Association of stomatin with lipid-protein complexes in the plasma membrane and the endocytic compartment

The 31 kDa integral membrane protein stomatin (protein 7.2b) has a monotopic structure and a cytofacial orientation, We have shown previously that stomatin is located in plasma membrane protruding structures and forms high-order homooligomers in the human epithelial cell line UAC, suggesting that this protein has a structural function in the cortical morphogenesis of the cells, It is also present in a pool of juxtanuclear vesicles. In this study, we show that stomatin colocalizes with the GPI-anchored proteins placental alkaline phosphatase (PLAP) and membrane folate receptor alpha (MFR alpha) endogenously expressed in UAC cells, This observation enabled us to demonstrate two different aspects of stomatin, First, using anti-FLAP antibody internalization, we show that the peri-centrosomal vesicles containing stomatin correspond to a subset of endosomes, which can also be labeled with the late endosomal/lysosomal marker LAMP-2, Secondly, we found that stomatin is partially present in detergentin-soluble membrane domains and co-patches with FLAP on the plasma membrane, after cross-linking of FLAP by antibodies. These data indicate that stomatin and GPI-anchored proteins are linked through lipid rafts and undergo the same sorting events. We propose that stomatin, through its affinity for lipid rafts, functions in concentrating GPI-anchored proteins in membrane microvillar structures. Consistent with this hypothesis, we found that stomatin is expressed exclusively in microvilli of the apical membrane in polarized Madin-Darby canine kidney (MDCK) cells.