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Liposomes have been of immense interest from the aspect of both fundamental and applied sciences. While their membrane mimicking properties enable them as excellent models to explore and understand the biophysics of cell membranes, they have also found extensive applicability in the fields of drug transport, DNA transfection and antimicrobial activity. The phase behaviour and dynamical features in the different phases of various biomembrane mimetic lipids/membrane systems are of immense interest. It is known that basic nature of the dynamical processes in these model membrane systems is very similar to that in the actual cell membrane. Dynamics of membrane is a prime determinant in a number of physiological processes such as cell signaling, permeability, cell fusion, etc. It is also possible to understand the interaction of nonsteroidal anti-inflammatory drugs (NSAIDs) and ionic liquids with lipid bilayers from the changes observed in their dynamical parameters. Combining neutron scattering and MD simulation techniques, we have successfully described the dynamical behaviour in various bio-mimetic membrane systems. Few results highlighting the effect of different NSAIDs and ionic liquids on the dynamics of the lipid membrane are also discussed. [ABSTRACT FROM AUTHOR]

Basal stem rot (BSR) disease is a devastating threat to Malaysia oil palm plantation, which is caused by Ganoderma boninense fungi. Dikaryon is the dominant stage of G. boninense and the pathogenic form of the fungus as compared to the monokaryon. Mating signalling response lead by mitogen-activated pathway (MAPK) responsible in the transformation of sterile monokaryon to pathogenic dikaryon. Expression of the STE3 gene involved in the autocrine activation of mating pathway. Mutations of STE3 gene in some pathogenic yeast resulted in inhibition of pheromone responses and blocked the signalling in the cell fusion. Identification of pheromone receptor, STE3 gene involved in mating signalling pathway is important to understand the mating response towards pathogenesis of G. boninense. Genome walking and polymerase chain reaction (PCR) were carried out to amplify isolate the full length of STE3 gene. Several in silico sequence analysis were conducted to characterise the STE3 gene. The full-length gene sequence including the upstream and downstream regions obtained could be used in future studies to understand pathogenesis of G. boninense. [ABSTRACT FROM AUTHOR]

Cell fusion is an essential eukaryotic process that occurs during mammalian development, including fertilization and muscle development. Despite the importance, the molecular mechanisms of fusion processes are still under active investigation. Saccharomyces cerevisiae, or budding yeast, provides an excellent model to study fusion, as these eukaryotic cells fuse during mating. Fusion begins when cells of the opposite mating type polarize their growth towards each other, creating a flat interface between the cells where fusion occurs. At this step, the cell walls between the mating pair must degrade before plasma membrane fusion and cytoplasmic mixing can occur. This step involves many key proteins that interact with each other to ensure successful cell wall degradation. Fus1 is a transmembrane protein hypothesized to act as a scaffold to bring other fusion regulators to the zone of cell fusion. However, its specific protein-protein interactions and function have not been uncovered. Fus1 contains a cytoplasmic, C-terminal SH3 domain, a highly conserved domain found in other proteins where it facilitates protein-protein interactions. To determine the function of this domain in Fus1, we analyzed a point mutation where a conserved histidine residue was mutated to alanine, H469A. Through microscopy-based assays, we found that this mutation causes a significant decrease in fusion. The Fus1-H469A protein has a decrease in localization to the fusion zone. We hypothesized that these phenotypes resulted from a block in protein-protein interactions, and investigated the genetic interaction between Fus1 and Fus2, another fusion regulator that localizes to the fusion zone. We found a decrease in co-localization of Fus2 and Fus1-H469A compared to the wildtype proteins and determined that the deletion of FUS2 had a smaller effect on Fus1-H469A than wildtype. We propose that the H469A mutation alters Fusl's ability to interact with Fus2, meaning the SH3 domain serves a key role for fusion. [ABSTRACT FROM AUTHOR]

