Endocytosis

1. The Pathway of Endocytosis.- 1. Introduction.- 2. Summary of the Pathway.- 3. History of Endocytosis.- 4. Ligands Internalized by Receptor-Mediated Endocytosis.- 5. Receptor Distribution.- 5.1. Receptor Mobility.- 5.2. Clustering of Ligand—Receptor Complexes in Coated Pits.- 5.3. Preclustered Receptors.- 6. Receptosomes.- 6.1. Mechanism of Receptosome Formation.- 6.2. Properties of Receptosomes.- 6.3. Rapid Speed of the Endocytic Event.- 6.4. Fusion of Receptosomes.- 7. Role of the Golgi System.- 7.1. Ligand Entry into the Golgi System.- 7.2. Sorting in the TR Golgi.- 7.3 Structure of the Golgi System.- 8. Down-Regulation of Receptors.- 9. Why Ligands Enter Cells at Different Rates.- 10. Functions of Receptor-Mediated Endocytosis.- 11. Conclusions and Future Prospects.- References.- 2. Receptors.- 1. Scope of Receptorology.- 2. Receptor Organization.- 3. The Study of Receptors.- 4. Techniques of Ligand Binding to Receptors.- 4.1. Measurement of Binding.- 4.2. Assay of Membrane Receptors.- 4.2.1. Filtration.- 4.2.2. Centrifugation.- 4.2.3. Equilibrium Dialysis and Flow Dialysis.- 4.2.4. Assay of Solubilized Receptors.- 4.2.5. “Nonspecific” Binding.- 5. Analysis of Binding Data.- 5.1. The Simple Noncooperative (Michaelian) Binding Pattern.- 5.1.1. The Direct Plot.- 5.1.2. The Semilogarithmic Plot.- 5.1.3. The Scatchard Plot.- 5.1.4. The Double-Reciprocal Plot.- 5.1.5. The Hill Plot.- 5.2. Displacement Experiments.- 5.3. Non-Michaelian Ligand Binding.- 5.4. Distinguishing Negative Cooperativity from Heterogeneous Population of Sites.- 5.4.1. Equilibrium Methods.- 5.4.2. The Kinetic Approach.- 6. Receptor-to-Effector Coupling.- 6.1. The Nicotinic System.- 6.2. Hormone-Dependent Analysis Cyclase.- 7. Receptor Desensitization and Down-Regulation.- 7.1. The Nicotinic Receptor System.- 7.2. The ?-Adrenergic System.- References.- 3. Chemical and Physical Properties of the Hepatic Receptor for Asialoglycoproteins.- 1. Introduction.- 2. Physical Properties.- 3. Requirement for Calcium.- 4. Determinants of Binding.- 5. Binding Kinetics.- 6. Dual Role of Sialic Acid.- 7. Receptor Distribution and Topology.- 8. Avian Hepatic Binding Protein.- 9. Perspectives.- References.- 4. The Structure of Clathrin-Coated Membranes: Assembly and Disassembly.- 1. Introduction.- 1.1. Coated Membranes in Cells.- 1.2. Isolated Coated Vesicles.- 2. Isolation, Extraction, and Fractionation of Coated Vesicles and Their Components.- 2.1. Purification of Coated Vesicles.- 2.2. Release of Coats from Vesicles.- 2.3. Fractionation of Coated Vesicle Extracts.- 3. Composition of Coated Vesicles.- 3.1. Clathrin Triskelions.- 3.1.1. Clathrin Triskelions: The Structural and Functional Unit.- 3.1.2. Clathrin Triskelions: Composition.- 3.1.3. Clathrin Triskelions: The Heavy Chain.- 3.1.4. Clathrin Triskelions: Light Chains.- 3.1.5. Clathrin Triskelions: Heavy-Chain—Light-Chain Interactions.- 3.2. Assembly Polypeptides.- 3.3. Tubulin and ?-Related Polypeptides.- 3.4. Calmodulin.- 3.5. Lipid and Carbohydrate.- 4. Clathrin Coat Dynamics.- 4.1. Assays for Coat Assembly.- 4.1.1. Electron Microscopy.- 4.1.2. Sedimentation Assays.- 4.1.3. Light Scattering.- 4.2. Coat Assembly: Triskelions.- 4.2.1. Coat Assembly: Clathrin Domains Required.- 4.3. Coat Reassembly: Role of Assembly Polypeptides.- 4.4. Coat Assembly: Clathrin Binding to Membranes.- 4.5. Coat Disassembly.- 5. Conclusions.- References.- 5. Transferrin: Receptor-Mediated Endocytosis and Iron Delivery.- 1. Introduction.- 2. Structure of Transferrin.