Characterization of lectin resistant cell populations derived from human colon carcinoma: Correlation of K-Ras with beta 1-6 branching of N-linked carbohydrate and CEA production

Previous studies of cell lines derived from human colon carcinoma showed th at the extent of beta 1-6 branching on N-linked carbohydrate was associated with the presence of K-ras mutation and Ras-activation. We observed that t he extent of Ras-activation in these cell lines depends not only upon the p resence of an activating mutation in K-ras, but also on the amount of total K-Ras protein produced. Here we examined whether negative selective pressu re by PHA-L against beta 1-6 branching could select for cells having a lowe r level of K-Ras protein and Ras-activation. PHA-L binds specifically to th e beta 1-6 branch in N-linked carbohydrate. We utilized a R-ras mutant colo n carcinoma cell line, HTB39, which had abundant beta 1-6 branching and hig h levels of K-Ras mutant protein. Lectin resistant cell populations of HTB3 9 were generated and found to have less beta 1-6 branching and less K-Ras p rotein than their parental counterpart. The lectin resistant cell populatio ns produced lower levels of highly glycosylated CEA, which contributed to t he lower level of beta 1-6 branching in these cells. PHA-L resistant cell p opulations were two-fold less sensitive than the parental line to an inhibi tor of farnesyl transferase (an enzyme essential for Pas processing and fun ction). This suggested a decrease in dependence on K-ras mediated signaling . Collectively, the data indicated that beta 1-6 branching of N-linked carb ohydrate and CEA production were linked to K-Ras protein synthesis and acti vation of the Ras-signaling pathway. (C) 1999 Academic Press.