Hanna Câmara da Justa , Antonielle Beatriz Baldissera , Mariana Izabele Machado , Samira Hajjar Souza , Nayanne Louise Costacurta Polli , Marianna Boia-Ferreira , Pedro Henrique de Caires Schluga , Lucelia Donatti , Ana Carolina M. Wille , João Carlos Minozzo , Luiza Helena Gremski , Silvio S. Veiga
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引用次数: 0
Abstract
Members of the phospholipase D (PLD) superfamily found in Loxosceles spider venoms are potent toxins with inflammatory and necrotizing activities. They degrade phospholipids in cell membranes, generating bioactive molecules that activate skin cells. These skin cells, in turn, activate leukocytes involved in dermonecrosis, characterized by aseptic coagulative necrosis. Although the literature has advanced in understanding the structure-function relationship, the cell biology resulting from the interactions of these molecules with cells remains poorly understood. In this study, we show that different cells exposed to recombinant PLDs bind these molecules to their plasma membrane, leading to the subsequent organization of extracellular microvesicles/ectosomes. The binding occurs as quickly as five minutes or less after exposure, increases over time, and eventually, the PLDs are expelled from the cell surface without generating cytotoxicity. PLDs are not endocytosed, nor do they spatially colocalize with acidic organelles in the intracellular environment. At least two regions of PLDs – the domain involved in magnesium ion coordination and the choline binding site – appear to play a role in cell surface binding and ectosome organization. However, the amino acids involved in catalysis do not participate in these events. The binding of these PLDs to the cell membrane, independent of catalytic activity, is sufficient to trigger intracellular signaling and enhance the expression of the pro-inflammatory IL-8 gene. These results are supported by the observation that isoforms of PLDs lacking catalytic activity induce an inflammatory response in vivo when injected into the skin of rabbits, without causing dermonecrosis. Our data indicate that these PLDs bind to the surface of target cells, promoting the organization of extracellular vesicles/ectosomes. Subsequently, these events activate pro-inflammatory genes and induce an inflammatory response in vivo. The binding to cells is not dependent on amino acids involved in catalysis but rather on amino acids involved in magnesium coordination. The binding of PLDs to the cell surface, formation of ectosomes, and activation of cells appear to initiate signals involved in inflammatory responses that can lead to dermonecrosis in accidents. This correlation is supported by experimental observations indicating that the events of toxin binding to cells, formation of microvesicles, and inflammatory responses observed both in vitro and in vivo are interconnected.
期刊介绍:
BBA Molecular and Cell Biology of Lipids publishes papers on original research dealing with novel aspects of molecular genetics related to the lipidome, the biosynthesis of lipids, the role of lipids in cells and whole organisms, the regulation of lipid metabolism and function, and lipidomics in all organisms. Manuscripts should significantly advance the understanding of the molecular mechanisms underlying biological processes in which lipids are involved. Papers detailing novel methodology must report significant biochemical, molecular, or functional insight in the area of lipids.