Imaging plasma membrane domains in signal transduction pathways

The cell membrane acts as a barrier that controls the passage of substances from the outside to the inside of a cell. It is composed of various lipids organized in a bilayer with proteins embedded. Experimental data suggested that lipids are organized in nanometer-sized structures called membrane domains. I study the existence and the role of domains in living cells through single-molecule fluorescence microscopy. This technique allows pinpointing the position of each molecule with high spatial accuracy. I apply it to study the distribution of a membrane-anchored protein, HRas, in the inner leaflet of the membrane. From the single-molecule positions a map of protein distribution is reconstructed. Statistical analysis revealed dynamic partitioning in membrane domains. A different approach relies on tracking single proteins diffusion in the membrane. With this method I studied the influence of domains in the assembly of a two-component receptor, type I interferon receptor. I observed confinements of the components in small domains, which makes assembly faster and more efficient. Further, I present an advanced technique, to track proteins at microsecond time scale. After validating the technique on DNA, I applied it to GPI-anchor protein diffusion. These data confirmed the existence of theoretically proposed, complex diffusive modes.LEI Universiteit LeidenStichting voor Fundamenteel Onderzoek der Materie (FOM), die financieel wordt gesteund door de Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO)Biological and Molecular Physic

Zebrafish xenografts as a fast screening platform for bevacizumab cancer therapy

Cancer is the second leading cause of death in the world. Given that cancer is a highly individualized disease, predicting the best chemotherapeutic treatment for individual patients can be difficult. Ex vivo models such as mouse patient-derived xenografts (PDX) and organoids are being developed to predict patient-specific chemosensitivity profiles before treatment in the clinic. Although promising, these models have significant disadvantages including long growth times that introduce genetic and epigenetic changes to the tumor. The zebrafish xenograft assay is ideal for personalized medicine. Imaging of the small, transparent fry is unparalleled among vertebrate organisms. In addition, the speed (5-7 days) and small patient tissue requirements (100-200 cells per animal) are unique features of the zebrafish xenograft model that enable patient-specific chemosensitivity analyses.info:eu-repo/semantics/publishedVersio

Effect of selenium enrichment on metabolism of tomato (Solanum lycopersicum) fruit during post‐harvest ripening

BACKGROUND Selenium (Se) enrichment of plants seems effective in enhancing the health‐related properties of produce, and in delaying plant senescence and fruit ripening. The current study investigated the effects of Se on tomato fruit ripening. Tomato (Solanum lycopersicum L.) plants were grown in hydroponics with different Se‐enriched nutrient solutions. Se, as sodium selenate, was added at rate of 0 mg L‐1 (control), 1 mg L‐1, and 1.5 mg L‐1. RESULTS Selenium was absorbed by roots and translocated to leaves and fruit. Se enrichment did not significantly affect the qualitative parameters of fruit at commercial harvest, instead it delayed ripening by affecting specific ripening‐related processes (respiration, ethylene production, color evolution) during postharvest. In the current experiment 100 g of tomato hydroponically grown with a 1.5 mg Se L‐1 enriched solution provided a total of 23.7 μg Se. Selenium recommended daily intake is 60 μg for women and 70 μg for men, thus the daily consumption of 100 g of enriched tomato would not lead to Se toxicity, but would provide a good Se diet supplementation. CONCLUSIONS The cultivation of tomato plants in a Se‐enriched solution appeared effective in producing tomato fruit with improved performances during storage and postharvest shelf life, and also with greater potential health‐promoting properties

Influence of zinc and manganese enrichments on growth, biosorption and photosynthetic efficiency of Chlorella sp

Treating biosolids from industrial, urban, and agricultural plants produces high amounts of water. After organic pollutants and non-essential heavy metals have been removed, these wastewaters are still rich in trace elements such as zinc (Zn), copper, or manganese (Mn) and have high conductivity and extremely variable pH. In this study, an isolated Chlorella sp. strain was grown for 21 days in nutrient solutions enriched with known amounts of Zn or Mn to obtain concentrations three (4.0 mg L−1)- and six (1.0 mg L−1)-fold higher than the basal medium levels, respectively, and over the limits permitted in aquatic environments. The green alga exhibited high tolerance to Zn and Mn, with the maximum abatement of Zn (28–30%) and Mn (60–63.5%) after 14 and 7 days of culture, respectively. Mn stimulated the growth rate and biomass production of Chlorella, which showed the highest carbon levels just in the first week. In both treatments, the nitrogen and protein contents remarkably increased. The photosynthetic pigments increased until the 14th day, with a higher extent in the Zn-enriched solution. An increasing photochemical efficiency was observed after 7 days of treatment, when the microalgae grown in Zn- and Mn-enriched solutions showed a slightly higher maximum photochemical efficiency than control. The autotrophic and controlled growth system adopted was designed to monitor the dynamic balance of Zn and Mn contents in the solutions and in the algal biomass. This system has proved to be useful in identifying the optimal nutritional conditions of the microalgae, along with the optimal temporal patterns of both metal biosorption capacity for water remediation and element bioaccumulation in the algal biomass

