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Mitochondrial proteomics of nasopharyngeal carcinoma metastasis.

Liu J, Zhan X, Li M, Li G, Zhang P, Xiao Z, Shao M, Peng F, Hu R, Chen Z - BMC Med Genomics (2012)

Bottom Line: The functional enrichment analyses of DEPs discovered five significant biological processes including cellular response to reactive oxygen species, hydrogen peroxide metabolic process, regulation of mitochondrial membrane potential, cell redox homeostasis and oxidation reduction, and five significant molecular functions including oxidoreductase activity, caspase inhibitor activity, peroxiredoxin activity, porin activity and antioxidant activity.Those 5-8F cells with suppression of PRDX3 showed an increased mobility potential.These data suggest that those mitochondrial DEPs are potential biomarkers for NPC metastasis, and their dysregulation would play important roles in mitochondria oxidative stress-mediated NPC metastatic process.

View Article: PubMed Central - HTML - PubMed

Affiliation: Key Laboratory of Cancer Proteomics of Chinese Ministry of Health, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan, 410008, PR China.

ABSTRACT

Background: Mitochondrial proteomic alterations of nasopharyngeal carcinoma metastasis remain unknown. Our purpose is to screen mitochondrial proteins for the elucidation of the molecular mechanisms of nasopharyngeal carcinoma metastasis and the discovery of metastasis-related biomarkers.

Methods: Mitochondria were isolated from nasopharyngeal carcinoma metastatic (5-8F) and nonmetastatic (6-10B) cell lines, respectively. After characterization of isolated mitochondria, mitochondrial differentially expressed proteins (DEPs) were quantified by two-dimensional difference in-gel electrophoresis (2D-DIGE), and identified by peptide mass fingerprint (PMF) and tandem mass spectrometry (MS/MS). A functional enrichment analysis and a protein-protein interaction sub-network analysis for DEPs were carried out with bioinformatics. Furthermore, siRNAs transient transfections were used to suppress expressions of some up-regulated DEPs in metastatic cells (5-8F), followed by Transwell Migration assay.

Results: Sixteen mitochondrial DEPs including PRDX3 and SOD2 were identified. Those 5-8F cells with suppression of PRDX3 showed an increased mobility potential. The functional enrichment analyses of DEPs discovered five significant biological processes including cellular response to reactive oxygen species, hydrogen peroxide metabolic process, regulation of mitochondrial membrane potential, cell redox homeostasis and oxidation reduction, and five significant molecular functions including oxidoreductase activity, caspase inhibitor activity, peroxiredoxin activity, porin activity and antioxidant activity. A protein-protein interaction sub-network of DEPs was generated with literature data. Ten mitochondrial DEPs including PRDX3, PRDX6, SOD2, ECH1, SERPINB5, COX5A, PDIA5, EIF5A, IDH3B, and PSMC4 were rationalized in the tumor-stroma co-evolution model that mitochondrial oxidative stress directly contributes to tumor metastasis.

Conclusions: Sixteen mitochondrial DEPs were identified with mass spectrometry and ten of them were rationalized in the tumor-stroma co-evolution model. Those 5-8F cells with suppression of PRDX3 showed an increased mobility potential. These data suggest that those mitochondrial DEPs are potential biomarkers for NPC metastasis, and their dysregulation would play important roles in mitochondria oxidative stress-mediated NPC metastatic process.

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Up-regulated expressions of PRDX3 in metastatic 5-8F cell mitochondria relative to nonmetastatic 6-10B cell mitochondria without significant expression difference in the level of whole cells.A. Western blotting image. B. The quantitative comparison of the digitized Western blotting image. COXIV, a marker protein of mitochondria, was used as a reference standard protein when mitochondrial protein sample was analyzed with Western blot. ACTIN that is mainly expressed in the cytoplasm was used a reference standard protein when whole-cell protein sample was analyzed with Western blot. Western blot experiments were carried out triply (n = 3).
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Figure 6: Up-regulated expressions of PRDX3 in metastatic 5-8F cell mitochondria relative to nonmetastatic 6-10B cell mitochondria without significant expression difference in the level of whole cells.A. Western blotting image. B. The quantitative comparison of the digitized Western blotting image. COXIV, a marker protein of mitochondria, was used as a reference standard protein when mitochondrial protein sample was analyzed with Western blot. ACTIN that is mainly expressed in the cytoplasm was used a reference standard protein when whole-cell protein sample was analyzed with Western blot. Western blot experiments were carried out triply (n = 3).

Mentions: Western blotting was conducted to confirm differential expression of PRDX3 in mitochondria of metastatic 5-8F cells compared with nonmetastic 6-10B cells. Equal amount (60 μg) of mitochondrial proteins of metastatic 5-8F and nonmetastatic 6-10B cells were applied to 12% SDS-PAGE and then transferred to PVDF membranes using COX IV as internal standard. The results showed that PRDX3 was significantly up-regulated (about 240%) in mitochondria of metastatic 5-8F cells relative to nonmetastatic 6-10B cells (Figure6). It was consistent with the 2D-DIGE result.


Mitochondrial proteomics of nasopharyngeal carcinoma metastasis.

