Chemical composition of the plasma membrane from epimastigote forms of Trypanosoma cruzi

The chemical composition of two plasma membrane fractions from epimastigote forms of Trypanosoma cruzi is reported. Fraction M, a preparation obtained by conventional methods of cell fractionation is composed of 31% proteins, 34% lipids, 16% carbohydrates and 3% of the lipopeptidophosphoglycan. Phospholipids and sterols account for 7.5 and 9%, respectively, of the total mass. Phosphatidylethanolamine is the major phospholipid in fraction M, representing 45% of the total membrane phospholipids. The other fraction, fraction V (vesicles), was obtained by treatment of the cell with a vesiculating agent. This fraction contains 42% lipids, 20% carbohydrates, 13% proteins and 21% of the lipopeptidophosphoglycan. Phospholipids and sterols make up 17 and 8%, respectively, of the total mass of this fraction. Phosphatidylcholine and phosphatidylethanolamine are the main phospholipids found in fraction V. Phosphonolipids and sialic acid have not been detected in either membrane fraction. Sodium dodecyl sulphate polyacrylamide gel electrophoretic analysis show that the glycoproteins ABC and the lipopeptidophosphoglycan are 50- and 10-times more concentrated, respectively, in fractions V and M than in the whole cell homogenate. The high molar sterol/phospholipid ratio found in fraction M suggests that this fraction is less fluid than fraction V, perhaps reflecting a migration of certain membrane components in the presence of the vesiculating agent. Hence, fraction M is, probably, more representative of the epimastigote plasma membrane as a whole than fraction V.     

Inhibition of phospholipid methyltransferase during zymosan induced secretion of platelet-activating factor in human polymorphonuclear leukocytes

Phagocytosis of zymosan particles coated with complement induces a time and dose dependent inhibition of the enzyme phospholipid methyltransferase in human polymorphonuclear cells. The extent of phospholipid methyltransferase inhibition induced by various concentrations of zymosan strongly correlates with the secretory process: liberation of platelet-activating factor (PAF) and β-glucuronidase. Zymosan also decreases the incorporation of ³H-methyl group into phospholipids in cells pre-labeled with (³H-methyl)-methionine. Finally, preincubation of cells with 3-deaza-adenosine and homocysteine thiolactone, inhibitors of phospholipid methyltransferase, decrease the incorporation of ³H-methyl group into phospholipids in cells pre-labeled with (³H-methyl)-methionine and modulate the release of PAF. These results suggest that phospholipid methylation plays an important role during the transduction of the secretory signal triggered by zymosan in human polymorphonuclear cells.     

Trypanosoma cruzi: defined medium for continuous cultivation of virulent parasites.

A liquid medium was developed for the continuous cultivation of Trypanosoma cruzi. Among the several highly purified macromolecules tested only bovine liver catalase, horseradish peroxidase, lactoperoxidase, and bovine hemoglobin supported the continuous growth, at high yield, of mice-virulent Trypanosoma cruzi; other hemoproteins were inactive. Bovine liver catalase showed optimal Trypanosoma cruzi growth-promoting activity, parasites reaching 20 × 10⁶ parasites/ml (95% epimastigotes) at about 10 days in most of the 45 subpassages to date. Furthermore, this protein in the incubation medium provided all the amino acid requirements of actively growing parasites, thus eliminating the need for exogeneous free amino acids. Additional experiments revealed that the hemoprotein's growth-promoting activity was independent of any enzymatic activity and that reconstituting the exact protein composition by means of exogeneous amino acids did not support parasite multiplication, suggesting the importance of the primary structure of the active proteins for growth-promoting activity. These active macromolecules supported the multiplication of five different strains of Trypanosoma cruzi, but did not support Leishmania brasiliensis or Leishmania mexicana proliferation, suggesting species specificity.     

