Differential involvement of endocytic compartments in the biosynthetic traffic of apical proteins.
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.
Renal-Electrolyte Division, University of Pittsburgh, Pittsburgh, PA 15261, USA.
Kerry O
KO
Cresawn
Beth A
BA
Potter
Asli
A
Oztan
Christopher J
CJ
Guerriero
Gudrun
G
Ihrke
James R
JR
Goldenring
Gerard
G
Apodaca
Ora A
OA
Weisz
D000818:Animals; D015415:Biomarkers / Q000378:metabolism*; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D016764:Cell Polarity; D004285:Dogs; D004705:Endocytosis / Q000502:physiology*; D051930:Endolyn / Q000235:genetics / Q000378:metabolism; D011992:Endosomes / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols / Q000378:metabolism; D019267:Hemagglutinin Glycoproteins, Influenza Virus / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000378:metabolism; D021962:Membrane Microdomains / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D021381:Protein Transport / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014168:Transferrin / Q000235:genetics / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000235:genetics / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000378:metabolism; D000099193:rab11 GTP-Binding Proteins
D015415:Biomarkers; D051930:Endolyn; D017261:Glycosylphosphatidylinositols; D019267:Hemagglutinin Glycoproteins, Influenza Virus; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D014168:Transferrin; D014759:Viral Envelope Proteins; D020691:rab GTP-Binding Proteins; D000099193:rab11 GTP-Binding Proteins; C028391:G protein, vesicular stomatitis virus