Identification of O-GlcNAc-Modified Proteins in Human Lung Cancer Cells

• Main Authors: Jia-Yi Peng, 彭佳儀
• Other Authors: Teh-Ying Chou
• Format: Others
• Language: zh-TW
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/75446738897212900521

碩士 === 國立陽明大學 === 生化暨分子生物研究所 === 100 === O-linked β-N-acetylglucosamine modification (O-GlcNAcylation) is a dynamic, reversible and inducible post-translational modification on nuclear and cytoplasmic proteins. O-GlcNAc has been reported on a large number of proteins which are involved in a wide range of biological processes, including metabolism, signal transduction, transcription, translation, and protein turnover. Studies have shown that O-GlcNAcylation plays a role in many human diseases, such as Alzheimer disease, diabetes and cardiovascular disease. Recently, the cellular O-GlcNAcylation level has been correlated with cancer progression and metastasis; however, little is known about the O-GlcNAc-modified proteins which are involved. In this study, we compared two human lung adenocarcinoma cell lines with different invasion ability, i.e., CL1-1 (less invasive) and CL1-5 (more invasive), to identify O-GlcNAc modified proteins which play an important role in tumor metastasis. Comparing on O-GlcNAc Western blotting, we found that CL1-1 and CL1-5 differentially express O-GlcNAc modified nuclear proteins. We used wheat germ agglutinin to purify O-GlcNAc modified nuclear proteins and identified these proteins by mass spectrometry to analyze different O-GlcNAc modified proteins between CL1-1 and CL1-5. We found two proteins that are involved in cancer progression and metastasis, i.e., IQGAP1, which likely expressed higher O-GlcNAcylation in CL1-1 and Sam68, which likely expressed higher O-GlcNAcylation in CL1-5. Furthermore, we confirmed that IQGAP1 and Sam68 were O-GlcNAcylated in CL1-1 and CL1-5 cell lines. We also identified four O-GlcNAc modification sites, Ser15, Ser18, Ser20 and Ser422, on protein Sam68 by mass spectrometry. We will further investigate the roles of Sam68 O-GlcNAcylation in cancer pathogenesis and metastasis.