Subsurface cisterns in paraventricular nuclei of the hypothalamus of the rat

Summary Structures identified as subsurface cisterns (SSC's) were found in neurons of the paraventricular nuclei of the rat hypothalamus. They appeared as cytoplasmic organelles consisting most often of stacks of parallel cisterns apposed to the neuronal plasmalemma. These SSC's were located in the interneurons of the parvocellular system, but not in neurosecretory cells and glial cells. SSC's were seen at zones of cytoplasm apposed to neuronal or glial cell processes, showing in some instances specific relationships with synaptic areas.The morphological features of these SSC's are described, and their possible functional significance is briefly discussed.     

Adenylyl cyclase deficient cr-1 (Crisp) mutant of Neurospora crassa: Cyclic AMP-dependent nutritional deficiencies

The inability to synthesize cyclic AMP drastically affects the nutritional metabolism of Neurospora crassa. The adenylyl cyclase-less mutant cr-1 (crisp) did not utilize several carbon sources, including glycerol, mannitol, arabinose, and casaminoacids. However, in glucose or acetate it grew as well as the wild type. The following evidence suggested that these nutritional deficiencies were a direct result of the cr-1 mutation: (i), in crosses to wild type they segregated together with the crisp morphological marker; (ii), cyclic AMP added to the cr-1 mutant growth medium overcame the nutritional deficiencies; (iii), the cyclic AMP effect was specific for the crisp mutant, for it was not observed with the wild type, nor with a spontaneous glycerol-utilizing cr-1 strain.     

Empirical examination of the threshold model of neuron firing

An elementary model of neuronal activity involves temporal and spatial summation of postsynaptic currents that are elicited by presynaptic spikes and that, in turn, elicit postsynaptic potentials at a trigger zone; when the potential at the trigger zone exceeds a threshold level, a postsynaptic spike is generated. This paper describes three methods of estimating the summation function, that is, the function of time that converts the synaptic current into potential at the trigger zone: namely, maximum likelihood, cross-correlation analysis and cross-spectral analysis. All three methods, when applied to input-output data collected on various neurons of Aplysia californica, give comparable results. As estimated, the summation function involved in the explored cells has an early positive-going swing that is large and brief. In the cell L5, but not in R2, there was also a late negative-going swing of longer duration.Prepared with the partial support of the National Science Foundation Grant MCS 77-22986 to DRB and NSF and NIH grants to JPS     

Inhibition of the respiration of dormant and swollenStreptomyces spores by chloramphenicol

Chloramphenicol produces a decrease in the respiratory quotient of dormant and swollenStreptomyces antibioticus spores of about 20–25% and 18–24%, respectively, in the absence of protein synthesis as measured by [³H]leucine incorporation and by chemical methods. Rifampin and streptomycin had no inhibitory effect on respiration, thus excluding the possibility of an effect of all protein synthesis inhibitors on respiration. The inhibition of respiration by chloramphenicol was not a consequence of an increase in mortality due to toxicity, nor was it influenced by the developmental stage of the spores. It seems that chloramphenicol affects the activity of some component(s) related to the electron transport chain ofS. antibioticus spores, situated before the cytochrome oxidase level.     

Synchronous growth of a freshwater diatom Melosira italica under natural environment

Summary A filamentous diatom Melosira italica was collected at the beginning of rainy season from a shallow lake in the tropical savanna region in Brazil. Even the sample taken from surface water contained empty cells in high percentages. The number of cells per filament of M. italica showed a peculiar pulse-like frequency distribution with peak values at 4, 8, 12 and 16. Evidences of the synchronous cell division in this planktonic diatom under natural environment are discussed.     

