Vesiculation of cellular membranes is a constitutive process, although it can be induced by external physico-chemical parameters. In this study we explored the blebbing and vesiculation of Madin-Darby canine kidney (MDCK) cells induced by fluorescently-tagged aegerolysin, OlyA-mCherry. The paper shows that vesiculation of MDCK cells induced by OlyA-mCherry is not decreased by low temperatures and does not depend on increased [Ca2+] ions. Our proteomic analysis revealed that these shed extracellular vesicles (EVs) contained 71 proteins. These proteins were mostly of cytosolic and nuclear origin, and only a few were assigned as membrane-associated or typical membrane raft residents. Furthermore, the cellular and EVs lipidome were characterized using liquid chromatography-mass spectrometry and high-resolution nuclear magnetic resonance, which included analysis of the phospholipids and ceramide-based glycosphingolipids. We identified 218 different lipid species in these cells, and 84 different lipid species in EVs. We concluded that shed EVs are potentially interesting model for biophysical research as well as tool for non-invasive sampling of cytosolic proteins from cells and thus metabolic fingerprinting.
COBISS.SI-ID: 3971151
The aim of this study was to investigate the differences in lipid profiles associated with different levels of urothelial cancer cell invasiveness. Comparative lipidomic studies were performed on the RT4 versus T24 urothelial cancer cell lines as models for noninvasive papillary urothelial neoplasm cells and invasive urothelial neoplasm cells. Our results demonstrate that the two types of cells, RT4 and T24 cells, express some significant differences of cellular membranes including changes in lipid composition. The differences between RT4 and T24 cells suggest significantly different organization of the cellular membranes, which can affect the membrane fluidity and membrane-dependent functions, and contribute to the lower stiffness of plasma membrane and reduced cell–cell adhesion required for movement and invasiveness of these T24 urothelial carcinoma cells with a high metastatic potential. The differences in lipid composition between both cell lines were additionally confirmed using membrane raft-sensing fluorescently-labeled aegerolysin, OlyA-mCherry that was recently developed in our laboratories.
COBISS.SI-ID: 3968591
The fungal genus Aspergillus is of critical importance to humankind. This widespread genus includes beneficial species with industrial applications, such as production of metabolites and enzymes, and a genetic model that has provided fundamental discoveries in biology. The genus also includes pathogenic species capable of causing major human and animal disease, and species which can cause devastating crop yield loss, post-harvest spoilage and contamination with potent carcinogenic toxins. The genome sequences of eight aspergilli have already been explored to investigate aspects of fungal biology, raising questions about evolution and specialization within this genus. In this research-community-driven study, we have generated genome sequences for ten novel, highly diverse Aspergillus species and compared these in detail to sister and more distant genera. Comparative studies of key aspects of fungal biology including primary and secondary metabolism, stress response, biomass degradation, and signal transduction revealed both conservation and diversity among the species. Comparative genomics was complemented with experimental studies to validate differences observed between the genomes. These revealed several highlights, such as the potential for sex in asexual species, organic acid production genes being a key feature of black aspergilli, alternative approaches for degrading plant biomass, and indications for the genetic basis of stress response. A genome-wide phylogenetic analysis demonstrated in detail the relationship of the newly genome sequenced species with other aspergilli. Many aspects of biological differences between fungal species cannot be explained by current knowledge obtained from genome sequences. The comparative genomics and experimental study presented here, allows for the first time a genus-wide view of the biological diversity of the aspergilli, and in many, but not all, cases linked genome differences to phenotype. These data provide many leads for functional studies into the evolution and biotope adaptation of individual species and furnish a reference, not only for the ongoing whole genus genome sequencing project of Aspergillus, but also for comparative studies in and between other fungal genera. Furthermore, insights gained can be exploited for biotechnological and medical applications of fungi, such as the improvement of production of enzymes and metabolites, or the development of novel anti-fungal strategies.
COBISS.SI-ID: 4239951
This review summarizes the so far obtained knowledge on distribution, structure and possible applications of proteins belonging to the aegerolysin family.
COBISS.SI-ID: 4324943
Understanding of the interactions between proteins and natural and artificially prepared lipid membrane surfaces and embedded nonpolar cores is important in studies of physiological processes and their pathologies and is applicable to nanotechnologies. In particular, rapidly growing interest in cellular droplets defines the need for simplified biomimetic lipid model systems to overcome in vivo complexity and variability. We present a protocol for the preparation of kinetically stable nanoemulsions with nanodroplets composed of sphingomyelin (SM) and cholesterol (Chol), as amphiphilic surfactants, and trioleoylglycerol (TOG), at various molar ratios. To prepare stable SM/Chol-coated monodisperse lipid nanodroplets, we modified a reverse phase evaporation method and combined it with ultrasonication. Lipid composition, ?-potential, gyration and hydrodynamic radius, shape, and temporal stability of the lipid nanodroplets were characterized and compared to extruded SM/Chol large unilamellar vesicles. Lipid nanodroplets and large unilamellar vesicles with theoretical SM/Chol/TOG molar ratios of 1/1/4.7 and 4/1/11.7 were further investigated for the orientational order of their interfacial water molecules using a second harmonic scattering technique, and for interactions with the SM-binding and Chol-binding pore-forming toxins equinatoxin II and perfringolysin O, respectively. The surface characteristics (?-potential, orientational order of interfacial water molecules) and binding of these proteins to the nanodroplet SM/Chol monolayers were similar to those for the SM/Chol bilayers of the large unilamellar vesicles and SM/Chol Langmuir monolayers, in terms of their surface structures. We propose that such SM/Chol/TOG nanoparticles with the required lipid compositions can serve as experimental models for monolayer membrane to provide a system that imitates the natural lipid droplets.
COBISS.SI-ID: 4754255