PolNet Analysis: a software tool for the quantification of network-level endothelial cell polarity and blood flow during vascular remodelling

In this paper, we present PolNet, an open source software tool for the study of blood flow and cell-level biological activity during vessel morphogenesis. We provide an image acquisition, segmentation, and analysis protocol to quantify endothelial cell polarity in entire in vivo vascular networks. In combination, we use computational fluid dynamics to characterise the haemodynamics of the vascular networks under study. The tool enables, for the first time, network-level analysis of polarity and flow for individual endothelial cells. To date, PolNet has proven invaluable for the study of endothelial cell polarisation and migration during vascular patterning, as demonstrated by our recent papers (Franco 2015, Franco 2016a). Additionally, the tool can be easily extended to correlate blood flow with other experimental observations at the cellular/molecular level. We release the source code of our tool under the LGPL licence

PolNet:A Tool to Quantify Network-Level Cell Polarity and Blood Flow in Vascular Remodeling

In this article, we present PolNet, an open-source software tool for the study of blood flow and cell-level biological activity during vessel morphogenesis. We provide an image acquisition, segmentation, and analysis protocol to quantify endothelial cell polarity in entire in vivo vascular networks. In combination, we use computational fluid dynamics to characterize the hemodynamics of the vascular networks under study. The tool enables, to our knowledge for the first time, a network-level analysis of polarity and flow for individual endothelial cells. To date, PolNet has proven invaluable for the study of endothelial cell polarization and migration during vascular patterning, as demonstrated by two recent publications. Additionally, the tool can be easily extended to correlate blood flow with other experimental observations at the cellular/molecular level. We release the source code of our tool under the Lesser General Public License

Reconstructing hotspots of genetic diversity from glacial refugia and subsequent dispersal in Italian common toads (Bufo bufo)

Animal science

The distributions of the six species constituting the smooth newt species complex (Lissotriton vulgaris sensu lato and L. montandoni) – an addition to the New Atlas of Amphibians and Reptiles of Europe

The ‘smooth newt’, the taxon traditionally referred to as Lissotriton vulgaris, consists of multiple morphologically distinct taxa. Given the uncertainty concerning the validity and rank of these taxa, L. vulgaris sensu lato has often been treated as a single, polytypic species. A recent study, driven by genetic data, proposed to recognize five species, L. graecus, L. kosswigi, L. lantzi, L. schmidtleri and a more restricted L. vulgaris. The Carpathian newt L. montandoni was confirmed to be a closely related sister species. We propose to refer to this collective of six Lissotriton species as the smooth newt or Lissotriton vulgaris species complex. Guided by comprehensive genomic data from throughout the range of the smooth newt species complex we 1) delineate the distribution ranges, 2) provide a distribution database, and 3) produce distribution maps according to the format of the New Atlas of Amphibians and Reptiles of Europe, for the six constituent species. This allows us to 4) highlight regions where more research is needed to determine the position of contact zones

Imaging plasma membrane domains in signal transduction pathways

The cell membrane acts as a barrier that controls the passage of substances from the outside to the inside of a cell. It is composed of various lipids organized in a bilayer with proteins embedded. Experimental data suggested that lipids are organized in nanometer-sized structures called membrane domains. I study the existence and the role of domains in living cells through single-molecule fluorescence microscopy. This technique allows pinpointing the position of each molecule with high spatial accuracy. I apply it to study the distribution of a membrane-anchored protein, HRas, in the inner leaflet of the membrane. From the single-molecule positions a map of protein distribution is reconstructed. Statistical analysis revealed dynamic partitioning in membrane domains. A different approach relies on tracking single proteins diffusion in the membrane. With this method I studied the influence of domains in the assembly of a two-component receptor, type I interferon receptor. I observed confinements of the components in small domains, which makes assembly faster and more efficient. Further, I present an advanced technique, to track proteins at microsecond time scale. After validating the technique on DNA, I applied it to GPI-anchor protein diffusion. These data confirmed the existence of theoretically proposed, complex diffusive modes.</p