Liu J, Zhan X, Li M, Li G, Zhang P, Xiao Z, Shao M, Peng F, Hu R, Chen Z - BMC Med Genomics (2012)

Up-regulated expressions of PRDX3 in metastatic 5-8F cell mitochondria relative to nonmetastatic 6-10B cell mitochondria without significant expression difference in the level of whole cells.A. Western blotting image. B. The quantitative comparison of the digitized Western blotting image. COXIV, a marker protein of mitochondria, was used as a reference standard protein when mitochondrial protein sample was analyzed with Western blot. ACTIN that is mainly expressed in the cytoplasm was used a reference standard protein when whole-cell protein sample was analyzed with Western blot. Western blot experiments were carried out triply (n = 3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3539862&req=5

Figure 6: Up-regulated expressions of PRDX3 in metastatic 5-8F cell mitochondria relative to nonmetastatic 6-10B cell mitochondria without significant expression difference in the level of whole cells.A. Western blotting image. B. The quantitative comparison of the digitized Western blotting image. COXIV, a marker protein of mitochondria, was used as a reference standard protein when mitochondrial protein sample was analyzed with Western blot. ACTIN that is mainly expressed in the cytoplasm was used a reference standard protein when whole-cell protein sample was analyzed with Western blot. Western blot experiments were carried out triply (n = 3).
Mentions: Western blotting was conducted to confirm differential expression of PRDX3 in mitochondria of metastatic 5-8F cells compared with nonmetastic 6-10B cells. Equal amount (60 μg) of mitochondrial proteins of metastatic 5-8F and nonmetastatic 6-10B cells were applied to 12% SDS-PAGE and then transferred to PVDF membranes using COX IV as internal standard. The results showed that PRDX3 was significantly up-regulated (about 240%) in mitochondria of metastatic 5-8F cells relative to nonmetastatic 6-10B cells (Figure6). It was consistent with the 2D-DIGE result.

Bottom Line: The functional enrichment analyses of DEPs discovered five significant biological processes including cellular response to reactive oxygen species, hydrogen peroxide metabolic process, regulation of mitochondrial membrane potential, cell redox homeostasis and oxidation reduction, and five significant molecular functions including oxidoreductase activity, caspase inhibitor activity, peroxiredoxin activity, porin activity and antioxidant activity.Those 5-8F cells with suppression of PRDX3 showed an increased mobility potential.These data suggest that those mitochondrial DEPs are potential biomarkers for NPC metastasis, and their dysregulation would play important roles in mitochondria oxidative stress-mediated NPC metastatic process.

View Article: PubMed Central - HTML - PubMed

Affiliation: Key Laboratory of Cancer Proteomics of Chinese Ministry of Health, Xiangya Hospital, Central South University, 87 Xiangya Road, Changsha, Hunan, 410008, PR China.

ABSTRACT

Background: Mitochondrial proteomic alterations of nasopharyngeal carcinoma metastasis remain unknown. Our purpose is to screen mitochondrial proteins for the elucidation of the molecular mechanisms of nasopharyngeal carcinoma metastasis and the discovery of metastasis-related biomarkers.

Methods: Mitochondria were isolated from nasopharyngeal carcinoma metastatic (5-8F) and nonmetastatic (6-10B) cell lines, respectively. After characterization of isolated mitochondria, mitochondrial differentially expressed proteins (DEPs) were quantified by two-dimensional difference in-gel electrophoresis (2D-DIGE), and identified by peptide mass fingerprint (PMF) and tandem mass spectrometry (MS/MS). A functional enrichment analysis and a protein-protein interaction sub-network analysis for DEPs were carried out with bioinformatics. Furthermore, siRNAs transient transfections were used to suppress expressions of some up-regulated DEPs in metastatic cells (5-8F), followed by Transwell Migration assay.

Results: Sixteen mitochondrial DEPs including PRDX3 and SOD2 were identified. Those 5-8F cells with suppression of PRDX3 showed an increased mobility potential. The functional enrichment analyses of DEPs discovered five significant biological processes including cellular response to reactive oxygen species, hydrogen peroxide metabolic process, regulation of mitochondrial membrane potential, cell redox homeostasis and oxidation reduction, and five significant molecular functions including oxidoreductase activity, caspase inhibitor activity, peroxiredoxin activity, porin activity and antioxidant activity. A protein-protein interaction sub-network of DEPs was generated with literature data. Ten mitochondrial DEPs including PRDX3, PRDX6, SOD2, ECH1, SERPINB5, COX5A, PDIA5, EIF5A, IDH3B, and PSMC4 were rationalized in the tumor-stroma co-evolution model that mitochondrial oxidative stress directly contributes to tumor metastasis.

Conclusions: Sixteen mitochondrial DEPs were identified with mass spectrometry and ten of them were rationalized in the tumor-stroma co-evolution model. Those 5-8F cells with suppression of PRDX3 showed an increased mobility potential. These data suggest that those mitochondrial DEPs are potential biomarkers for NPC metastasis, and their dysregulation would play important roles in mitochondria oxidative stress-mediated NPC metastatic process.

Show MeSH
Related in: MedlinePlus