Interaction of melittin with dimyristoyl phosphatidylcholine liposomes. Evidence for boundary lipid by raman spectroscopy

The interaction of melittin, a polypeptide consisting of 26 amino acid residues, with dimyristoyl phosphatidylcholine bilayers was investigated by vibrational Raman spectroscopy. Spectral peak height intensity ratios, involving vibrational transitions in both the 3000 cm^?1 acyl chain methylene carbon-hydrogen stretching mode region and the 1100 cm^?1 acyl chain carbon-carbon skeletal stretching mode interval, served as temperature profile indices for monitoring the bilayer order-disorder processes. For a lipid : melittin molar ratio of 14 : 1 two order-disorder transitions were observed. In comparison to a gel to liquid crystalline phase transition of 22.5°C for the pure lipid, the lower transition, exhibiting a 2°C width, is centered at 17°C and is associated with a depression of the main lipid phase transition of dimyristoyl phosphatidylcholine. The second thermal transition, displaying a 7°C interval, occurs at approx. 29°C and is associated with the melting behavior of approximately seven immobilized boundary lipids which surround the inserted hydrophobic segment of the polypeptide. For a lipid : melittin molar ratio of 10 : 1 two thermal transitions are also observed at 11 and 30°C. As before, they represent, respectively, the main gel to liquid crystalline phase transition and the melting behavior of approximately four boundary lipids attached to melittin. From these data alternative schemes are suggested for disposing the immobilized lipids around the hydrophobic portion of the polypeptide within the bilayer.     

Occurrence of subcutaneous zygomycosis (Entomophthoromycosis Basidiobolae) caused by Basidiobolus haptosporus with pulmonary involvement

A case of a two-year-old boy with multiple subcutaneous lesions caused by Basidiobolus haptosporus is presented.The child had also a non-toxic familial goiter and clinical and radiological features of a pulmonary illness. The pulmonary manifestations only disappeared with the treatment with potassium iodide. The authors think that the pulmonary lesions must have arisen by direct spread of the fungus from the subcutaneous lesions of the chest.     

Infectivity of Proviral DNA from Avian Sarcoma Virus-Transformed Mammalian Cells

The number of Rous viral genomes in the cellular DNA from two subclones (RS2/3, RS2/6) derived from the same clone of hamster BHK-21 cells transformed by Rous sarcoma virus was determined by hybridization with viral complementary DNA made in vitro, and the capacity of the cellular DNA to infect (transfect) chicken embryo fibroblasts was compared before and after shearing this DNA to about the size of the provirus (6 x 10(6) to 7 x 10(6) daltons). The two subclones differed widely both in their capacity to give rise to virus (inducibility) after fusion with chicken embryo fibroblasts and in level of expression of viral proteins. It was shown that cells of both subclones contain a single copy of Rous DNA and yield infectious DNA. However, whereas transfection of chicken embryo fibroblasts was successful with both unsheared (>/=18 x 10(6) daltons) and sheared DNA from the most inducible subclone (RS2/3 subclone), which also expresses viral proteins to an appreciable amount, transfection with DNA from the least inducible subclone (RS2/6 subclone), in which viral proteins are not expressed, succeeded only with sheared DNA. It was then about as successful as with sheared or unsheared RS2/3 DNA. The lack of infectivity of unsheared RS2/6 DNA may be explained by the hypothesis proposed by Cooper and Temin (G. M. Cooper and H. T. Temin, J. Virol. 17:422-430, 1976) to explain the lack of infectivity of DNA from certain chicken cells producing spontaneously low amounts of RAV-0 and resistant to exogenous RAV-0 infection, that is, that the viral genome (proviral DNA) is linked to a cis-acting control element which blocks its expression. This linkage might originate, in RS2/6 cells, from translocation of cellular DNA containing the single proviral copy.     

Construction of a colony bank of E. coli containing hybrid plasmids representative of the Bacillus subtilis 168 genome. Expression of functions harbored by the recombinant plasmids in B. subtilis.

Summary A collection of about 2500 clones containing hybrid plasmids representative of nearly the entire genome of B. subtilis 168 was established in E. coli SK1592 by using the poly(dA)·poly(dT) joining method with randomly sheared DNA fragments and plasmid pHV33, a bifunctional vector which can replicate in both E. coli and B. subtilis. Detection of cloned recombinant DNA molecules was based on the insertional inactivation of the Tc gene occurring at the unique BamHI cleavage site present in the vector plasmid.Thirty individual clones of the collection were shown to hybridize specifically with a B. subtilis rRNA probe. CCC-recombinant plasmids extracted from E. coli were pooled in lots of 100 and used to transform auxotrophic mutants of B. subtilis 168. Complementation of these auxotrophic mutations was observed for several markers such as thr, leuA, hisA, glyB and purB. In several cases, markers carried by the recombinant plasmids were lost from the plasmid and integrated into the chromosomal DNA. Loss of genetic markers from the hybrid plasmids did not occur when a rec ^- recipient strain of B. subtilis was used.Abbreviations Ap^R resistance to ampicillin - Tc^R resistance to tetracycline - Cm^R resistance to chloramphenicol - CCC covalently closed circular duplex - Mdal magadalton     

Bactericidal activity of tuftsin

Summary The biological activities of the phagocytosis stimulating tetrapeptide, Thr-Lys-Pro-Arg are discussed. A brief account on the stimulation by tuftsin of phagocytosis of various particles, including bacteria was reported. Stimulation of bactericidal activity by this tetrapeptide was investigated in vitro as well as in vivo. The potency of tuftsin to enhance blood clearing of Staphylococcus aureus, Listeria monocytogenes, Escherichia coli and Serratia marcescens by mouse peritoneal macrophages was demonstrated.Bactericidal activity and effects of tuftsin on this phenomenon were studied in liver and spleen of mice. Tuftsin stimulates these activities. Same experiments were performed in infected leukemic mice by Serratia marcescens or Escherichia coli. Results on blood clearing and bactericidal activities in liver and spleen were reported and compared to those of healthy and leukemic untreated animals. Tuftsin was found to present interesting stimulatory effects on the bactericidal activity of phagocytes.     

