Sub-cellular Localization of Calcium in Azolla-Anabaena Symbiosis by Chlortetracycline,ESI and EELS

The subcellular localization of calcium in cells of symbiotic partners located within leaf cavities of Azolla was investigated by using chlorotetracycline, ESI and EELS analysis. Loosely membrane-bound calcium was evidenced by using CTC or EGTA and CTC, in cytoplasmic regions of Azolla hair cells and in cytoplasm of the cyanobiont. Tightly membrane-bound calcium revealed by CTC, and ESI and EELS analysis, was observed in cyanophycin granules and carboxysomes of the cyanobiont. A third calcium type, revealed by ESI and EELS analysis, was localized at the level of cell walls of simple and branched Azolla hairs, in the envelope of heterocysts, and in the cell walls of the cyanobiont.     

The genomic instability of yeast cdc6-1/cdc6-1 mutants involves chromosome structure and recombination

When diploid cells of Saccharomyces cerevisiae homozygous for the temperature-sensitive cell division cycle mutation cdc6-1 are grown at a semipermissive temperature they exhibit elevated genomic instability, as indicated by enhanced mitotic gene conversion, mitotic intergenic recombination, chromosomal loss, chromosomal gain, and chromosomal rearrangements. Employing quantitative Southern analysis of chromosomes separated by transverse alternating field gel electrophoresis (TAFE), we have demonstrated that 2N-1 cells monosomic for chromosome VII, owing to the cdc6-1 defect, show slow growth and subsequently yield 2N variants that grow at a normal rate in association with restitution of disomy for chromosome VII. Analysis of TAFE gels also demonstrates that cdc6-1/cdc6-1 diploids give rise to aberrant chromosomes of novel lengths. We propose an explanation for the genomic instability induced by the cdc6-1 mutation, which suggests that hyper-recombination, chromosomal loss, chromosomal gain and chromosomal rearrangements reflect aberrant mitotic division by cdc6-1/cdc6-1 cells containing chromosomes that have not replicated fully.     

Conformation and interactions of bombolitin I analogues with SDS micelles and phospholipid vesicles: CD,fluorescence, two-dimensional NMR and computer simulations

Bombolitins are five structurally related heptadecapeptides acting at the membrane level able to lyse erythrocytes and liposomes and to enhance the activity of phospholipase A ₂(PLA₂). In the presence of SDS or phospholipid vesicles bombolitins are able to form amphiphilic α-helical structures and this property seems to be the major determinant of bioactivity. In order to test the model of interaction between bombolitin I and membranes, an analogue was synthesized in which all the lysines were replaced by arginines: ([Arg^2,9,12, Ile¹⁰] bornbolitin I). The design ofthis sequence allowed the synthesis of a second analogue through a specijic postsynthetic dansylation at the ?-amino group qf a lysine residue replacing the original leucine residue at position 7. The, first analogue was, fiilly characterized by CD and two-dimensional nmr in the presence of SDS or phospholipid vesicles. The peptide, folds into an amphiphilic α-helical confbrrnation with the helical segment spanning the central part of the sequencefrom Ile³ to His¹⁶. This behavior is identical to that observed for the native sequence. The replacement of Iysine residues by arginine hus no detectable effect on the conformational prderence of the peptide chain. By CD and fluorescence spectroscopy measurements, the fluorophore-containing analogue [Arg^2,9,12, Lys⁷(?-dansyl)] bombolitin I also folded into the α-helical conformation in the presence of SDS micelles or phospholipid vesicles. In particular, the dansyl fluorophore, which is located approximately in the middle of the apolar surface ojthe amphiphilic helix, is clearly buried in a hydrophobic environment when the peptide is bound to phospholipid vesicles. These findings support the hypothesis that the peptide helices are oriented parallel to the vesicle surface. © 1995 John Wiley & Sons, Inc.     

