Exocyst requirement for endocytic traffic directed toward the apical and basolateral poles of polarized MDCK cells

• Published in: Molecular biology of the cell Vol. 18; no. 10; pp. 3978 - 3992
• Main Authors: Oztan, Asli, Silvis, Mark, Weisz, Ora A, Bradbury, Neil A, Hsu, Shu-Chan, Goldenring, James R, Yeaman, Charles, Apodaca, Gerard
• Format: Journal Article
• Language: English
• Published: United States The American Society for Cell Biology 01.10.2007
• Subjects: Animals; Cell Membrane Permeability; Cell Polarity; Dogs; Down-Regulation - genetics; Endocytosis; Endosomes - metabolism; Epithelial Cells - cytology; Epithelial Cells - metabolism; Immunoglobulin A - metabolism; Membrane Proteins - chemistry; Membrane Proteins - metabolism; Protein Binding; Protein Subunits - metabolism; Protein Transport; rab GTP-Binding Proteins - metabolism; Rabbits; Rats; Recombinant Fusion Proteins - metabolism; trans-Golgi Network - metabolism; Transferrin - metabolism; Vesicular Transport Proteins - metabolism
• DOI: 10.1091/mbc.e07-02-0097
• ISSN: 1059-1524

The octameric exocyst complex is associated with the junctional complex and recycling endosomes and is proposed to selectively tether cargo vesicles directed toward the basolateral surface of polarized Madin-Darby canine kidney (MDCK) cells. We observed that the exocyst subunits Sec6, Sec8, and Exo70 were localized to early endosomes, transferrin-positive common recycling endosomes, and Rab11a-positive apical recycling endosomes of polarized MDCK cells. Consistent with its localization to multiple populations of endosomes, addition of function-blocking Sec8 antibodies to streptolysin-O-permeabilized cells revealed exocyst requirements for several endocytic pathways including basolateral recycling, apical recycling, and basolateral-to-apical transcytosis. The latter was selectively dependent on interactions between the small GTPase Rab11a and Sec15A and was inhibited by expression of the C-terminus of Sec15A or down-regulation of Sec15A expression using shRNA. These results indicate that the exocyst complex may be a multipurpose regulator of endocytic traffic directed toward both poles of polarized epithelial cells and that transcytotic traffic is likely to require Rab11a-dependent recruitment and modulation of exocyst function, likely through interactions with Sec15A.