Biomimetic microscale platforms for the visualization of biological processes : from GUVs towards artificial cells

First in order to explore the possibilities of building a synthetic artificial cell following a biomimetic approach, we engineered synthetic polymer-based giant unilamellar vesicles (GUVs) with selective membrane permeability. Since the membranes of polymeric GUVs present a high impermeability compared to natural lipid membranes, membranes are selectively permeabilized in a biomimetic approach by the insertion of the small pore-forming peptide gramicidin (gA) as gA biopores are known to allow the transport of protons and monovalent ions. Whilst gA has been inserted in lipid membranes in numerous research studies, the challenge of inserting the bacterial pore into polymer membranes is greater because of the significant difference between the pore length and the thickness of the polymer membrane (more than 3.5 times). Confocal laser scanning microscopy (CLSM) was used to show that neither the size, nor the morphology of the GUVs was affected by successful insertion of gA and further to visualize the pH change inside the cavity of GUVs in real-time by recording videos. In order to demonstrate the successful insertion of gA, a pH-sensitive dye is encapsulated inside the cavity of GUVs and proton gradients between the environment of GUVs and their inner cavity serves to assess the exchange of protons across the membrane upon gA insertion. The results showed that gA was successfully inserted and remained functional in polymer membranes with thickness of 9.2–12.1 nm. Larger membrane thicknesses did not allow gA insertion, and 12.1 nm represents a limit for the mismatch between the pore length and the membrane thickness. Our gA-GUVs are therefore pH-regulating and maintain their integrity in different pH conditions in a cell-like manner. This bio-mimetic approach to use ion channels with specific selectivity for insertion in polymer membranes is an elegant strategy to develop mimics of biomembranes or for supporting the design of bioreactors. Next, a functional cell mimetic compartment is developed by the insertion of the bacterial membrane protein (OmpF) in thick synthetic polymer membranes of an artificial GUV compartment that encloses the oxidative enzyme horseradish peroxidase. In this manner a simple and robust cell mimic is designed, that supports a rudimental form of metabolism. The biopore serves as a gate, which allows substrates to enter the cavities of the GUVs, where they are converted into the resorufin-like products by the encapsulated enzyme, and then released in the environments of GUVs. Our bio-equipped GUVs facilitate the control of specific catalytic reactions in confined micro-scale spaces mimicking cell size and architecture and thus provide a straightforward approach serving to obtain deeper insights in the real-time of biological processes inside cells. This elegant strategy of equipping both GUV membranes and GUV cavities with biomolecules, opens the way towards cell-like compartments as novel materials with bio-functionality is the combination of synthetic micrometer-sized giant unilamellar vesicles (GUVs) with biomolecules because it enables studying the behavior of biomolecules and processes within confined cavities. Finally a visionary strategy for creating the first bioinspired molecular factory with functionality as a real cell-mimic based on micrometer-sized giant plasma membrane vesicles (GPMVs) is addressed. GPMVs are cell-derived giant vesicles consisting of an outer compartment architecture (membrane) and an inner composition, which both directly mirror the composition of cells from which they originate except the larger organelles like for example nuclei and Golgi apparatus making measurements easier and are the closest cell-mimic available on the market up to now. In a step towards the development of bioinspired molecular factories with functionality as cell mimics, we generate the next generation of cell mimics by the production of sophisticated hybrid molecular factories based on GPMVs, which are equipped with a synthetic molecular machinery inside their cavities that provides functionality. Such a hierarchical approach in compartmentalization allows the lower-level synthetic functional compartments encapsulated within the cavity of the GPMV to act as independent anatomically discreet units that specialize in their own function, making them nanoscale versions of nature own organelles. Towards the first bioinspired molecular factory enzyme-equipped polymersomes with a reconstituted membrane protein (OmpF) are encapsulated inside the GPMVs as enzymatic nanocompartment spaces, where they retain their structure and functionality. When substrates were added to the outer solution of the GPMVs, it was shown that they could penetrate both the membrane of the GPMVs and the inner compartment membranes of the synthetic nanoreactors equipped with OmpF pores. In this respect the equipment of the catalytic nanocompartment spaces with OmpF was essential as it allowed the enzyme to perform in the inner cavities. Successful substrate conversion was visualized by following the fluorescent product of the enzymatic reaction (resorufin-like product), which could leave the polymersome and diffuse inside the GPMV cavity. Finally we demonstrate that equipped GPMVs can act as artificial cell mimics – retaining their membrane and inner composition if they are injected into multicellular organisms – Zebrafish embryos. To the best of our knowledge, this is the first time that a molecular factory functioning as a cell-like mimic has been be constructed by using a top down-bottom up approach and has been tested in vivo by taking advantage of the fundamental nature of GPMVs