Use of sequence characterised amplified region and RAPD markers in the identification of the white truffle Tuber magnatum Pico

Isolates of white truffles were identified as Tuber magnatum Pico species using a pair of primers selected from a sequence characterised amplified region (SCAR) and a specific random amplified polymorphic DNA (RAPD) marker. The present study reveals that PCR-fragment-pattern polymorphisms, the construction of probes and couples of primers from one or more of these polymorphic fragments may provide a useful and rapid tool for identifying species of ectomycorrhizal fungi in addition to conventional methods (morphological parameters).     

Calcium uptake in mosses and its role in stone biodeterioration

The biodeterioration of stone substrata by bryophytes (Musci and Hepaticae) is due to biogeochemical and biogeophysical mechanisms. In this work the biogeochemical damage caused by epilithic moss species, found in archaeological sites of Rome, was investigated. The cellular calcium content in specimens of Grimmia pulvinata (Hedw.) Sm. was analysed using a sequential elution technique. The analyses were carried out on moss specimens sampled from different lithotypes and in different seasons in order to measure the calcium concentration both in relation to the mineral composition of the stone and to the plant physiology. The highest calcium concentration of the extracellular exchangeable fraction was detected in the samples grown on marble and in the waterlogged specimen sampled in spring time.     

A common β hexosaminidase gene mutation in adult Sandhoff disease patients

-Hexosaminidase gene mutations were analyzed in two adult-onset Sandhoff disease Italian patients by PCR analysis of a common known mutation (5) and by heteroduplex analysis of genomic and RT-PCR DNA fragments, covering the whole gene. The patients' genotypes were 5/C1214T, and G890A/C1214T, respectively. As mutation C1214T (Pro405Leu) is also present in the other two late-onset cases so far described, we suggest that C1214T is a common mutation in this type of Sandhoff disease. Mutation G890A (Cys297Tyr) is a novel mutation which presumably causes altered processing of the pro chain.     

Synthesis of optically active diols using an efficient polymer bound cinchona alkaloid derivative

A new insoluble polymer containing a Cinchona alkaloid derivative has been synthesized and used as chiral ligand in the heterogeneous enantioselective dihydroxylation of olefins. It is shown that the enantioselectivity of the optically active diols obtained from both aliphatic and aromatic substrates is always comparable to that observed in the homogeneous phase under the same reaction conditions. A method for evaluating the enantiomeric excesses of the optically active products is also described. © 1995 Wiley-Liss, Inc.     

Recombination and Chromosome Segregation during the Single Division Meiosis in SPO12–1 and SPO13–1 Diploids

This paper reports a study of chromosome segregation and recombination during sporulation of spo12–1 and spo13–1 diploid strains of S. cerevisiae. These strains undergo a single division to form asci containing two diploid or near-diploid spores. The segregation of centromere-linked markers in the two-spored (dyad) products indicates that the division is generally equational. However, in a small percentage of the spo12–1 and spo13–1 cells, it appears that a meiosis I-like division occurs. Aberrant segregation of the MAT locus on chromosome III, yielding a monosomic and a trisomic spore pair, occurs in 12% of all dyads. The segregation patterns of markers at various distances from their centromeres and several pairs of markers on the same chromosome indicate that recombination takes place in both strains at nearly standard meiotic levels.     

Probing protein structure by solvent perturbation of nuclear magnetic resonance spectra. Nuclear magnetic resonance spectral editing and topological mapping in proteins by paramagnetic relaxation filtering.

Soluble spin labels, which "bleach" the surface proton resonances of a protein to n.m.r. measurements, can provide useful information about protein conformation and dynamics. The use of the soluble nitroxide, TEMPOL, has been explored to show the correlation of the paramagnetic perturbations of protein two-dimensional n.m.r. data with proton exposure to the free radical in hen egg-white lysozyme. The results demonstrate that the nitroxide approaches the protein randomly, and that the extent of the observed paramagnetic effects reflects the native folding pattern of the protein. A correlation of spectral simplification with the known tertiary structure establishes the feasibility of new strategies for topological mapping of surface and buried protons of the protein. Application to the elucidation of protein structure and to the study of dynamical processes is discussed.