Dynamics of MAL2 during glycosylphosphatidylinositol-anchored protein transcytotic transport to the apical surface of hepatoma HepG2 cells.
Delivery of glycosylphosphatidylinositol (GPI)-anchored proteins to the apical surface takes place by transcytosis in hepatocytes and also probably in epithelial Madin-Darby canine cells. The integral protein MAL2 was demonstrated to be essential for basolateral-to-apical transcytosis in hepatoma HepG2 cells. Reduction of endogenous MAL2 levels impedes cargo delivery to the apical membrane, but, paradoxically, cargo does not accumulate in the subapical compartment where MAL2 predominantly resides but in distant endosome elements. To understand how transcytosis can be apparently mediated at a distance, we have analyzed the dynamics of machinery and cargo by live-cell imaging of MAL2 and transcytosing CD59, a GPI-anchored protein, in HepG2 cells. MAL2 was revealed as being a highly dynamic protein. Soon after basolateral endocytosis of CD59, a fraction of MAL2 redistributed into peripheral vesicular clusters that concentrated CD59 and that were accessible to transferrin (Tf) receptor, a basolateral recycling protein. Following Tf receptor segregation, the clusters fused in a MAL2(+)globular structure and moved toward the apical surface for CD59 delivery. All these processes were impaired in cells with reduced MAL2 content. Other GPI-anchored proteins examined behave similarly. As MAL2 is expressed by many types of epithelia, the sorting events described herein are probably of quite general utility.
de Marco
María C
MC
Centro de Biología Molecular Severo Ochoa, Universidad Autónoma de Madrid and Consejo Superior de Investigaciones Científicas, Cantoblanco, Madrid, Spain.
Puertollano
Rosa
R
Martínez-Menárguez
José A
JA
Alonso
Miguel A
MA
D018957:CD59 Antigens / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000378:metabolism*; D045744:Cell Line, Tumor; D016764:Cell Polarity; D003600:Cytosol / Q000378:metabolism*; D004847:Epithelial Cells / Q000737:chemistry / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D062806:Myelin and Lymphocyte-Associated Proteolipid Proteins; D021381:Protein Transport / Q000502:physiology; D011510:Proteolipids / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism*
D018957:CD59 Antigens; D017261:Glycosylphosphatidylinositols; C438606:MAL2 protein, human; D062806:Myelin and Lymphocyte-Associated Proteolipid Proteins; D011510:Proteolipids; D033921:Vesicular Transport Proteins; D049452:Green Fluorescent Proteins