Carbohydrate Metabolism in Cultured Cells

It is perhaps obvious to any student of Biology that the discovery of chemical processes in whole organisms has usually preceded the elucidation of the compo nent steps. However, it is perhaps less obvious that the unravelling of the se quences in which those chemical steps occur in living matter, of the precise mechanisms involved, and of the manner in which they are regulated, would have been achieved neither by the study of intact plants and animals nor even of extracts derived from them. Our ability to understand the nature and regulation of metabolism rests on two main premises: the postulate that life processes can indeed be validly investigated with individual cells and cell-free extracts, and the thesis that there is an essential "unity in biochemistry" (as Kluyver put it, 60 years ago) that enables events in one organism to be legitimately studied in another. Of particular utility in this latter respect has been the use of cultures of single-celled organisms, growing in defined media-especially prokaryotes, such as Escherichia coli, and eukaryotes, such as Neurospora and Sac charomyces sp. , to which both biochemical and genetical techniques could be applied. It was, of course, Pasteur's observations of bacterial fermentations that first overthrew the belief that oxygen was essential for all energy-yielding pro cesses: his recognition that "La fermentation . . . . . c' est La vie sans air" laid the foundations of our knowledge of glycolysis.

Inhalt
1 Studies of Regulation of Hexose Transport into Cultured Fibroblasts.- 1. Introduction.- 2. Hexose Uptake or Transport Tests.- 2.1. Effects of Cycloheximide on Hexose Transport Regulation.- 2.2. Is the Release of the Mediated Curb of the Hexose Transport System Dependent on Transcription?.- 3. Metabolic Pathways.- 4. Characterization of Certain Cellular Macromolecules and Structures.- 4.1. Enzyme Assays of the Hexose Uptake System in Lysed Cells.- 4.2. Membrane-Associated Glucose-Binding Proteins Released without Cell Lysis.- 4.3. Studies on Isolated Plasma Membrane Preparations in Regard to Their Hexose Transport Population and Identification of Specific Transport Proteins.- 5. Effects of Glucose Starvation on a Variety of Plasma Membrane Proteins.- 6. The Glucose-Mediated Curb of Hexose Transport Requires Oxidative Energy.- 7. Nucleoside Triphosphate Levels in Cultured Fibroblasts as a Function of General Metabolism and Nutrition.- 8. Hexose Transport Regulation and Oncogenic Transformation of Cultured Fibroblasts.- 9. Evolutionary Aspects.- References.- 2 The Utilization of Carbohydrates by Animal Cells: An Approach to Their Biochemical Genetics.- 1. The Utilization of Carbohydrates.- 2. Glycolysis.- 2.1. Glucokinase.- 2.2. Hexokinase.- 2.3. Phosphoglucose Isomerase.- 2.4. Phosphofructokinase.- 2.5. Glyceraldehyde-3-phosphate Dehydrogenase.- 2.6. Phosphoglycerate Kinase.- 2.7. Other Glycolytic Enzymes.- 3. The Provision of Energy.- 3.1. Pyruvate Metabolism.- 4. Pentose Phosphate Pathway.- 4.1. Glucose-6-phosphate Dehydrogenase.- 5. Differentiation.- 5.1. Alternative Carbon Sources.- 5.2. Gluconeogenesis.- 6. Other Effects of Carbohydrates.- 6.1. Effects on Morphology.- 6.2. Glucose-Regulated Proteins.- 6.3. Hypergravity.- 7. Concluding Remarks.- References.- 3 Biochemical Genetics of Respiration-Deficient Mutants of Animal Cells.- 1. Introduction.- 2. The Selection of Respiration-Deficient Mammalian Cell Mutants.- 2.1. Characterization of the First Mutant.- 2.2. Protocol for the Isolation of Additional Mutants.- 3. Glycolysis and Respiration in Wild-Type Parents and res Mutants.- 4. Biochemical Characterization of Mutants.- 4.1. Defect in NADH-CoQ Reductase.- 4.2. Defect in Succinate Dehydrogenase.- 4.3. Defect in Mitochondrial Protein Synthesis.- 5. Genetic Characterization of Mutants.- 6. Work in Progress and Future Prospects.- 7. Summary.- References.- 4 Glutaminolysis in Animal Cells.- 1. Glutamine Metabolism in Mammals.