- 2.1. Chemical Characterization.- 2.2. Carbohydrate Chains.- 2.3. Iron Binding.- 2.4. Iron Release.- 3. Function of Transferrin.- 3.1. Ubiquity of Transferrin Receptors.- 3.2. Transferrin—Reticulocyte Interactions.- 4. Role of Transferrin in Biology and Medicine.- 4.1. Requirements for Cell Growth and Proliferation.- 4.2. Relationship to Malignant Transformation.- 4 3 Immunological Surveillance of Cancer and the Transferrin Receptor.- 4.4. Use of the Transferrin Receptor in Chemotherapy.- 5. The Transferrin Receptor: Biochemical Characterization.- 5.1. Transferrin Receptor Structure.- 5.2. Transferrin Receptor Biosynthesis.- 6. Cellular Binding and Uptake of Transferrin: Kinetic and Inhibitor Studies.- 7. Prelysosomal Divergence of EGF and Transferrin During Endocytosis.- 7.1. Characterization of Binding Sites for EGF and Transferrin.- 7.2. Release and Degradation of EGF and Transferrin from Cells at 37°C.- 7.3. Density Gradient Centrifugation of Cell Fractions on Colloidal Silica.- 7.4. Fluorescence Microscopy.- 7.5. Electron Microscopy.- 8. Role of a Prelysosomal Compartment in Transferrin-Bound Iron Release and Receptor-Bound Ligand Release.- 9. Biosynthesis and Recycling of Receptors: Two Roles for Secretion in Endocytosis?.- 10. Summary and Future Prospects.- References.- 6. POLYMERIC IgA AND GALACTOSE-SPECIFIC PATHWAYS IN RAT HEPATOCYTES: EVIDENCE FOR INTRACELLULAR LIGAND SORTING.- 1. Introduction.- 2. Receptor-Mediated Endocytosis in Rat Hepatocytes.- 2.1. Diversity of Recognition Systems.- 2.2. Diversity of the Fates of Ligands and Receptors.- 3. Methodology.- 3.1. Tagging of Ligands and Double-Labeling Experiments.- 3.2. Assessment of Polymeric IgA Derivatives.- 3.3. Assessment of Galactose-Exposing Derivatives.- 3.4. Independence of Ligand Processing.- 4. Fate of Secretory Component and Galactose-Specific Receptors.- 4.1. Biosynthesis and Properties of the Secretory Component.- 4.2. Endocytosis of Secretory Component and Postendocytotic Events.- 4.3. Properties of the Galactose-Specific Receptors.- 4.4. Endocytosis of Galactose-Specific Receptors and Postendocytotic Events.- 5. Pathways of Polymeric IgA and of Galactose-Exposing Derivatives in Rat Hepatocytes: Ultrastructural Studies.- 5.1. The Polymeric IgA-Specific Pathway.- 5.2. The Galactose-Specific Pathway.- 5.3. Cointernalization of Polymeric IgA and Galactose-Exposing Derivatives.- 6. Intracellular Ligand Sorting in Rat Hepatocytes.- 6.1. The DAB-Induced Density Shift.- 6.2. Concomitant Density Shift of Ligands.- 6.3. Combined Differential and Isopycnic Centrifugation Studies.- 7. Mechanism of Ligand and Receptor Sorting.- 7.1. Acidification Mediates a Two-Phase Partition.- 7.2. Phase Sorting.- 7.3. Receptor Sorting and Specific Addressing.- 7.4. Current Model and Implications.- 8. Properties of Ligand-Sorting Organelles.- 8.1. Physical and Morphological Properties.- 8.2. Membrane Composition.- 8.3. Absence of Proteolysis.- 8.4. Cholesterol-Rich Membrane.- 9. Conclusions and Perspectives.- 9.1. Identification of Sorting Organelles.- 9.2. Perspectives on the Sorting Mechanism.- 9.3. Ligand-Containing Structures as Transient or Stable Organelles.- References.- 7. Toxins.- 1. Introduction.- 2. Toxin Structure.- 2.1. The Plant Lectins Ricin, Abrin, Modeccin, and Viscumin.- 2.2. Diphtheria Toxin and Pseudomonas aeruginosa Exotoxin A.- 2.3. Cholera Toxin, E. coli Heat-Labile Toxin, Pertussis Toxin, and Shigella Toxin.- 2.4. Toxin Conjugates.- 3. Intracellular Action.- 3.1. Diphtheria Toxin and Pseudomonas Toxin.