Sialidase NEU3 is a peripheral membrane protein localized on the cell surface and in endosomal structures
Résumé
Sialidase NEU3 is also known as the plasma membrane-associated form of mammalian sialidases, exhibiting a high substrate specificity toward gangliosides. In this respect, sialidase NEU3 modulates cell surface biological events and plays a pivotal role in different cellular processes, including cell adhesion, recognition and differentiation. At the moment, no detailed studies concerning the subcellular localization of NEU3 are available and the mechanism of its association to cellular membranes is still unknown. Here we demonstrate that sialidase NEU3, besides its localization at the plasma membrane, is present in intracellular structures at least partially represented by a subset of the endosomal compartment. Moreover, we show that NEU3 present at the plasma membrane is internalized and locates then to the recycling endosomal compartment. The enzyme is associated to the outer leaflet of the plasma membrane, as shown by selective cell surface protein biotinylation. This evidence is in agreement with the ability of NEU3 in degrading gangliosides inserted in the plasma membrane of adjacent cells. Moreover, the association mechanism of the protein to the lipid bilayer was elucidated by carbonate extraction. Under these experimental conditions we succeeded in solubilizing NEU3, thus demonstrating that the enzyme is a peripheral membrane protein. In addition, Triton X-114 phase separation further demonstrates the hydrophilic nature of the protein. Overall, these results provide important information about the biology of NEU3, the most studied member of the mammalian sialidase family.
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