Enhancing Antibacterial Activity Against Escherichia coli K-12 of Peptide Ib-AMP4 with Synthetic Analogues

A family of Ib-AMP4 peptide analogues was obtained by solid phase synthesis, modifying the net charge and hydrophobicity of C-terminal domain by replacing certain amino acidic residues by arginine and tryptophan. Additionally, disulfide bonds were eliminated by replacing the cysteine residues by methionine, which resulted in a decrease in the number of synthesis byproducts, and consequently diminished the subsequent purification steps. The obtained peptides were purified by RP-HPLC and their molecular mass was determined by MALDI-TOF mass spectrometry. The peptide analogues (IC₅₀ between 1 and 50 μM) presented a higher antibacterial activity against Escherichia coli K-12 than the native peptide (IC₅₀ > 100 μM). The hemolytic activity of the peptide with the highest antibacterial efficacy presented no degradation of erythrocytes for a concentration of 1 μM that corresponds to its IC₅₀ value. The results show that the synthesized peptides are good candidates for the treatment of diseases caused by E. coli.     

Schizosaccharomyces pombe encodes a mutated AP endonuclease 1

Mutagenic and cytotoxic apurinic/apyrimidinic (AP) sites are among the most frequent lesions in DNA. Repair of AP sites is initiated by AP endonucleases and most organisms possess two or more of these enzymes. Saccharomyces cerevisiae has AP endonuclease 1 (Apn1) as the major enzymatic activity with AP endonuclease 2 (Apn2) being an important backup. Schizosaccharomyces pombe also encodes two potential AP endonucleases, and Apn2 has been found to be the main repair activity, while Apn1 has no, or only a limited role in AP site repair. Here we have identified a new 5' exon (exon 1) in the apn1 gene and show that the inactivity of S. pombe Apn1 is due to a nonsense mutation in the fifth codon of this new exon. Reversion of this mutation restored the AP endonuclease activity of S. pombe Apn1. Interestingly, the apn1 nonsense mutation was only found in laboratory strains derived from L972 h(-) and not in unrelated isolates of S. pombe. Since all S. pombe laboratory strains originate from L972 h(-), it appears that all experiments involving S. pombe have been conducted in an apn1(-) mutant strain with a corresponding DNA repair deficiency. These observations have implications both for future research in S. pombe and for the interpretation of previously conducted epistatis analysis.     

Calcium signal transmission between ryanodine receptors and mitochondria

Control of energy metabolism by increases of mitochondrial matrix [Ca(2+)] ([Ca(2+)](m)) may represent a fundamental mechanism to meet the ATP demand imposed by heart contractions, but the machinery underlying propagation of [Ca(2+)] signals from ryanodine receptor Ca(2+) release channels (RyR) to the mitochondria remains elusive. Using permeabilized cardiac (H9c2) cells we investigated the cytosolic [Ca(2+)] ([Ca(2+)](c)) and [Ca(2+)](m) signals elicited by activation of RyR. Caffeine, Ca(2+), and ryanodine evoked [Ca(2+)](c) spikes that often appeared as frequency-modulated [Ca(2+)](c) oscillations in these permeabilized cells. Rapid increases in [Ca(2+)](m) and activation of the Ca(2+)-sensitive mitochondrial dehydrogenases were synchronized to the rising phase of the [Ca(2+)](c) spikes. The RyR-mediated elevations of global [Ca(2+)](c) were in the submicromolar range, but the rate of [Ca(2+)](m) increases was as large as it was in the presence of 30 microm global [Ca(2+)](c). Furthermore, RyR-dependent increases of [Ca(2+)](m) were relatively insensitive to buffering of [Ca(2+)](c) by EGTA. Therefore, RyR-driven rises of [Ca(2+)](m) appear to result from large and rapid increases of perimitochondrial [Ca(2+)]. The falling phase of [Ca(2+)](c) spikes was followed by a rapid decay of [Ca(2+)](m). CGP37157 slowed down relaxation of [Ca(2+)](m) spikes, whereas cyclosporin A had no effect, suggesting that activation of the mitochondrial Ca(2+) exchangers accounts for rapid reversal of the [Ca(2+)](m) response with little contribution from the permeability transition pore. Thus, rapid activation of Ca(2+) uptake sites and Ca(2+) exchangers evoked by RyR-mediated local [Ca(2+)](c) signals allow mitochondria to respond rapidly to single [Ca(2+)](c) spikes in cardiac cells.     

Fragment quality and matrix affect epiphytic performance in a Mediterranean forest landscape

Destruction and fragmentation of habitats represent one of the most important threats for biodiversity. Here, we examined the effects of fragmentation in Mediterranean forests on the epiphytic lichen Lobaria pulmonaria (Lobariaceae). We tested the hypothesis that not only the level of fragmentation affects L. pulmonaria populations, but also the quality of the habitat and the nature of the surrounding matrix affect them. The presence and abundance of the lichen was recorded on 2039 trees in a total of 31 stands. We recorded habitat quality and landscape variables at three hierarchical levels: tree, plot, and patch. We found that L. pulmonaria tends to occur in trees with larger diameters in two types of surveyed forests. In Quercus pyrenaica patches, the mean diameter of colonized trees was smaller, suggesting the importance of bark roughness. Factors affecting the presence and cover of the lichen in each type of forest were different. There was a strong positive influence of distance from a river in beech forests, whereas proximity to forest edge positively affected in oak forests. The influence of the surrounding matrix was also an important factor explaining the epiphytic lichen abundance.     

Asymmetric cell division and neoplastic growth

Asymmetric division was formed as an evolutionary conserved mechanism of self-maintenance of cellular populations and creation of a variety of cell types during ontogenesis. Asymmetric division enables a special mechanism of determinant segregation, which further defines development of daughter cells. As a result two unequal cells are developed. Recent research demonstrates the interplay of disturbed asymmetric division of stem cells and tumorigenesis. Genes implicated in cell’s polarization and normal progression of asymmetric mitosis were identified in Drosophila. Other genes regulating asymmetric mitosis were described as tumor suppressors, and their mutations were shown to initiate neoplastic growth. Comparative study of gene expression suggests that the disturbance of asymmetric division might be one of the reasons for neoplasm progression in vertebrates.     

Mechanosensitive Closed-Closed Transitions in Large Membrane Proteins: Osmoprotection and Tension Damping

Multiconformation membrane proteins are mechanosensitive (MS) if their conformations displace different bilayer areas. Might MS closed-closed transitions serve as tension buffers, that is, as membrane “spandex”? While bilayer expansion is effectively instantaneous, transitions of bilayer-embedded MS proteins are stochastic (thermally activated) so spandex kinetics would be critical. Here we model generic two-state (contracted/expanded) stochastic spandexes inspired by known bacterial osmovalves (MscL, MscS) then suggest experimental approaches to test for spandex-like behaviors in these proteins. Modeling shows: 1), spandex kinetics depend on the transition state location along an area reaction coordinate; 2), increasing membrane concentration of a spandex right-shifts its midpoint (= tension-Boltzmann); 3), spandexes with midpoints below the activating tension of an osmovalve could optimize osmovalve deployment (required: large midpoint, barrier near the expanded state); 4), spandexes could damp bilayer tension excursions (required: midpoint at target tension, and for speed, barrier halfway between the contracted and expanded states; the larger the spandex Δ-area, the more precise the maintenance of target tension; higher spandex concentrations damp larger amplitude strain fluctuations). One spandex species could not excel as both first line of defense for osmovalve partners and tension damper. Possible interactions among MS closed-closed and closed-open transitions are discussed for MscS- and MscL-like proteins.