Garlic (Allium sativum) is an edible plant used as food and spice. In popular medicine it is considered, along with the onions, a real remedy due to the rich content of vitamins and minerals. Pentimethalin is the active component of the herbicide Stomp 330 EC; this is frequently used by the farmers to control weeds in the garlic cultures. Cytogenetic effects of Pentimethalin have been tested on the root tip cells of Allium sativum L., with a series of concentrations (1.5; 2 and 3 ppm), for 6, 12 and 24 hours. The control has been untreated. Garlic bulbs have been grown in water at room temperature (20 ± 2 °C); when the newly emerged roots are 2 cm in length, they are used in the test. Microscopic slides have been prepared by using Feulgen-Rossenbeck method for coloring the chromosomes and squash technique. The measurements were made under an optic microscope (two models have been used: MBL-2000 Kruss and Celestron 44340 LCD Digital) by considering 15 microscopically areas per slide. Two parameters were studied: the mitotic index and chromosomal abnormalities. The number of abnormal cells was counted in each phase of mitosis. The analysis of variance was used to assess the significant differences between control and each treatment. The results were analyzed using statistical program for Windows. The effects of increasing concentrations of Pentimethalin were: decrease in mitotic index and induction of a high level of chromosomal abnormalities. Thus, the mitotic index decreased with increasing the herbicide concentration at each exposure time; on the other hand, various chromosomal abnormalities like rings, fragments, bridges, stickiness, laggards and micronucleus were observed. From this point of view, the bridges were generally observed in all treatment alternatives. The frequencies of chromosomal abnormalities increase with increasing the herbicide concentration. In this regard, the differences among the concentrations have been significant, when compared with untreated control. Our study reveals a direct correlation between herbicide concentration, exposure time and mutagenic effects observed in exposed Allium sativum root cells. Cytologic effects of Pentimethalin to garlic, suggest prudence regarding abusive usage of this substance. However, supplementary studies are necessary to determine the molecular mechanisms whereby Pentimethalin induce genotoxic effects to plants. Besides, there are still many chemical substances for plant protection whose possible genotoxic effects are unknown. [ABSTRACT FROM AUTHOR]

We employed femtosecond (fs) laser pulses to solve important biomedical and biotechnology problems, embryo biopsy and cell fusion respectively. We report on the results of fully contactless laser-mediated polar body (PB) and trophectoderm (TE) biopsy of early mammalian embryos. In the former case the fs laser scalpel (Cr:Forsterite seed oscillator and a regenerative amplifier, 100 fs, 10 Hz) was initially used to drill an opening in the outer covering of the embryo, and then the PB was extracted out of the zygote by means of optical tweezers (cw fiber laser, 1064 nm). In the latter case the laser scalpel was employed to dissect 5-7 TE cells that had just left the zona pellucida (ZP) during the hatching. The energy of laser pulses was thoroughly optimized to prevent cell damage and provide high viability of treated cells. Morphological and fluorescent analysis showed that femtosecond laser-based embryo biopsy did not compromise further in vitro embryo development. We also demonstrated the possibilities of using fs laser pulses for cell fusion. The contact border of the blastomeres of two-cell mouse embryos was perforated by a single fs-laser pulse with energy of 30-50 nJ. The blastomeres fusion process was usually complete within ∼60 min. In conclusion, the proposed techniques of laser microprocessing of living cells enable accurate, contamination-free, simple and quick performance of the procedures, and thus show a great potential for using fs lasers as a microsurgical tool. [ABSTRACT FROM AUTHOR]

Pheochromocytoma is a tumor of the adrenal gland which originates from chromaffin cells and is characterized by the secretion of excessive amounts of neurotransmitter which lead to high blood pressure and palpitations. Pheochromocytoma contain membrane bound granules that store neurotransmitter. The release of these stored molecules into the extracellular space occurs by fusion of the granule membrane with the cell plasma membrane, a process called exocytosis. The molecular mechanism of this membrane fusion is not well understood. It is proposed that the so called SNARE proteins [1] are the pillar of vesicle fusion as their cleavage by clostridial toxin notably, Botulinum neurotoxin and Tetanus toxin abrogate the secretion of neurotransmitter [2]. Here, I describe how physical principles are applied to a biological cell to explore the role of the vesicle SNARE protein synaptobrevin-2 in easing granule fusion. The data presented here suggest a paradigm according to which the movement of the C-terminal of synaptobrevin-2 disrupts the lipid bilayer to form a fusion pore through which molecules can exit. [ABSTRACT FROM AUTHOR]

