CONICET | Buscador de Institutos y Recursos Humanos

Activated leukocyte cell adhesion molecule (ALCAM/CD166) is a member of the immunoglobulin superfamily, primary involved in cell-cell adhesion. Cell surface ALCAM expression has been shown to be downregulated at the tumor cell surface in prostate cancer, and it has been proposed as a cancer biomarker in prostate, colorectal, oral, ovarian and pancreatic cancer. In endothelial cells, ALCAM can interact with galectin-8 (Gal-8), a tandem-repeat type galectin. Surface plasmon resonance studies revealed differential Gal-8 binding affinity towards endogenous ALCAM isolated from MDA-MB-231 breast cancer cells (ALCAMend) and human recombinant ALCAM expressed in HEK-293 cells (ALCAMrec). In this work, we compared the N-glycosylation profile for ALCAMend and ALCAMrec in order to identify structural differences influencing Gal-8 selective interactions. Glycoprofiling was performed by bi-dimensional chromatography combining WAX-HPLC and HILIC-UPLC, and further structural analysis by exoglycosidase sequencing. Our results rule out the presence of sulfated N-glycans as a possible source of differential affinity, and demonstrated that while ALCAMrec presents a high percentage of α(2,3)sialylation in its N-glycans, MDA-MB231-purified ALCAMend has a considerable percentage of α(2,6)sialylated N-glycans, being the latter a non-permissive modification for Gal-8 binding. Further structural analysis revealed interesting structural differences between ALCAM samples from different sources, and highlighted the fact that ALCAM N-glycosylation depends on the cellular expression system and that this differential glycoprofiling affects lectin interactions and biological responses.

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