CEA is the major PHA-L-reactive glycoprotein in colon carcinoma cell linesand tumors: Relationship between K-ras activation and beta 1-6 branching of N-linked carbohydrate on CEA

Previously we have shown that a positive correlation existed between the pr esence of beta 1-6 branching of N-linked carbohydrate (detected as PHA-L re activity) and the level of Ras activation in colon carcinoma cell lines. In these cell lines the major PHA-L-reactive species was found to be 180 kDa. Here we identified this species to be carcinoembryonic antigen (CEA) by de monstrating that: (a) CEA immunoreactivity and PHA-L reactivity colocalized on blots of crude cellular membranes from these cell lines, and that (b) i mmunoprecipitation of CEA resulted in quantitative coprecipitation of PHA-L reactivity at 180 kDa. Metabolic labeling of cell line HTB39 with [H-3]man nose revealed that CEA was the predominantly labeled glycoprotein. This ind icated that CEA was the major PHA-L-reactive species due its high level of expression. The amount of PHA-L reactivity present on CEA, expressed as the PHA-L/CEA ratio, was found to vary between cell lines. This ratio was foun d to correlate closely with the level of Ras activation in these cells. In cellular membrane isolated from primary colon carcinoma, the major PHA-L-re active species was also 180 kDa. This reactivity colocalized with CEA immun oreactivity, indicating that the major beta 1-6-branching glycoprotein in m embranes from primary colon carcinoma was CEA. Similar to that seen in cell lines, the amount of PHA-L reactivity on CEA in human tumor samples varied , suggesting that a similar paradigm of Ras-induced expression of beta 1-6 branching may occur in human colon carcinoma, (C) 2000 Academic Press.