Shaping Giant Membrane Vesicles in 3D-Printed Protein Hydrogel Cages

Giant unilamellar phospholipid vesicles are attractive starting points for constructing minimal living cells from the bottom-up. Their membranes are compatible with many physiologically functional modules and act as selective barriers, while retaining a high morphological flexibility. However, their spherical shape renders them rather inappropriate to study phenomena that are based on distinct cell shape and polarity, such as cell division. Here, a microscale device based on 3D printed protein hydrogel is introduced to induce pH-stimulated reversible shape changes in trapped vesicles without compromising their free-standing membranes. Deformations of spheres to at least twice their aspect ratio, but also toward unusual quadratic or triangular shapes can be accomplished. Mechanical force induced by the cages to phase-separated membrane vesicles can lead to spontaneous shape deformations, from the recurrent formation of dumbbells with curved necks between domains to full budding of membrane domains as separate vesicles. Moreover, shape-tunable vesicles are particularly desirable when reconstituting geometry-sensitive protein networks, such as reaction-diffusion systems. In particular, vesicle shape changes allow to switch between different modes of self-organized protein oscillations within, and thus, to influence reaction networks directly by external mechanical cues

Correlates of genetic monogamy in socially monogamous mammals: insights from Azara's owl monkeys

Understanding the evolution of mating systems, a central topic in evolutionary biology for more than 50 years, requires examining the genetic consequences of mating and the relationships between social systems and mating systems. Among pair-living mammals, where genetic monogamy is extremely rare, the extent of extra-group paternity rates has been associated withmale participation in infant care, strength of the pair bond and length of the breeding season. This study evaluated the relationship between two of those factors and the genetic mating system of socially monogamous mammals, testing predictions that male care and strength of pair bond would be negatively correlated with rates of extra-pair paternity (EPP). Autosomal microsatellite analyses provide evidence for genetic monogamy in a pair-living primate with bi-parental care, the Azara’s owl monkey (Aotus azarae). A phylogenetically corrected generalized least square analysis was used to relate male care and strength of the pair bond to their genetic mating system (i.e. proportions of EPP) in 15 socially monogamous mammalian species. The intensity of male care was correlated with EPP rates in mammals, while strength of pair bond failed to reach statistical significance. Our analyses showthat, once social monogamy has evolved, paternal care, and potentially also close bonds, may facilitate the evolution of genetic monogamy.German Science Foundation (HU 1746/2-1); Wenner-Gren Foundation; L.S.B. Leakey Foundation;National Geographic Society; National Science Foundation (BCS-0621020, 1219368, and 1232349); the University of Pennsylvania Research Foundation; the Zoological Society of San Dieg

Setting targets for HIV/AIDS-What lessons can be learned from other disease control programmes?

Our analysis of experience from programmes targeting malaria, leprosy and TB shows the importance of drawing broadly on research and implementation expertise, and civil society more broadly, when setting targets for HIV control. The engagement of stakeholders from the highest burden settings, including affected populations, is crucial, to ensure that disease control efforts uphold human rights and tackle HIV-related stigma and discrimination. An appropriate balance is needed between ambitious, galvanising global targets that drive funding and political/public engagement, and targets that reflect the complexities and local epidemiological variations in disease profile. Ethical issues and unintended consequences need to be considered when setting targets—particularly around local effects and opportunity costs of having foregone other areas of disease control and public health. Intermediate and adaptable targets are needed that allow for course corrections to programmes. Overly burdensome reporting requirements for individual local programmes and countries should be avoided, as well as potential for overlapping and sometimes conflicting targets both within and across vertical disease programmes. Process targets should be distinguished from outcome targets, which should be measurable and based on high-quality data. Retention of expert healthcare worker skills and specialist services is vital, while moving towards integrated health systems if effective disease control programmes are to be maintained. Target development should seek areas of programme delivery where an opportunity to codevelop targets and integrate services exists. Global efforts to move to universal health coverage (UHC), for example, could be factored in when developing targets. Sustaining investment and continuing political interest in the end phase of any elimination or eradication strategy, once incidence and prevalence are low, are critical to achieve success. Equity- and access-based service delivery targets become increasingly important as the elimination strategy nears its end and should be factored into planning. Achieving disease elimination and/or eradication is only possible with sufficient investment in research to develop new prevention tools such as vaccines, point-of-care diagnostics, and treatments to counteract the effects of increasing drug resistance and the challenging latency period of diseases; public health infrastructure upgrades that address wider determinants of health; and health and surveillance systems that allow for equitable delivery and access to services

