Differential involvement of endocytic compartments in the biosynthetic traffic of apical proteins

• Published in: The EMBO journal Vol. 26; no. 16; pp. 3737 - 3748
• Main Authors: Cresawn, Kerry O, Potter, Beth A, Oztan, Asli, Guerriero, Christopher J, Ihrke, Gudrun, Goldenring, James R, Apodaca, Gerard, Weisz, Ora A
• Format: Journal Article
• Language: English
• Published: Chichester, UK John Wiley & Sons, Ltd 22-08-2007; Blackwell Publishing Ltd; Nature Publishing Group
• Subjects: Animals; Biomarkers - metabolism; biosynthetic delivery; Cell Line; Cell Membrane - chemistry; Cell Membrane - metabolism; Cell Polarity; Cellular biology; Dogs; Endocytosis - physiology; Endolyn - genetics; Endolyn - metabolism; endosome; Endosomes - metabolism; Glycosylphosphatidylinositols - metabolism; Hemagglutinin Glycoproteins, Influenza Virus - genetics; Hemagglutinin Glycoproteins, Influenza Virus - metabolism; Humans; Inactivation; Kidneys; lipid raft; MDCK; Membrane Glycoproteins - genetics; Membrane Glycoproteins - metabolism; Membrane Microdomains - metabolism; Membrane Proteins - genetics; Membrane Proteins - metabolism; Molecular biology; polarized; Protein Transport - physiology; Proteins; rab GTP-Binding Proteins - metabolism; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Transferrin - genetics; Transferrin - metabolism; Vesicular stomatitis virus; Viral Envelope Proteins - genetics; Viral Envelope Proteins - metabolism
• ISSN: 0261-4189; 1460-2075
• DOI: 10.1038/sj.emboj.7601813

Newly synthesized basolateral markers can traverse recycling endosomes en route to the surface of Madin–Darby canine kidney cells; however, the routes used by apical proteins are less clear. Here, we functionally inactivated subsets of endocytic compartments and examined the effect on surface delivery of the basolateral marker vesicular stomatitis virus glycoprotein (VSV‐G), the raft‐associated apical marker influenza hemagglutinin (HA), and the non‐raft‐associated protein endolyn. Inactivation of transferrin‐positive endosomes after internalization of horseradish peroxidase (HRP)‐containing conjugates inhibited VSV‐G delivery, but did not disrupt apical delivery. In contrast, inhibition of protein export from apical recycling endosomes upon expression of dominant‐negative constructs of myosin Vb or Sec15 selectively perturbed apical delivery of endolyn. Ablation of apical endocytic components accessible to HRP‐conjugated wheat germ agglutinin (WGA) disrupted delivery of HA but not endolyn. However, delivery of glycosylphosphatidylinositol‐anchored endolyn was inhibited by >50% under these conditions, suggesting that the biosynthetic itinerary of a protein is dependent on its targeting mechanism. Our studies demonstrate that apical and basolateral proteins traverse distinct endocytic intermediates en route to the cell surface, and that multiple routes exist for delivery of newly synthesized apical proteins.