The mechanisms that direct positive-stranded RNA virus replication complexes to plant and animal cellular membranes are poorly understood. We describe a specific interaction between a replication protein of an RNA plant virus and membranes in vitro and in live cells. The tobacco etch virus (TEV) 6 kDa protein associated with membranes as an integral protein via a central 19 amino acid hydrophobic domain. In the presence or absence of other viral proteins, fluorescent fusion proteins containing the 6 kDa protein associated with large vesicular compartments derived from the endoplasmic reticulum (ER). Infection by TEV was associated with a collapse of the ER network into a series of discrete aggregated structures. Viral RNA replication complexes from infected cells were also associated with ER-like membranes. Targeting of TEV RNA replication complexes to membranous sites of replication is proposed to involve post-translational interactions between the 6 kDa protein and the ER.

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Formation of plant RNA virus replication complexes on membranes: role of an endoplasmic reticulum-targeted viral protein.

Nitrogen partitioning among proteins in chloroplasts and mitochondria was examined in pea (Pisum sativum L.) and wheat (Triticum aestivum L.) grown hydroponically with different nitrogen concentrations. In pea leaves, chloroplast nitrogen accounted for 75 to 80% of total leaf nitrogen. We routinely found that 8% of total ribulose-1,5-bisphosphate carboxylase/oxygenase adhered to thylakoids during preparation and could be removed with Triton X-100. With this precaution, the ratio of stroma nitrogen increased from 53 to 61% of total leaf nitrogen in response to the nitrogen supply, but thylakoid nitrogen remained almost constant around 20% of total. The changes in the activities of the stromal enzymes and electron transport in response to the nitrogen supply reflected the nitrogen partitioning into stroma and thylakoids. On the other hand, nitrogen partitioning into mitochondria was appreciably smaller than that in chloroplasts, and the ratio of nitrogen allocated to mitochondria decreased with increasing leaf-nitrogen content, ranging from 7 to 4% of total leaf nitrogen. The ratio of mitochondrial respiratory enzyme activities to leaf-nitrogen content also decreased with increasing leaf-nitrogen content. These differences in nitrogen partitioning between chloroplasts and mitochondria were reflected in differences in the rates of photosynthesis and dark respiration in wheat leaves measured with an open gas-exchange system. The response of photosynthesis to nitrogen supply was much greater than that of dark respiration, and the CO2 compensation point decreased with increasing leaf-nitrogen content.

Effects of Nitrogen Nutrition on Nitrogen Partitioning between Chloroplasts and Mitochondria in Pea and Wheat.

Membrane flow and interconversions among endomembranes

Sites in maize (Zea mays L.) coleoptile homogenates that reversibly bind naphthalene-1-acetic acid with high affinity and may represent receptor sites for auxins are located primarily on cellular membranes that show the enzymic and buoyant density characteristics of membranes of the rough endoplasmic reticulum. The sites remain attached to the endoplasmic reticulum (ER) membranes after the ribosomes have been stripped off them. Binding sites for naphthylphthalamic acid, an inhibitor of auxin transport, are located on membranes different from those that carry the naphthalene-1-acetic-acid (NAA)-binding sites, and which are probably plasma membrane. The two kinds of binding sites can be largely separated by appropriate density gradient centrifugation. The results raise the possibility that primary auxin action occurs at ER membranes and could represent facilitation of the transfer of hydrogen ions and nascent secretory protein into the ER lumen followed by secretory transport of these products to the cell exterior via the Golgi system.

Auxin-binding Sites of Maize Coleoptiles Are Localized on Membranes of the Endoplasmic Reticulum.

