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Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins.

ABSTRACT: BACKGROUND:A limiting factor in performing proteomics analysis on cancerous cells is the difficulty in obtaining sufficient amounts of starting material. Cell lines can be used as a simplified model system for studying changes that accompany tumorigenesis. This study used two-dimensional gel electrophoresis (2DE) to compare the whole cell proteome of oral cancer cell lines vs normal cells in an attempt to identify cancer associated proteins. RESULTS:Three primary cell cultures of normal cells with a limited lifespan without hTERT immortalization have been successfully established. 2DE was used to compare the whole cell proteome of these cells with that of three oral cancer cell lines. Twenty four protein spots were found to have changed in abundance. MALDI TOF/TOF was then used to determine the identity of these proteins. Identified proteins were classified into seven functional categories - structural proteins, enzymes, regulatory proteins, chaperones and others. IPA core analysis predicted that 18 proteins were related to cancer with involvements in hyperplasia, metastasis, invasion, growth and tumorigenesis. The mRNA expressions of two proteins - 14-3-3 protein sigma and Stress-induced-phosphoprotein 1 - were found to correlate with the corresponding proteins' abundance. CONCLUSIONS:The outcome of this analysis demonstrated that a comparative study of whole cell proteome of cancer versus normal cell lines can be used to identify cancer associated proteins.

SUBMITTER: Karsani SA

PROVIDER: S-EPMC3974152 | biostudies-literature | 2014 Jan

REPOSITORIES: biostudies-literature

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Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins.

Karsani Saiful Anuar SA Saihen Nor Afiza NA Zain Rosnah Binti RB Cheong Sok-Ching SC Abdul Rahman Mariati M

Proteome science 20140115 1

<h4>Background</h4>A limiting factor in performing proteomics analysis on cancerous cells is the difficulty in obtaining sufficient amounts of starting material. Cell lines can be used as a simplified model system for studying changes that accompany tumorigenesis. This study used two-dimensional gel electrophoresis (2DE) to compare the whole cell proteome of oral cancer cell lines vs normal cells in an attempt to identify cancer associated proteins.<h4>Results</h4>Three primary cell cultures of ...[more]

PMID: 24422745

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Project description:BackgroundChemoresistance remains a significant challenge in chronic myelogenous leukemia (CML) management, which is one of the most critical prognostic factors. Elucidation the molecular mechanisms underlying the resistance to chemoresistance may lead to better clinical outcomes.ResultsIn order to identify potential protein targets involved in the drug-resistant phenotype of leukemia, especially the chronic myelogenous leukemia (CML), we used a high-resolution "ultra-zoom" 2DE-based proteomics approach to characterize global protein expression patterns in doxorubicin-resistant myelogenous leukemia cells compared with parental control cells. Ultra-high resolution of 2DE was achieved by using a series of slightly overlapping narrow-range IPG strips during isoelectric focusing (IEF) separation. A total number of 44 proteins with altered abundances were detected and identified by MALDI-TOF or LC-MS/MS. Among these proteins, enolase, aldolase, HSP70 and sorcin were up-regulated in doxorubicin-resistant myelogenous leukemia cell line, whereas HSP27 was down-regulated. Some of the results have been validated by Western blotting. Both enolase and aldolase were first reported to be involved in chemoresistance, suggesting that process of glycolysis in doxorubicin-resistant myelogenous leukemia cells was accelerated to some extent to provide more energy to survive chemical stress. Possible roles of most of the identified proteins in development of chemoresistance in myelogenous leukemia cells were fully discussed. The results presented here could provide clues to further study for elucidating the mechanisms underlying drug resistance in leukemia.ConclusionsAs a whole, under the chemical stress, the doxorubicin-resistant myelogenous leukemia cells may employ various protective strategies to survive. These include: (i) pumping the cytotoxic drug out of the cells by P-glycoprotein, (ii) increased storage of fermentable fuel, (iii) sophisticated cellular protection by molecular chaperones, (iv) improved handling of intracellular calcium, (v) increased glucose utilization via increased rates of glycolysis. In the present study, proteomic analysis of leukemia cells and their drug resistant variants revealed multiple alterations in protein expression. Our results indicate that the development of drug resistance in doxorubicin-resistant myelogenous leukemia cells is a complex phenomenon undergoing several mechanisms.

Project description:Helicobacter pylori (H. pylori) is a spiral-shaped Gram-negative bacterium that causes the most common chronic infection in the human stomach. Approximately 1%-3% of infected individuals develop gastric cancer. However, the mechanisms by which H. pylori induces gastric cancer are not completely understood. The available evidence indicates a strong link between the virulence factor of H. pylori, cytotoxin-associated gene A (CagA), and gastric cancer. To further characterize H. pylori virulence, we established three cell lines by infecting the gastric cancer cell lines SGC-7901 and AGS with cagA+ H. pylori and transfecting SGC-7901 with a vector carrying the full-length cagA gene. We detected 135 differently expressed proteins from the three cell lines using proteome technology, and 10 differential proteins common to the three cell lines were selected and identified by LC-MS/MS as well as verified by western blot: β-actin, L-lactate dehydrogenase (LDH), dihydrolipoamide dehydrogenase (DLD), pre-mRNA-processing factor 19 homolog (PRPF19), ATP synthase, calmodulin (CaM), p64 CLCP, Ran-specific GTPase-activating protein (RanGAP), P43 and calreticulin. Detection of the expression of these proteins and genes encoding these proteins in human gastric cancer tissues by real-time PCR (RT-qPCR) and western blot revealed that the expression of β-ACTIN, LDH, DLD, PRPF19 and CaM genes were up-regulated and RanGAP was down-regulated in gastric cancer tissues and/or metastatic lymph nodes compared to peri-cancerous tissues. High gene expression was observed for H. pylori infection in gastric cancer tissues. Furthermore, the LDH, DLD and CaM genes were demethylated at the promoter -2325, -1885 and -276 sites, respectively, and the RanGAP gene was highly methylated at the promoter -570 and -170 sites in H. pylori-infected and cagA-overexpressing cells. These results provide new insights into the molecular pathogenesis and treatment targets for gastric cancer with H. pylori infection.

Dataset Information

Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins.

Publications

Comparative proteomics analysis of oral cancer cell lines: identification of cancer associated proteins.

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