The architectural complexity of the hepatocyte canalicular surface has prevented examination of apical membrane dynamics with methods used for other epithelial cells. By adopting a pharmacological approach, we have documented for the first time the internalization of membrane proteins from the hepatic apical surface. Treatment of hepatocytes or WIF-B cells with phosphoinositide 3-kinase inhibitors, wortmannin or LY294002, led to accumulation of the apical plasma membrane proteins, 5′-nucleotidase and aminopeptidase N in lysosomal vacuoles. By monitoring the trafficking of antibody-labeled molecules, we determined that the apical proteins in vacuoles came from the apical plasma membrane. Neither newly synthesized nor transcytosing apical proteins accumulated in vacuoles. In wortmannin-treated cells, transcytosing apical proteins traversed the subapical compartment (SAC), suggesting that this intermediate in the basolateral-to-apical transcytotic pathway remained functional. Ultrastructural analysis confirmed these results. However, apically internalized proteins did not travel through SAC en route to lysosomal vacuoles, indicating that SAC is not an intermediate in the apical endocytic pathway. Basolateral membrane protein distributions did not change in treated cells, uncovering another difference in endocytosis from the two domains. Similar effects were observed in polarized MDCK cells, suggesting conserved patterns of phosphoinositide 3-kinase regulation among epithelial cells. These results confirm a long-held but unproven assumption that lysosomes are the final destination of apical membrane proteins in hepatocytes. Significantly, they also confirm our hypothesis that SAC is not an apical endosome.
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31 May 1999
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May 31 1999
Evidence for Apical Endocytosis in Polarized Hepatic Cells: Phosphoinositide 3-Kinase Inhibitors Lead to the Lysosomal Accumulation of Resident Apical Plasma Membrane Proteins
Pamela L. Tuma,
Pamela L. Tuma
Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
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Catherine M. Finnegan,
Catherine M. Finnegan
Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
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Ji-Hyun Yi,
Ji-Hyun Yi
Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
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Ann L. Hubbard
Ann L. Hubbard
Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
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Pamela L. Tuma
Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
Catherine M. Finnegan
Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
Ji-Hyun Yi
Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
Ann L. Hubbard
Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205
Address correspondence to Ann L. Hubbard, Department of Cell Biology and Anatomy, Johns Hopkins University School of Medicine, Baltimore, MD 21205. Tel.: (410) 955-2333. Fax: (410) 955-1013. E-mail: [email protected]
Received:
March 18 1999
Revision Received:
April 15 1999
Online ISSN: 1540-8140
Print ISSN: 0021-9525
1999
J Cell Biol (1999) 145 (5): 1089–1102.
Article history
Received:
March 18 1999
Revision Received:
April 15 1999
Citation
Pamela L. Tuma, Catherine M. Finnegan, Ji-Hyun Yi, Ann L. Hubbard; Evidence for Apical Endocytosis in Polarized Hepatic Cells: Phosphoinositide 3-Kinase Inhibitors Lead to the Lysosomal Accumulation of Resident Apical Plasma Membrane Proteins . J Cell Biol 31 May 1999; 145 (5): 1089–1102. doi: https://doi.org/10.1083/jcb.145.5.1089
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