Biosynthesis and glycosylation of the carcinoembryonic antigen

Abstract

Human gastric adenocarcinoma MKN-45 cells were found to synthesize actively carcinoembryonic antigen (CEA). The biosynthesis and carbohydrate processing of CEA were studied in these cells by means of metabolic labelling followed by immunoadsorption with a specific polyclonal-antibody preparation and gel electrophoresis. Pulse-chase studies with [14C]leucine and [3H]mannose (shortest pulse 3 min) showed that N-linked oligosaccharide side chains are added to the protein co-translationally, producing a high-mannose immature CEA; the average molecular mass of this form is 145 kDa. The protein is later translocated to the Golgi apparatus and here undergoes additional processing; these modifications are visible in our system as a broadening of the CEA band and require about 4 h. The upper limit of mature CEA band reaches 200 kDa, but radioactivity is maximally incorporated at 168 kDa. The extent of co-translational glycosylation was measured by treating the cells with tunicamycin; in the presence of this inhibitor, a 74 kDa aglyco-CEA was produced and was still recognized by the antibody. Monensin, an ionophore which interferes with glycoprotein maturation and terminal sugar addition, blocked broadening of the CEA band, producing a sharp 141 kDa peak. In conclusion, CEA appears to be synthesized as a 145 kDa high-mannose immature form, the protein core accounting for about half of its molecular mass. Full maturation results in a broad band at 168 kDa.

The influence of metastatic site on the expression of CEA and cellular localization of β‐catenin in colorectal cancer

“…Importantly, CEA was shown previously to undergo partial glycosylation resulting in the appearance of similar mol. mass species of CEA observed in our studies. Additionally, as described by the American Tissue and Cell Culture Collection, the LS180 and HT29 cells are known to express high levels of CEA when compared with HCT116 or ATRFLOX cells.…”

Intracellular Targeting of CEA Results in Th1-Type Antibody Responses Following Intradermal Genetic Vaccination by a Needle-Free Jet Injection Device

“…1). Cells transfected with the wild-type construct also gave rise to a band of around 150 kDa, matching that of an early, not yet fully glycosylated, form of CEA [19]. In contrast, detection of CEA in samples from cells expressing the truncated CEA proteins CEA6 and CEA66 (lacking signal peptides) revealed a band of about 80 kDa, corresponding to the reported size of nonglycosylated CEA (78 kDa) [20].…”

“…The sequence of wild-type CEA contains both N-and C-terminal signal peptides. The N-terminal signal peptide ensures proper cotranslational translocation of the wild-type CEA polypeptide into the ER, where it is heavily glycosylated, and its subsequent transport to the plasma membrane [19]. During posttranslational processing of the protein, the C-terminal signal sequence is removed and replaced with a glycosylphosphatidylinositol (GPI) membrane anchor [21,28].…”

Site-Specific N-Linked Glycosylation Analysis of Human Carcinoembryonic Antigen by Sheathless Capillary Electrophoresis–Tandem Mass Spectrometry

“…18,19 Human carcinoembryonic antigen (CEA) (UniProt entry P06731, CEACAM5_HUMAN) is a highly N-glycosylated protein of approximately 180 kDa with sugars accounting for about half of its molecular mass. 20 CEA can be found in normal human colonic epithelial cells and is reported to be upregulated in tumor forming and colonic adenocarcinogenic cell lines; 21,22 elevated CEA levels are valued as a progression and outcome biomarker in several forms of human cancers, 23 including colorectal cancer (CRC). 24 Continuous measurement of CEA serum levels is clinically used for the postoperative and posttherapy recurrence monitoring of CRC patients.…”