Neolignans from an Aniba species

A benzene extract of the trunk of an Aniba species (Lauraceae) contained benzyl benzoate, benzyl salicylate, sitosterol and the neolignans (2S,3S,3aR)-3a-allyl-5-methoxy-3-methyl-2-piperonyl-2,3,3a,6-tetrahydro-6-oxobenzofuran (burchellin); (2S,3S,3aR)-3a-allyl-5-methoxy-3-methyl-2-veratryl-2,3,3a,6-tetrahydro-6-oxobenzofuran; (2S,3S,3aR)-3a-allyl-5,7-dimethoxy-3-methyl-2-veratryl-2,3,3a,6-tetrahydro-6-oxobenzofuran; (2S,3S,5S)-5-allyl-5-methoxy-3-methyl-2-veratryl-2,3,5,6-tetrahydro-6-oxo-benzofuran; (2R,3R)-7-methoxy-3-methyl-5-propenyl-2-veratryl-2,3-dihydrobenzofuran; rel-(1R,5R,6R,7R,8S)-1-allyl-8-hydroxy-3-methoxy-7-methyl-4-oxo-6-piperonylbicyclo[3,2,1]oct-2-ene (guianin); rel-(1S,5S,6S,7R,8R)-1-allyl-8-hydroxy-3,5-dimethoxy-7-methyl-4-oxo-6-piperonylbicyclo[3,2,1]oct-2-ene; rel-(1S,5S,6S,7R,8R)-8-acetoxy-1-allyl-3-hydroxy-5-methoxy-7-methyl-4-oxo-6-piperonyl-bicyclo[3,2,1]oct-2-ene; rel-1S,5S,6S,7R,8R)-8-acetoxy-3,5-dimethoxy-7-methyl-4-oxo-6-piperonylbicyclo[3,2,1]oct-2-ene; rel-(1R,5S,6R,7R)-1-allyl-3-methoxy-7-methyl-4,8-dioxo-6-piperonylbicyclo[3,2,1]oct-2-ene.     

Membrane adenosine triphosphatase of Micrococcus lysodeikticus. Isolation of two forms of the enzyme complex and correlation between enzymatic stability,latency and activity.

Summary Two new forms of the plasma membrane ATP-ase ofMicrococcus lysodeikticus NCTC 2665 were isolated from a sub-strain of the microorganism by polyacrylamide gel electrophoresis. One of them had a mol.wt of 368,000 and a very low specific activity (0.80 µ mol.min^–1.mg protein^–1) that could not be stimulated by trypsin. This form has been called B^I (strain B, inactive). If the electrophoresis was carried out in the presence of reducing agents (i.e., dithiothreitol) and the pH of the effluent maintained at a value of 8.5 another form of the enzyme was obtained. This had a mol.wt of 385,000 and a specific activity of 2.5–5.0 µ mol.min^–1.mg protein^–1 that could be stimulated by trypsin to 5–10 µ mol.min^–1.mg protein^–1. This preparation of the ATPase has been called form B_A (strain B, enzyme active). The subunit composition of both forms has been studied by sodium dodecyl sulphate and urea gel electrophoresis and compared to that of the enzyme previously purified from the original strain (form A). The three forms of the enzyme had similar and subunits, with mol.wt of about 50,000 and 30,000 dalton, respectively. They also had in common the component(s) of relative mobility 1.0, whose status as true subunit(s) of the enzyme remains yet to be established. However, subunit, that had a mol.wt of about a 52,500 in form A (Andreu et al. Eur. J. Biochem. (1973) 37, 505–515), had a mol.wt similar to in form B_I and about 60,000 in form B_A. Furthermore B_A usually showed two types of this subunit ( and) and an additional peptide chain () with a mol.wt of about 25,000 dalton. This latter subunit seemed to account for the stimulation by trypsin of form B_A.Forms B_A could be converted to B_I by storage and freezing and thawing. Conventional protease activity could not be detected in any of the purified ATPase forms and addition of protease inhibitors to form B_A failed to prevent its conversion to form B_I. The low activity form (B_I) was more stable than the active forms of the enzyme and also differed in its circular dichroism. These results show thatM. lysodeikticus ATPase can be isolated in several forms. Although these variations may be artifacts caused by the purification procedures, they provide model systems for understanding the structural and functional relationships of the enzyme and for drawing some speculations about its functionin vivo.