Proteomic studies of post-translational O-GlcNAcylation in colon cancerThanong Phueaouan
( M.Sc. )
M.R. Jisnuson Svasti
O-GlcNAcylation is a novel post-translational modification which plays important roles in diseases including cancer. Although several proteins were reported to be modified by O-GlcNAc, this modification in cancer biology is still largely unexplored. Therefore, the aim of this research was to study the expression of O-GlcNAc protein levels and identify these proteins in human colon cancer. Fifteen tissue specimens of colon cancer patients including 5 samples of colon cancer grade I, 8 samples of grade II and 2 samples of grade III, were used in this study. O-GlcNAc proteins between cancer and normal adjacent specimens were compared by using one- and two- dimensional polyacrylamide gel electrophoresis (1D- and 2D-PAGE) followed by immunoblotting with anti-O-GlcNAc antibody (CTD110.6). CTD110.6 immonublotting revealed many O-GlcNAc proteins in both cancer and normal tissues. The densitometric analysis of the expression of total O-GlcNAc proteins showed no different expression levels between cancer grade I and normal, however, an increase of this modification was observed in cancer grade II and grade III when compared to normal. Using 2D-PAGE followed with CTD110.6 immunoblotting, O-GlcNAc protein patterns of individual and pooled cancer grade I, II and III, and normal samples were mapped and compared. From the pooled cancer and normal samples, 48 protein spots were selected to determine their O-GlcNAc expression levels and only 22 proteins of 18 spots were successfully identified. Thirteen proteins were highly O-GlcNAc expressed in all cancer grades compared to normal. Whereas nine proteins were O-GlcNAc over expressed in cancer grade I and II, except grade III, when compared to normal. Among of these identified proteins, cytokeratin 18 and annexin A2 were further confirmed to be O-GlcNAc overexpressed in colon cancer by using O-GlcNAc immunoprecipitation. These data demonstrate that O-GlcNAc modification is essential for cancer and altered O-GlcNAc proteins identified are likely to play important roles in colon cancer development. The identified O-GlcNAc proteins may serve as novel markers for colon cancer and for the study of cellular signaling pathway in cancer.