Characterization of Lectin Resistant Cell Populations Derived from Human Colon Carcinoma: Correlation of K-Ras with β1-6 Branching of N-Linked Carbohydrate and CEA Production

Abstract

Previous studies of cell lines derived from human colon carcinoma showed that the extent of β1-6 branching on N-linked carbohydrate was associated with the presence of K-ras mutation and Ras-activation. We observed that the extent of Ras-activation in these cell lines depends not only upon the presence of an activating mutation in K-ras, but also on the amount of total K-Ras protein produced. Here we examined whether negative selective pressure by PHA-L against β1-6 branching could select for cells having a lower level of K-Ras protein and Ras-activation. PHA-L binds specifically to the β1-6 branch in N-linked carbohydrate. We utilized a K-ras mutant colon carcinoma cell line, HTB39, which had abundant β1-6 branching and high levels of K-Ras mutant protein. Lectin resistant cell populations of HTB39 were generated and found to have less β1-6 branching and less K-Ras protein than their parental counterpart. The lectin resistant cell populations produced lower levels of highly glycosylated CEA, which contributed to the lower level of β1-6 branching in these cells. PHA-L resistant cell populations were two-fold less sensitive than the parental line to an inhibitor of farnesyl transferase (an enzyme essential for Ras processing and function). This suggested a decrease in dependence on K-ras mediated signaling. Collectively, the data indicated that β1-6 branching of N-linked carbohydrate and CEA production were linked to K-Ras protein synthesis and activation of the Ras-signaling pathway.