchymal transition (EMT) as assessed by changes in CDH1, VIM, ZEB1 and ZEB2 expression and altered growth properties in Matrigel. In adapted cell lines exhibiting increased FGF2 and FGFR1 expression, measures of growth and signaling, but not EMT, were blocked by FGFR-specific TKIs, an FGF-ligand trap and FGFR1 silencing with RNAi. In parental HCC4006 cells, cell growth was strongly inhibited by gefitinib, although drug-resistant clones progress within 10 days. Combined treatment with gefitinib and AZD4547, an FGFR-specific TKI, prevented the outgrowth of drug-resistant clones. Thus, induction of FGF2 and FGFR1 following chronic adaptation to EGFR-specific TKIs provides a novel autocrine receptor tyrosine kinase-driven bypass pathway in a subset of lung cancer cell lines that are initially sensitive to EGFR-specific TKIs. The findings support FGFR-specific TKIs as potentially valuable additions to existing targeted therapeutic strategies with EGFR-specific TKIs to prevent or delay acquired resistance in EGFR-driven NSCLC. Examination of mRNA levels in DMSO and gefitinib-resistant cultures of HCC4006 and HCC827. Each group has two replicates.

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