Characterization of lectin resistant cell populations derived from human colon carcinoma: correlation of K-Ras with beta1-6 branching of N-linked carbohydrate and CEA production.
Previous studies of cell lines derived from human colon carcinoma showed that the extent of beta1-6 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 beta1-6 branching could select for cells having a lower level of K-Ras protein and Ras-activation. PHA-L binds specifically to the beta1-6 branch in N-linked carbohydrate. We utilized a K-ras mutant colon carcinoma cell line, HTB39, which had abundant beta1-6 branching and high levels of K-Ras mutant protein. Lectin resistant cell populations of HTB39 were generated and found to have less beta1-6 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 beta1-6 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 beta1-6 branching of N-linked carbohydrate and CEA production were linked to K-Ras protein synthesis and activation of the Ras-signaling pathway.
Wojciechowicz
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Department of Oncology/Immunology, Wyeth-Ayerst Research, Pearl River, New York, 10965, USA.
Mallon
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D002272:Carcinoembryonic Antigen / Q000096:biosynthesis*; D003110:Colonic Neoplasms / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D037102:Lectins / Q000378:metabolism*; D008715:Methionine / Q000031:analogs & derivatives / Q000494:pharmacology; D010835:Phytohemagglutinins / Q000494:pharmacology; D011233:Precipitin Tests; D014407:Tumor Cells, Cultured; D018631:ras Proteins / Q000378:metabolism*
D002272:Carcinoembryonic Antigen; D004791:Enzyme Inhibitors; C096856:FTI 277; D037102:Lectins; D010835:Phytohemagglutinins; C100156:leukoagglutinins, plants; D008715:Methionine; D018631:ras Proteins