I. General Considerations and Background.- I.1. Perspectives.- I.2. Definitions.- I.2.1. General Functional Categories.- I.2.2. Outline of Lysosome Functioning in Phagocytes.- I.2.3. Additional Terms: Heterophagy and Autophagy.- I.3. Characterization of Lysosomes.- I.3.1. Basic Biochemical Characteristics of Lysosomes.- I.3.1.1. Key Features of Lysosomes.- I.3.1.2. The Lysosomal Enzymes.- I.3.1.3. Digestion in Lysosomes.- I.3.1.4. Lysosomal "Permeability".- I.3.2. Some Cytochemical and Morphological Characteristics.- I.3.2.1. Cytochemical Methods.- I.3.2.2. Cytochemical Studies of Exogenous Tracers.- I.3.2.3. Additional Cytochemical Features of Lysosomes: "Matrix" Materials and Some Other Non-Enzymatic Components.- I.4. Morphological Categories of Lysosomes.- II. Lysosome Formation, Functioning and Fate.- II. 1. Heterophagy.- II.1.1. Background.- II.1.2. Specificity.- II.1.3. The Transport of Acid Hydrolases.- II.1.3.1. The Formation of Primary Lysosomes in PMN Leukocytes.- II.1.3.2. Primary Lysosomes in Macrophages.- II.1.4. Fusion Phenomena, Lysosome "Reuse" and Lysosome Membranes.- II.1.4.1. Lysosome "Recycling".- II.1.4.2. Energetics and Control of Movements in Heterophagy.- II.1.4.3. Lysosome Membranes Fusion.- II.1.4.4. Other Features of the Lysosome Surface: Enzymes, Changes and Stability.- II.1.4.5. Acidification.- II.2. Hydrolase Transport in Cells Other than Phagocytes.- II.2.1. GERL.- II.2.2. Endoplasmic Reticulum and Lysosomes Some Biochemical Findings.- II.3. Autophagy.- II.3.1. Basic Morphology.- II.3.2. Mode of Formation of Autophagie Vacuoles.- II.3.2.1. Source of the Delimiting Membranes.- II.3.2.2. Source of the Hydrolases.- II.3.3. Crinophagy.- II.3.4. Some General Aspects of the Control and Specificity of Autophagy.- II.4. Multivesicular Bodies (MVB's).- II.4.1. Heterophagic Roles.- II.4.2. Participation in Autophagy.- II.4.2.1. Incorporation of Secretory Material and of Intact Vesicles.- II.4.2.2. Degradation of Membranes Participating in Endocytosis.- II.4.2.3. Possible Roles in Degrading Other Types of Membrane.- II.4.2.4. Microautophagy.- II.5. The Fate of Lysosomes.- II.5.1. Release vs. Retention.- II.5.2. "Telolysosomes" and Lipofuscin.- II.6. Lysosome Heterogeneity.- III. Lysosomes in Turnover and Modulation.- III.1. Turnover of Cells and Tissues of Higher Animals and Features of Developmental Remodelling.- III.1.1. Background and an Example: The Red Blood Cell Life History.- III.1.1.1. Lysosomes in the Maturation of Red Blood Cells.- III.1.1.2. The Destruction of Red Blood Cells.- III.1.1.3. Iron Storage and Release.- III.1.2. Developmental "Remodelling".- III.1.2.1. Case Histories: Cell Destruction in Insect Metamorphosis.- III.2. Turnover of Extracellular Materials.- III.2.1. Connective Tissue Components.- III.2.1.1. Hydrolase Release in Cartilage.- III.2.1.2. Osteoclasts.- III.2.1.3. Collagenases.- III.3. Turnover of Circulating Macromolecules.- III.3.1. Selectivity.- III.4. Intracellular Turnover.- III.4.1. Some Methodological Perspectives and Problems.- III.4.2. Turnover in Bacteria.- III.4.3. Turnover of Organelles and Intracellular Macromolecules in Eucaryotes.- III.4.3.1. Some General Points and Some Experiments Paralleling those Done with Procaryotes.- III.4.3.2. Turnover of Macromolecules that are not Components of Membrane-Delimited Organelles.- III.4.3.3. Aspects of the Turnover of Membrane-Delimited Cytoplasmic Organdies.- III.5. Turnover of Photoreceptor Membranes.- IV. Pathology.- IV.1. Lysosomal Storage Diseases.- IV.1.1. Lipidoses and Polysaccharidoses.- IV.1.2. Etiological Aspects.- IV.1.3. Some Interesting Disorders of Uncertain Status.- IV.2. Lysosomes and Infection.- IV.2.1. The Entry of Structures with Macromolecular Dimensions into Cells: Viruses and Toxic Proteins.- IV.2.2. Potentially Instructive Failures of Defenses.- IV.2.3. Malaria.- IV.3. The "Pathological" Release of Enzymes to Extracellular Spaces Arthritis, Inflammation, and Related Phenomena.- IV.4. The Intracellular Release of Hydrolases: Lysosome Fragility, Labilizers, and Stabilizers.- IV.4.1. Methodological Problems.- IV.4.2. Silica and Uric Acid.- IV.4.3. Labilizers, Stabilizers, Drugs, and Inhibitors.- IV.5. Lysosomes in Immune Responses.- IV.5.1. Macrophages and Some Other Cells in the "Processing" or "Presentation" of Antigens.- IV.5.2. Lysosomes in Lymphocyte Activation.- IV.5.3. The Transfer of Maternal Antibodies.- V. Some Special Topics and Some Loose Ends.- V.1. Lysosomes in Plant Cells.- V.1.1. Senescence and Cell Death.- V.1.2. Autophagy.- V.1.3. Extracellular Hydrolases.- V.2. Hydrolases in Secretory Cells.- V.2.1. Lysosomes and Secretory Processes the Thyroid Gland.- V.2.2. Hydrolases in the Golgi Apparatus and Secretion Granules.- V.2.3. Melanin.- V.3. Lysosomes in Animal Gametes.- V.3.1. Sperm.- V.3.2. Eggs.- V.4. Closing Comments.- Acknowledgements.

Lysosomes: A Survey

The properties of a discrete membranous fraction isolated on sucrose gradients from castor bean endosperm have been examined. This fraction was previously shown to be the exclusive site of phosphorylcholine-glyceride transferase. The distribution of NADPH-cytochrome c reductase and antimycin insensitive NADH-cytochrome c reductase across the gradient followed closely that of the phosphorylcholine-glyceride transferase. This fraction also had NADH diaphorase activity and contained cytochromes b5 and P 450. On sucrose gradients containing 1 mM EDTA this fraction had a mean isopycnic density of 1.12 g/cm3 and sedimented separately from the ribosomes; electron micrographs showed that it was comprised of smooth membranes. When magnesium was included in the gradients to prevent the dissociation of membrane-bound ribosomes, the isopycnic density of the membrane fraction with its associated enzymes was increased to 1.16 g/cm3 and under these conditions the electron micrographs showed that the membranes had the typical appearance of rough endoplasmic reticulum. Together these data show that the endoplasmic reticulum is the exclusive site of lecithin formation in the castor bean endosperm and establish a central role for this cytoplasmic component in the biogenesis of cell membranes.

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T. S. Moore

Papers

Endoplasmic reticulum as the site of lecithin formation in castor bean endosperm