Rab6 and Rab11 Regulate Chlamydia trachomatis Development and Golgin-84-Dependent Golgi Fragmentation

Many intracellular pathogens that replicate in special membrane bound compartments exploit cellular trafficking pathways by targeting small GTPases, including Rab proteins. Members of the Chlamydiaceae recruit a subset of Rab proteins to their inclusions, but the significance of these interactions is uncertain. Using RNA interference, we identified Rab6 and Rab11 as important regulators of Chlamydia infections. Depletion of either Rab6 or Rab11, but not the other Rab proteins tested, decreased the formation of infectious particles. We further examined the interplay between these Rab proteins and the Golgi matrix components golgin-84 and p115 with regard to Chlamydia-induced Golgi fragmentation. Silencing of the Rab proteins blocked Chlamydia-induced and golgin-84 knockdown-stimulated Golgi disruption, whereas Golgi fragmentation was unaffected in p115 depleted cells. Interestingly, p115-induced Golgi fragmentation could rescue Chlamydia propagation in Rab6 and Rab11 knockdown cells. Furthermore, transport of nutrients to Chlamydia, as monitored by BODIPY-Ceramide, was inhibited by Rab6 and Rab11 knockdown. Taken together, our results demonstrate that Rab6 and Rab11 are key regulators of Golgi stability and further support the notion that Chlamydia subverts Golgi structure to enhance its intracellular development

Atmospheric greenhouse gases retrieved from SCIAMACHY: comparison to ground-based FTS measurements and model results

SCIAMACHY onboard ENVISAT (launched in 2002) enables the retrieval of global long-term column-averaged dry air mole fractions of the two most important anthropogenic greenhouse gases carbon dioxide and methane (denoted XCO_2 and XCH_4). In order to assess the quality of the greenhouse gas data obtained with the recently introduced v2 of the scientific retrieval algorithm WFM-DOAS, we present validations with ground-based Fourier Transform Spectrometer (FTS) measurements and comparisons with model results at eight Total Carbon Column Observing Network (TCCON) sites providing realistic error estimates of the satellite data. Such validation is a prerequisite to assess the suitability of data sets for their use in inverse modelling. It is shown that there are generally no significant differences between the carbon dioxide annual increases of SCIAMACHY and the assimilation system CarbonTracker (2.00 ± 0.16 ppm yr^(−1) compared to 1.94 ± 0.03 ppm yr−1 on global average). The XCO_2 seasonal cycle amplitudes derived from SCIAMACHY are typically larger than those from TCCON which are in turn larger than those from CarbonTracker. The absolute values of the northern hemispheric TCCON seasonal cycle amplitudes are closer to SCIAMACHY than to CarbonTracker and the corresponding differences are not significant when compared with SCIAMACHY, whereas they can be significant for a subset of the analysed TCCON sites when compared with CarbonTracker. At Darwin we find discrepancies of the seasonal cycle derived from SCIAMACHY compared to the other data sets which can probably be ascribed to occurrences of undetected thin clouds. Based on the comparison with the reference data, we conclude that the carbon dioxide data set can be characterised by a regional relative precision (mean standard deviation of the differences) of about 2.2 ppm and a relative accuracy (standard deviation of the mean differences) of 1.1–1.2 ppm for monthly average composites within a radius of 500 km. For methane, prior to November 2005, the regional relative precision amounts to 12 ppb and the relative accuracy is about 3 ppb for monthly composite averages within the same radius. The loss of some spectral detector pixels results in a degradation of performance thereafter in the spectral range currently used for the methane column retrieval. This leads to larger scatter and lower XCH_4 values are retrieved in the tropics for the subsequent time period degrading the relative accuracy. As a result, the overall relative precision is estimated to be 17 ppb and the relative accuracy is in the range of about 10–20 ppb for monthly averages within a radius of 500 km. The derived estimates show that the SCIAMACHY XCH_4 data set before November 2005 is suitable for regional source/sink determination and regional-scale flux uncertainty reduction via inverse modelling worldwide. In addition, the XCO2 monthly data potentially provide valuable information in continental regions, where there is sparse sampling by surface flask measurements

In situ microfluidic cryofixation for cryo Focused Ion Beam milling and cryo electron tomography.