- 3.2. Ricin, Abrin, Modeccin, Viscumin and Shigella Toxin.- 3.3. Cholera Toxin, E. coli Heat-Labile Toxin, and Pertussis Toxin.- 3.4. Anthrax Toxin.- 4. Function of the Cell Surface Binding Sites.- 4.1. Characterization of the Binding Sites.- 4.1.1. Binding Sites for Ricin, Abrin, Modeccin, and Viscumin.- 4.1.2. Diphtheria Toxin Receptor.- 4.1.3. Receptor for Cholera Toxin and E. coli Toxin.- 4.1.4. Binding Sites for Other Toxins.- 4.2. Characteristics of the Binding.- 4.3. Ability of Binding Sites to Facilitate Toxin Entry.- 5. Endocytosis and Transport of Toxin-Containing Vesicles.- 5.1. Morphological Studies.- 5.2. Importance of Endocytosis.- 5.3. Intracellular Transport of Toxin-Containing Vesicles.- 5.4. Properties of Vesicular Compartments Relevant to Toxin Entry.- 6. Requirements for Toxin Exit from Intracellular Vesicles.- 6.1. Role of Low pH.- 6.1.1. Penetration of Diphtheria Toxin at Low pH.- 6.1.2. Requirement for Low pH for Entry of Other Toxins.- 6.1.3. Other pH Effects on Toxin Entry.- 6.2. Ion Requirements.- 6.2.1. Role of Calcium.- 6.2.2. Role of Chloride.- 6.3. Energy Requirements.- 6.4. Role of the Disulfide Bond.- 6.5. Studies of Toxin Entry Using Photoreactive Compounds.- 7. Conclusions.- References.- 8. Acidification of Endocytic Vesicles and Lysosomes.- 1. Introduction.- 1.1. Historical Background.- 1.2. Acidification in Various Cell Types.- 2. Measurement of pH.- 2.1 Definition of pH and Principles of Measurement.- 2.2 Donnan Effects.- 2.3 Methods for the Measurement of pH within Endocytic Vesicles and Lysosomes.- 2.3.1. Distribution of Weak Bases.- 2.3.2. Spectroscopic Methods.- 2.3.3. Other Methods.- 3. Lysosomal pH.- 3.1. Perturbation of Lysosomal pH.- 3.2. Role of Lysosome Acidification.- 4. Endocytic Vesicle pH.- 4.1. Consequences of Endocytic Vesicle Acidification.- 4.1.1. Receptor Recycling.- 4.1.2. Iron Release from Transferrin.- 4.1.3. Cytoplasmic Penetration by Viruses.- 4.1.4. Diphtheria Toxin Penetration.- 4.1.5. Summary of Biological Effects.- 5. Mechanism of Acidification.- 6. Summary.- References.- 9. Mathematical Modeling of Receptor-Mediated Endocytosis.- 1. Introduction.- 2. General Considerations of the Endocytic Pathway.- 2.1. Binding and Internalization.- 2.2. Ligand Degradation and Receptor Reutilization.- 3. General Models of the Endocytosis of Asialoglycoproteins.- 4. Surface Events and Internalization.- 4.1. The Interaction of Receptors with Coated Pits.- 4.2. The Interaction of Ligands with Receptors.- References.- 10. Morphologic Methods in the Study of Endocytosis in Cultured Cells.- 1. Introduction.- 2. Cytochemical Markers.- 2.1. Antibodies to Ligands and Receptors.- 2.2. Ligand Conjugates to Fluorochromes.- 2.3. Ligand Conjugates to Electron Microscopic Markers.- 2.3.1. Horseradish Peroxidase.- 2.3.2. Ferritin.- 2.3.3. Colloidal Gold.- 3. Light Microscopic Fluorescence and Image Intensification Methods.- 4. Electron Microscopic Morphologic Methods.- 4.1. Direct Embedding Technique for Cultured Cells.- 4.2. Membrane Contrast Enhancement Techniques.- 4.3. Serial Section Techniques.- 4.4. Stereo Analysis of Thin Sections.- 5 Immunocytochemistry.- 5.1. Light Microscopic Fluorescence.- 5.2. Electron Microscopic Immunocytochemical Methods.- 5.2.1. General Approaches.- 5.2.2. EGS and GBS Fixation and Processing Methods.- 5.2.3. Horseradish Peroxidase Labeling.- 5.2.4. Ferritin Bridge Labeling.- 6. Direct Mechanical Microinjection Methods.- 7. Experimental Protocols for the Study of Endocytosis.- References.