Quantitative membrane proteomics of esophageal squamous cell carcinoma

Abstract

Membranous proteins have been the attractive target in different cancers e.g. EGFR, VEGF, and CXCR4. We used ESCC (esophageal squamous cell carcinoma) as a prototype and employed the stable isotope labeling with amino acids in cell culture (SILAC) methodology to compare membrane proteome of cells with different type of differentiation. Normal epithelial (Het‐1A), well (TE1) & poorly differentiated (TE2) cell lines were labeled with light, medium or heavy arginine plus lysine, respectively. The membrane proteome was isolated by means of Na2CO3 treatment. The samples were resolved on SDS‐PAGE, excised into 24 bands. In‐gel trypsin digestion was carried out and samples were analyzed on LTQ‐Orbitrap Velos mass spectrometer. The data was processed and quantified with Proteome Discoverer (Version 1.3) using 1% FDR and unique peptides were considered for quantitation. We identified 1,950 unique peptides from =20,000 peptide spectrum matches (PSMs) were obtained. This analysis led to identification of 625 differentially expressed proteins between normal and ESCC derived cell lines. There was large number of known and novel membranous proteins e.g. EGFR, BAK1, C2orf18 and BCL2L13.