Cologne Of The Greats

Pages 39-4

Let Me Tell You

Page 3

Transnational traders: El Salvador’s women couriers in historical perspective

An estimated one-third to one-half of Salvadorans who carry remittances and goods between El Salvador and the United States are women. Scholars studying these viajeras argue that their work simultaneously represents a break from traditional gender relations confining women to the home and an extension of gender traits that favor women in developing social ties. Although social ties are crucial to the courier trade, this argument ignores antecedents to viajeras’ work in El Salvador and suggests that transnationalism pushes women into realms of labor and physical mobility that have been gendered masculine. Using ethnographic methods, I examine the relationship between women’s historical work in El Salvador and their current work as viajeras, as well the relationship between viajeras’ experiences and those of women transnational traders in other parts of the world. My findings contribute to a small but growing body of research suggesting that instead of merely being excluded from or manipulated by global processes, many women in the Global South have expanded the realm of their activities to help shape variable forms of global capitalism. Studying how they do so sheds light on local mechanisms for combating gender inequality and promoting development

Geophysical Investigations for Groundwater in a Complex Subsurface Terrain, Wadi Fatima, KSA: A Case History

Geological contacts in a complex subsurface terrain have favorable potentiality for groundwater; particularly in semiarid/or arid areas. Magnetic and DC resistivity surveys were proposed and conducted at Wadi Fatima for the groundwater exploration in this geologically complex media. Magnetic survey was used to delineate the basement structures which control the groundwater flow. On the other hand, DC resistivity profiling and sounding techniques were used to investigate and ensure the potentiality of the groundwater occurrence in the structurally complex basement. Magnetic data were subjected to analytic signal analysis, where it showed clearly the complexity of the study area. Many magnetic contacts were delineated at the maximum amplitudes of the analytic signal curves. Two selected profiles along the Wadi have been quantitatively interpreted using analytic signal, spectral analysis, horizontal and vertical derivatives. These analyses delineate the geological structures that may accommodate the groundwater. DC resistivity profiling technique was used along one profile, to define the effect of the basement structures on the ground water distribution under this profile. On the other hand, Vertical Electrical Sounding (VES) was conducted at selected sites along the magnetic and DC profiles. Ground magnetic and DC resistivity techniques led to a better understanding of the study area. Thus, the integration of the obtained results from these two methods was successful to define new sites for drilling wells for the groundwater supply that would be at VES9 and VES7. It was recommended to conduct more detailed studies in the southwestern part of the study area

Broadcasting in Saudi Arabia in the era of globalization : a study of local constraints on television development

This study examines the reasons for the Saudi media mdustry's dependence on imported foreign productions. In a departure from traditional dependency theory, which emphasises the role of external factors in the context of the world system, this study explains the state of dependency and underdevelopment in a more locally grounded analysis which evaluates the role of Saudi media policies and regulatory functions in perpetuatmg this dependency status. Two methodologies were applied, firstly, content analyses of a two-week period of Saudi television programming on Channel 1 were earned out to examine the quantity and quality of both local and imported television fare in terms of genre and format, Secondly, mterviews were conducted with Saudi media officials, media pohcy makers, and mdependent local producers to ascertain, from their perspective, what exactly constrains the Saudi media industry and limits its potential, and why the Saudi media is dependent on imported television fare. The results of the content analyses and interviews showed that political, professional and economic constraints handicap STV's performance. This has led to output which is considered to be irrelevant to the needs and mterests of the Saudi viewing population. It has also led to an increase in imported foreign programming and DBS populanty, thus creating a cause of concern among culturalists and Islamists who object to content which, they argue, conflicts with the basic principles of the Islamic faith. Recommendations are proposed to Saudi media policy makers in order to counteract the foreign competition and enhance mdigenous, self-reliant development

Increased heavy metal tolerance of cowpea plants by dual inoculation of an arbuscular mycorrhizal fungi and nitrogen-fixer Rhizobium bacterium

Through biological inoculation technology, the bacterial-mycorrhizal-legume tripartite symbiosis in artificially heavy metal polluted soil was documented and the effects of dual inoculation with arbuscular mycorrhizal (AM) fungus and Rhizobium (N-fixing bacteria, NFB) on the host plant cowpea (Vigna sinensis) in pot cultures were investigated at six concentrations of Zn (0.0 – 1000 mg/kg dry soil) and Cd (0.0 – 100 mg/kg dry soil). From a number of physiological indices measured in this study, microsymbionts significantly increased dry weight, root : shoot ratios, leaf number and area, plant length, leaf pigments, total carbohydrates, N and P content of infected plants as compared with non infected controls at all levels of heavy metal concentrations. Tolerance index of cowpea plants was increased in the presence of microsymbionts than in their absence in polluted soil. Microsymbionts dependencies of cowpea plants tended to be increased at higher levels of Zn and Cd in polluted soil. Metals accumulated by microsymbionts-infected cowpea plant were mostly distributed in root tissues, suggesting that an exclusion strategy for metal tolerance widely exists in them. This study provides evidence for benefits of NFB to AM fungi in the protection of host plants against the detrimental effects of heavy metals. If so, bacterial-AM-legume tripartite symbiosis could be a new approach to increase the heavy metal tolerance of legumes plants under heavy metal polluted soil.Keyword: Mycorrhiza, Vigna sinesis, heavy metals, microsymbiosis, Rhizobiu