Distribution of membrane­bound sialidase NEU3: the effect of lipid raft patching

The plasma membrane­associated sialidase NEU3 removes sialic acid from oligosaccharides, glycosphingolipids and glycoproteins directly on the cell surface and shows high substrate specificity toward gangliosides that are abundant in lipid rafts. Lipid rafts are small, dynamic domains enriched in cholesterol, sphingomyelin, gangliosides and proteins involved in biological processes with pivotal physio pathological implications. In this study the distribution of sialidase NEU3 inside and outside lipid rafts, its influence on the ganglioside composition, and its behaviour upon lipid raft patching has been described in COS7 and HeLa cells transiently transfected with the murine NEU3 cDNA. Upon NEU3 expression, the enzyme reaches a maximum activity that is 10/13­fold higher compared to non­transfected cells and equally distributes between the lipid raft and the non­raft membranes. The enzyme modifies the ganglioside content of the lipid bilayer with a decrease in GD1a and an increase in GM1. In NEU3 transfected cells, lipid raft patching induced by cholera toxin subunit­B treatment results in the recruitment of the enzyme inside lipid rafts. Confocal microscopy analysis of HeLa cells confirms NEU3 differential distribution between non­raft membranes and lipid rafts of the enzyme and its recruitment in lipid rafts upon raft patching. Aggregation of lipid rafts has been demonstrated in several biological events such as the regulation of haematopoiesis, the response to death receptor agonists in endothelial cells, the vesicular transport in polarized cells and, finally, in the series of events leading to malignancy. In this perspective, the recruitment of NEU3 from non­raft membranes into clustered lipid rafts may be relevant to explain the functional implications of the enzyme in raft biology. Moreover, NEU3 could be a possible novel target to modulate the behaviour of lipid rafts in cell biology.