S-adenosyl methionine requiring mutants in Saccharomyces cerevisiae: evidences for the existence of two methionine adenosyl transferases.

Summary Mutants requiring S-adenosyl methionine (SAM) for growth have been selected in Saccharomyces cerevisiae. Two classes of mutants have been found. One class corresponds to the simultaneous occurrence of mutations at two unlinked loci SAM1 and SAM2 and presents a strict SAM requirement for growth on any medium. The second class corresponds to special single mutations in the gene SAM2 which lead to a residual growth on minimal medium but to normal growth on SAM supplemented medium or on a complex medium like YPGA not containing any SAM. These genetic data can be taken as an indication that Saccharomyces cerevisiae possesses two isoenzymatic methionine adenosyl transferases (MAT). In addition, SAM1 and SAM2 loci have been identified respectively with the ETH-10 and ETH2 loci previously described.Biochemical evidences corroborate the genetic results. Two MAT activities can be dissociated in a wild type extract (MATI and MATII) by DEAE cellulose chromatography. Mutations at the SAM1 locus lead to the absence or to the modification of MATII whereas mutations at the SAM2 locus lead to the absence or to the modification of MATI. Moreover, some of our results seem to show that MATI and MATII are associated in vivo.     

Analyse quantitative des remaniements intestinaux et évolution de la synthese de l'ADN

Résumé Les remainements intestinaux affectant la paroi intestinale sont quantifiés chez les larves d'un Amphibien Anoure,Alytes obstetricans, traitées par la thyroxine. La fraction correspondant à chaque zone composant la paroi est déterminée sur coupe. La répartition et la fréquence des noyaux marqués sont recherchées chez des larves ayant reçu de la thymidine tritiée.L'indice de marquage nucléaire de l'épithélium primaire tend vers zéro durant les 3 premiers jours du traitement. Ces résultats reflètent un ralentissement de la croissance du tissue, précédant sa dégénérescence et son élimination complète au 14ème jour d'immersion dans la solution hormonale.Dès de 3ème jour de traitement, un épithélium secondaire se développe à partir de cellules-souches basales. Son indice de marquqge croît, culmine au 9ème jour avec une valeur proche de 50%, puis s'abaisse. La prolifération cellulaire est plus modérée dans la zone extra-épithéliale, qui s'épaissit toutefois au cours de cette métamorphose provoquée.

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Quantitative analysis of intestinal changes and the evolution of DNA synthesis in thyroxine-treated larvae ofAlytes obstetricans. (Amphibia, Anura)

Summary Changes in the intestinal wall ofAlytes obstetricans (Amphibia, Anura) were quantified during thyroxine treatment. The relative proportion of each intestinal wall component was determined in tissue sections. Autoradiographic studies on intestine with³H-thymidine revealed the distribution and frequency of labeled nuclei.The labeling index in the primary epithelium falls to zero by 3 days. Thyroxine treatment induces a decrease in the growth rate of this tissue, just before its degeneration and complete elimination by 14 days.Three days after T₄-treatment, a secondary epithelium develops from basal stem cells. Its labeling index rises to a maximum of about 50% by 9 days, and thereafter declines. Cell proliferation is less marked in the extraepithelial zone, which nevertheless thickens during this thyroxine-induced metamorphosis.

     

Mitochondrial DNA evolution in lagomorphs: Origin of systematic heteroplasmy and organization of diversity in European rabbits

Summary A characterization was conducted on mitochondrial DNA (mtDNA) molecules extracted separately from 107 European rabbits (Oryctolagus cuniculus) both wild and domestic, 13 European hares (Lepus capensis), and 1 eastern cottontail (Sylvilagus floridanus). Experimentally this study took into account restriction site polymorphism, overall length variation of the noncoding region, and numbers of repeated sequences. Nucleotide divergences indicate that the mtDNAs from the three species derived from a common ancestor some 6–8 million years (Myr) ago. Every animal appeared heteroplasmic for a set of molecules with various lengths of the noncoding region and variable numbers of repeated sequences that contribute to them. This systematic heteroplasmy, most probably generated by a rate of localized mtDNA rearrangements high enough to counterbalance the cellular segregation of rearranged molecules, is a shared derived character of leporids.The geographic distribution of mtDNA polymorphism among wild rabbit populations over the western European basin shows that two molecular lineages are represented, one in southern Spain, the second over northern Spain, France, and Tunisia. These two lineages derived from a common ancestor some 2 Myr ago. Their present geographical distribution may be correlated to the separation of rabbits into two stocks at the time of Mindel glaciation.Finally the distribution of mtDNA diversity exhibits a mosaic pattern both at inter- and intrapopulation levels.     

Interactions of surfactin,a biosurfactant fromBacillus subtilis,with inorganic cations

Summary The properties of surfactin, a biosurfactant lipopeptide, were highly modified in the presence of inorganic cations. The micellization of surfactin was favoured by monovalent and, especially by divalent cations with a modification of the molecular area at the air-water interface. Haemolysis of erythrocytes by surfactin was enhanced by low concentrations of divalent cations with an increase of the binding of the lipopeptide to membrane. Inorganic ions induced conformational rearrangements probably due to ion-surfactin associations which modify the surface-active properties.