Cell fusion is an important process in a wide range of living organisms. However, limited information is known about this mechanical process. One mechanism that involves cell fusion is mating of the yeast Saccharomyces cerevisiae by having two haploid yeast cells fuse to become one diploid cell. The cell walls of the two opposing yeast cells come into contact and slowly start degrading at the specific contact area so that the two plasma membranes can fuse. If the cell wall starts to degrade in an improper area, it can cause significant harm to the cells, but without any degradation, the cells will never fuse. Cell fusion is therefore regulated by genes that promote fusion properly to avoid causing harm to the cell. One gene that has a direct impact on yeast cell fusion is FUS1. Fus1 is a transmembrane protein that regulates cell fusion by degrading the cell walls between mating pairs. Fus1 is thought to localize mating-specific vesicles that contain hydrolases to break down the cell wall. Thus, a complete deletion of FUS1 blocks fusion before cell wall degradation. There are two known protein domains in Fus1, a transmembrane domain and a SH3 domain. SH3 domains are known to mediate protein-protein interactions, however how this domain specifically functions in Fus1 is unknown. I therefore created a single point mutation using site-directed mutagenesis in a highly conserved residue of the SH3 domain in Fus1. This single point mutation changes residue 469, a Histidine, into an Alanine. Using a Quantitative mating assay, I found that fusion in cells containing fus1-H469A was significantly blocked, shown by a lack of plasma membrane fusion between mating pairs. Preliminary data visualizing Fus1 protein localization in mating-induced haploid cells hints that the H469A mutation may affect the efficiency and location of Fus1 during fusion. [ABSTRACT FROM AUTHOR]

Myoblast migration represents a key step in muscle regeneration following microdamage after exercise (1). Myosatellite cell fusion with the damaged myofibre and subsequent differentiation pilots muscle remodelling (2). While increased production of Reactive Oxygen Species (ROS) is crucial in exercise-induced muscle damage (3). In line with this, antioxidant supplementation, such as the polyphenol resveratrol, may suppress skeletal muscle damage and improve anti-oxidant capacity (4). However, ROS is also crucial for exercise-mediated adaptation and repair (5). The way at which ROS and/or resveratrol modulate myoblast function in muscle repair remains unclear. Here, we explored the individual and combined effect of different concentrations of resveratrol (RS; 10-20-40-60 µM) and hydrogen peroxide (H2O2; 2-1 mM, 800-500-100-50-10µM) on non-differentiated (myoblasts) and differentiated (myotubes) mouse skeletal muscle-derived cells (C2C12). The impact of RS on myoblasts regenerative capacity was tested by assessing cell morphology, proliferation (MTS assay), migration (wound healing), sprouts formation (spheroids assay) and myotube formation and/or shrinking. In myoblasts, we tested the effect of RS plus H2O2 on cell migration. In 8-days differentiated myotubes we studied the impact of RS plus H2O2 on the oxidative capacity [succinate dehydrogenase (SDH) activity] and myosin ATPase activity and composition (total and type1-myosin). Data analysed by ANOVA in five replicates. The effect of RS on cell mitosis (24-48h) and proliferation (72h) was dose dependent; RS 10-20µM did not show significant effects, but at higher doses (40-60µM) it impaired both cell processes (p<0.01 ). RS 10-20µM enhanced (p=0.03) migration and the formation of myoblast sprouts, but not cell fusion. RS 40-60µM almost blocked myoblast regenerative capacity. In parallel, low doses of H2O2(10-500µM) enhanced motility with the highest efficiency at 100µM, while higher doses showed damaging effects. Notably, 24h of RS pre-conditioning (10-20 µM) prevented the deleterious effects of H2O2(1mM) and further enhanced cell migration induced by H2O2100µM. Evidence suggests that RS may increase fibre oxidative metabolism (4). Surprisingly, we did not find any effect of RS on myotube oxidative capacity (SDH), while we found a reduction (p<0.01) in myosin type1-ATPase activity and increased total myosin ATPase content. Finally, H2O2 (1mM) induced a reduction (p<0.01) in total and myosin type1 ATPase activity, that was prevented by RS in a dose-dependent manner. In conclusion, our data support the notion that low concentrations of ROS enhance myoblast regenerative capacity while it is impaired at high concentrations. High doses of anti-oxidants may also impair regenerative capacity, while low doses may abolish the detrimental impact of high concentrations of ROS on muscle regeneration. [ABSTRACT FROM AUTHOR]