We present a microfluidic platform for studying structure-function relationships at the cellular level by connecting video rate live cell imaging with in situ microfluidic cryofixation and cryo-electron tomography of near natively preserved, unstained specimens. Correlative light and electron microscopy (CLEM) has been limited by the time required to transfer live cells from the light microscope to dedicated cryofixation instruments, such as a plunge freezer or high-pressure freezer. We recently demonstrated a microfluidic based approach that enables sample cryofixation directly in the light microscope with millisecond time resolution, a speed improvement of up to three orders of magnitude. Here we show that this cryofixation method can be combined with cryo-electron tomography (cryo-ET) by using Focused Ion Beam milling at cryogenic temperatures (cryo-FIB) to prepare frozen hydrated electron transparent sections. To make cryo-FIB sectioning of rapidly frozen microfluidic channels achievable, we developed a sacrificial layer technique to fabricate microfluidic devices with a PDMS bottom wall <5 µm thick. We demonstrate the complete workflow by rapidly cryo-freezing Caenorhabditis elegans roundworms L1 larvae during live imaging in the light microscope, followed by cryo-FIB milling and lift out to produce thin, electron transparent sections for cryo-ET imaging. Cryo-ET analysis of initial results show that the structural preservation of the cryofixed C. elegans was suitable for high resolution cryo-ET work. The combination of cryofixation during live imaging enabled by microfluidic cryofixation with the molecular resolution capabilities of cryo-ET offers an exciting avenue to further advance space-time correlative light and electron microscopy (st-CLEM) for investigation of biological processes at high resolution in four dimensions

Towards space based verification of CO₂ emissions from strong localized sources: fossil fuel power plant emissions as seen by a CarbonSat constellation

Carbon dioxide (CO₂) is the most important man-made greenhouse gas (GHG) that cause global warming. With electricity generation through fossil-fuel power plants now being the economic sector with the largest source of CO₂, power plant emissions monitoring has become more important than ever in the fight against global warming. In a previous study done by Bovensmann et al. (2010), random and systematic errors of power plant CO₂ emissions have been quantified using a single overpass from a proposed CarbonSat instrument. In this study, we quantify errors of power plant annual emission estimates from a hypothetical CarbonSat and constellations of several CarbonSats while taking into account that power plant CO₂ emissions are time-dependent. Our focus is on estimating systematic errors arising from the sparse temporal sampling as well as random errors that are primarily dependent on wind speeds. We used hourly emissions data from the US Environmental Protection Agency (EPA) combined with assimilated and re-analyzed meteorological fields from the National Centers of Environmental Prediction (NCEP). CarbonSat orbits were simulated as a sun-synchronous low-earth orbiting satellite (LEO) with an 828-km orbit height, local time ascending node (LTAN) of 13:30 (01:30 p.m. LT) and achieves global coverage after 5 days. We show, that despite the variability of the power plant emissions and the limited satellite overpasses, one CarbonSat has the potential to verify reported US annual CO₂ emissions from large power plants (&ge;5 Mt CO₂ yr⁻¹) with a systematic error of less than ~4.9% and a random error of less than ~6.7% for 50% of all the power plants. For 90% of all the power plants, the systematic error was less than ~12.4% and the random error was less than ~13%. We additionally investigated two different satellite configurations using a combination of 5 CarbonSats. One achieves global coverage everyday but only samples the targets at fixed local times. The other configuration samples the targets five times at two-hour intervals approximately every 6th day but only achieves global coverage after 5 days. From the statistical analyses, we found, as expected, that the random errors improve by approximately a factor of two if 5 satellites are used. On the other hand, more satellites do not result in a large reduction of the systematic error. The systematic error is somewhat smaller for the CarbonSat constellation configuration achieving global coverage everyday. Therefore, we recommend the CarbonSat constellation configuration that achieves daily global coverage