Quantitative Proteomic Analysis Identifies MAPK15 as a Potential Regulator of Radioresistance in Nasopharyngeal Carcinoma Cells.

Zhanzhan Li,Na Li,Liangfang Shen,Jun Fu

doi:10.3389/fonc.2018.00548

Abstract

Since resistance to radiotherapy remains refractory for the clinical management of nasopharyngeal cancer (NPC), further understanding the mechanisms of radioresistance is necessary in order to develop more effective NPC treatment and improve prognosis. In this study, an integrated quantitative proteomic approach involving tandem mass tag labeling and liquid chromatograph-mass spectrometer was used to identify proteins potentially responsible for the radioresistance of NPC. The differential radiosensitivity in NPC model cells was examined through clonogenic survival assay, CCK-8 viability assay, and BrdU incorporation analysis. Apoptosis of NPC cells after exposure to irradiation was detected using caspase-3 colorimetric assay. Intracellular reactive oxygen species (ROS) was detected by a dichlorofluorescin diacetate fluorescent probe. In total, 5,946 protein groups were identified, among which 5,185 proteins were quantified. KEGG pathway analysis and protein-protein interaction enrichment analysis revealed robust activation of multiple biological processes/pathways in radioresistant CNE2-IR cells. Knockdown of MAPK15, one up-regulated protein kinase in CNE2-IR cells, significantly impaired clonogenic survival, decreased cell viability and increased cell apoptosis following exposure to irradiation, while over-expression of MAPK15 promoted cell survival, induced radioresistance and reduced apoptosis in NPC cell lines CNE1, CNE2, and HONE1. MAPK15 might regulate radioresistance through attenuating ROS accumulation and promoting DNA damage repair after exposure to irradiation in NPC cells. Quantitative proteomic analysis revealed enormous metabolic processes/signaling networks were potentially involved in the radioresistance of NPC cells. MAPK15 might be a novel potential regulator of radioresistance in NPC cells, and targeting MAPK15 might be useful in sensitizing NPC cells to radiotherapy.

Highlights

Nasopharyngeal carcinoma (NPC) is one of the most prevalent cancers in South China and Southeast Asia
CNE2-IR showed much higher clonogenic potential than CNE2 after exposure to irradiation (Figure 1A). These two cell lines were used for quantitative proteomic analysis using tandem mass tag (TMT) labeling and LC-MS/MS
Several radioresistance-related proteins [e.g., superoxide dismutase 2 (SOD2), heat shock protein 1 (HSPB1), peroxiredoxin 1 (PRDX1), nucleoside diphosphate kinase 1 (NME1), nucleophosmin 1 (NPM1)] identified in previous 2-DE/MALDI-TOF-MS studies in a CNE2-IR/CNE2 cell model were observed in our dataset (Table S2)

Summary

Introduction

Nasopharyngeal carcinoma (NPC) is one of the most prevalent cancers in South China and Southeast Asia. Radioresistance in Nasopharyngeal Carcinoma local control, the local recurrence and distant metastasis of NPC are still the major causes of deaths and morbidity in advanced stages [2]. Since a common cause of local recurrence and poor survival in NPC is radioresistance [3], understanding the mechanisms of radioresistance is necessary in order to develop more specific NPC treatment for and improve prognosis. As key regulators of cell functions, are one of the largest and most functionally diverse gene families. An integrated quantitative proteomic approach involving tandem mass tag (TMT) labeling and liquid chromatograph-tandemmass spectrometer (LC-MS/MS) was used to identify potential proteins responsible for the radioresistance of NPC [7, 8]. Our data suggest MAPK15 might be an important regulator of radioresistance in NPC cells, which may warrant MAPK15 as a potential therapeutic target in future investigations

Methods

Results

Conclusion