- 2. Glutaminolysis in Tissues.- 2.1. Liver and Kidney.- 2.2. Brain.- 2.3. Pancreas.- 2.4. Mammary Gland.- 2.5. The Intestine.- 2.6. Embryonic and Placental Tissue.- 2.7. Tumors.- 3. Glutaminolysis in Isolated Tissues and Primary Cell Suspensions.- 3.1. Enterocytes.- 3.2. Blood Cells.- 3.3. Lens.- 3.4. Germ Cells.- 3.5. Calvaria.- 3.6. Astrocytes.- 4. Glutaminolysis in Normal and Tumor Cells in Culture.- 4.1. Tumor-Derived Cells and Transformed Cell Lines.- 4.2. Untransformed Normal Cells in Culture.- 5. Glutaminolysis-The Pathway of Glutamine Oxidation.- 5.1. Enzymology.- 5.2. Compartmentation.- 5.3. Regulation of Glutaminolysis.- 6. Glutaminolysis and Glycolysis in Cell Growth and Function.- 6.1. The Role of Glycolysis in Cell Proliferation.- 6.2. Can Cells Proliferate in the Absence of Glutaminolysis?.- 6.3. The Role of Glutaminolysis in Cell Specialization.- 6.4. Potential Impact of Tumor Glutaminolysis on Host Glutamine and Glucose Metabolism.- 7. Conclusions.- 8. Addendum.- References.- 5 The Metabolism and Utilization of Carbohydrates by Suspension Cultures of Plant Cells.- 1. Introduction.- 2. Carbon Sources for Culture Growth.- 2.1. The Range of Carbon Sources Tested.- 2.2. Effects on Growth (Physiology and Biochemistry).- 2.3. Effects on Natural Product Synthesis.- 3. Uptake Mechanisms for Carbon Sources.- 3.1. Differential Mechanisms.- 3.2. Cellular Location.- 3.3. Effect of Internal Pools.- 4. Intracellular Fate of Carbon Source-Biochemistry: Oxidation, Biosynthesis, Storage.- 5. Summary Comments.- References.- 6 Carbohydrate Metabolism in African Trypanosomes, with Special Reference to the Glycosome.- 1. Introduction.- 1.1. General Background and Scope.- 1.2. Biology of the Kinetoplastida.- 1.3. Medical and Economic Importance of African Trypanosomiasis.- 2. Life Cycle.- 3. Methods for Cultivation.- 3.1. Mammalian Forms of T. brucei.- 3.2. Insect Forms of T. brucei.- 4. Substrates and End Products of Metabolism.- 4.1. Mammalian Forms.- 4.2. Insect Forms of the brucei Subgroup.- 5. Terminal Respiratory Systems.- 5.1. Mammalian Forms.- 5.2. Insect Forms.- 6. The Glycosome.- 6.1. Occurrence.- 6.2. Isolation and Purification.- 6.3. General Properties.- 6.4. Glycosomal Enzymes in T. brucei.- 7. Pathways of Glucose Metabolism.- 7.1. Long-Slender Trypomastigotes.- 7.2. Short-Stumpy Trypomastigotes.- 7.3. Procyclic Trypomastigotes.- 8. Glycolysis as a Target for Chemotherapy.- 8.1. Energy Metabolism.- 8.2. Action of Trivalent Aromatic Arsenicals.- 8.3. Action of Suramin.- 8.4. Action of SHAM-Glycerol.- 9. Summary and Outlook.- References.- 7 Sugar Transport Systems of Baker's Yeast and Filamentous Fungi.- 1. Yeast.- 1.1. Monosaccharide Transport Systems.- 1.2. Disaccharide Transport Systems.- 2. Filamentous Fungi.- 2.1. Aspergillus nidulans.- 2.2. Neurospora crassa.- References.- 8 Carbohydrate Metabolism in Yeast.- 1. Introduction.- 2. Methodological Approaches.- 2.1. Determination of Enzymatic Activities.- 2.2. Determination of Metabolite Levels.- 2.3. Fate of Labeled Substrates: Calculation of the Proportion of Carbohydrates Used through Different Pathways.- 2.4. Continuous Cultures versus Batch Cultures.- 2.5. Use of Mutants.- 3. Overview of the Pathways of Carbohydrate Metabolism.- 3.1. Glycolytic Pathway.- 3.2. Metabolism of Carbohydrates Other Than Glucose.- 3.3. Pentose Phosphate Pathway.- 3.4. Citric Acid Cycle.- 3.5. Reserve Carbohydrates.- 4. Regulatory Mechanisms.- 4.1. Catabolite Repression.- 4.2. Catabolite Inactivation.- 4.3. Pasteur Effect and Other Effects.- 4.4. Futile Cycles.- 5. Conclusions.- References.- 9 Regulation of Carbon Metabolism in Filamentous Fungi.- 1. Introduction.- 2. Extracellular Formation of Hexoses from Polysaccharides.- 2.1. Cellulose Degradation.- 2.2. Hemicellulose (Xylan) Degradation.- 2.3. Starch Degradation.- 3. Growth on Glucose and Related Carbon Sources.- 3.1. General Considerations.- 3.2. Enzymes of Hexose Metabolism.- 3.3. Synthesis of Reserve Carbohydrates.- 3.4. Pathways of O…