Osmotic tolerance of avian erythrocytes to complete hemolysis in solute free water.

Osmotic behavior of erythrocytes is not only important clinically, but is also significant in understanding of material transport across biological membranes. It is most commonly studied through fragiligrams - plots of the degree of hemolysis as a function of extracellular osmolarity. A fundamental assumption in experimental and theoretical studies on osmolarity driven transport of water across the plasma membranes of all cells is the sigmoidal nature of their osmotic behavior. Sigmoidal data is mathematically monotonic showing either a decreasing only or an increasing only trend, but not both, within certain thresholds; beyond these thresholds the data is asymptotic or flat. Fragiligrams of erythrocytes are usually sigmoidal, with maximal hemolysis in plain solute-free water and often up to a certain extracellular hypotonic environment. In this work, we report a new discovery of non-monotonic osmotic behavior of avian erythrocytes. In contrast to the expected monotonic fragiligrams obtained for mammalian erythrocytes, fragiligrams of avian erythrocytes show non-monotonic curves. Maximal hemolysis of avian erythrocytes was not observed at the most hypotonic conditions - instead, maximal hemolysis was observed at mild hypotonic conditions. Hemolysis of avian erythrocytes first increases then decreases with increasing extracellular osmolarity. We also report that the non-monotonic fragiligrams of chicken erythrocytes are converted to the expected monotonic sigmoids subsequent to controlled extracellular trypsinization. While possibly having profound evolutionary implications for vertebrates, the findings reported in this work have a direct impact on understanding of avian physiology. Our results also compel revisiting of experimental and theoretical models for understanding material transport across biological membranes under different osmotic conditions.

Cell-penetrating and cargo-delivery ability of a spider toxin-derived peptide in mammalian cells.

Cell-penetrating peptides are short cationic peptides with inherent ability to cross the plasma membrane barrier as well as intracellularly deliver cargo molecules conjugated to them. Venoms from snakes, scorpions and spiders are rich in membrane-active peptides. Crotamine from snake venom as well as maurocalcine and imperatoxin isolated from scorpion venoms have been reported to possess cell-penetrating property in mammalian cells. Latarcins, a group of spider venom toxins, has also been reported to possess antimicrobial property. However, cell-penetrating ability of Latarcins is still not elucidated. This is the first report where cell-penetrating ability of a peptide derived from spider toxin, Latarcin 1 has been demonstrated. Interestingly, the structurally minimized sequence of Latarcin 1 (LDP - Latarcin-derived peptide) when conjugated with nuclear localization sequence from Simian Virus T40 antigen (LDP-NLS) translocates across cell membrane in HeLa cells. The chimeric LDP-NLS peptide also did not exhibit cytotoxicity towards mammalian cells in contrast to the LDP that showed lesser uptake and higher cytotoxicity. LDP-NLS also successfully delivered macromolecular protein cargo inside the cells.

CyLoP-1: Membrane-active peptide with cell-penetrating and antimicrobial properties.

CyLoP-1 is a cysteine-rich cell-penetrating peptide derived from nuclear localization sequence of snake toxin, crotamine. The peptide has shown cytoplasmic uptake in mammalian cells at lower concentrations. In the present study, the cell-penetrating and antimicrobial activity of the peptide has been studied by employing mammalian cells, plant cells as well as bacterial and fungal pathogens. The study shows that the peptide acts as an effective CPP and a cargo-delivery vector for not only mammalian cells but also for plant cells. Besides this, the peptide also possesses antimicrobial activity against representative pathogens tested. It is shown to be effective in killing methicillin-resistant Staphylococcus aureus. We have observed that the presence of cysteine residues in the peptide play a major role in conferring cell-penetrating as well as antimicrobial activity to the peptide since there is a significant decline in these activities when cysteine residues are replaced with serine residues. Our findings are significant for the proposition that CyLoP-1 is an efficient membrane-active peptide with both cell-penetrating and antimicrobial activity. Hence, it can be further evaluated for its application in the field of drug-delivery, plant biotechnology and as a peptide-antibiotic.

Membrane-active peptides from marine organisms--antimicrobials, cell-penetrating peptides and peptide toxins: applications and prospects.

Marine organisms are known to be a rich and unique source of bioactive compounds as they are exposed to extreme conditions in the oceans. The present study is an attempt to briefly describe some of the important membrane-active peptides (MAPs) such as antimicrobial peptides (AMPs), cell-penetrating peptides (CPPs) and peptide toxins from marine organisms. Since both AMPs and CPPs play a role in membrane perturbation and exhibit interchangeable role, they can speculatively fall under the broad umbrella of MAPs. The study focuses on the structural and functional characteristics of different classes of marine MAPs. Further, AMPs are considered as a potential remedy to antibiotic resistance acquired by several pathogens. Peptides from marine organisms show novel post-translational modifications such as cysteine knots, halogenation and histidino-alanine bridge that enable these peptides to withstand harsh marine environmental conditions. These unusual modifications of AMPs from marine organisms are expected to increase their half-life in living systems, contributing to their increased bioavailability and stability when administered as drug in in vivo systems. Apart from AMPs, marine toxins with membrane-perturbing properties could be essentially investigated for their cytotoxic effect on various pathogens and their cell-penetrating activity across various mammalian cells. The current review will help in identifying the MAPs from marine organisms with crucial post-translational modifications that can be used as template for designing novel therapeutic agents and drug-delivery vehicles for treatment of human diseases.