Oxysterols direct immune cell migration via EBI2

Epstein-Barr virus-induced gene 2 (EBI2, also known as GPR183) is a G-protein-coupled receptor that is required for humoral immune responses; polymorphisms in the receptor have been associated with inflammatory autoimmune diseases. The natural ligand for EBI2 has been unknown. Here we describe the identification of 7α,25-dihydroxycholesterol (also called 7α,25-OHC or 5-cholesten-3β,7α,25-triol) as a potent and selective agonist of EBI2. Functional activation of human EBI2 by 7α,25-OHC and closely related oxysterols was verified by monitoring second messenger readouts and saturable, high-affinity radioligand binding. Furthermore, we find that 7α,25-OHC and closely related oxysterols act as chemoattractants for immune cells expressing EBI2 by directing cell migration in vitro and in vivo. A critical enzyme required for the generation of 7α,25-OHC is cholesterol 25-hydroxylase (CH25H). Similar to EBI2 receptor knockout mice, mice deficient in CH25H fail to position activated B cells within the spleen to the outer follicle and mount a reduced plasma cell response after an immune challenge. This demonstrates that CH25H generates EBI2 biological activity in vivo and indicates that the EBI2-oxysterol signalling pathway has an important role in the adaptive immune response.     

In vivo genome editing restores haemostasis in a mouse model of haemophilia

Editing of the human genome to correct disease-causing mutations is a promising approach for the treatment of genetic disorders. Genome editing improves on simple gene-replacement strategies by effecting in situ correction of a mutant gene, thus restoring normal gene function under the control of endogenous regulatory elements and reducing risks associated with random insertion into the genome. Gene-specific targeting has historically been limited to mouse embryonic stem cells. The development of zinc finger nucleases (ZFNs) has permitted efficient genome editing in transformed and primary cells that were previously thought to be intractable to such genetic manipulation. In vitro, ZFNs have been shown to promote efficient genome editing via homology-directed repair by inducing a site-specific double-strand break (DSB) at a target locus, but it is unclear whether ZFNs can induce DSBs and stimulate genome editing at a clinically meaningful level in vivo. Here we show that ZFNs are able to induce DSBs efficiently when delivered directly to mouse liver and that, when co-delivered with an appropriately designed gene-targeting vector, they can stimulate gene replacement through both homology-directed and homology-independent targeted gene insertion at the ZFN-specified locus. The level of gene targeting achieved was sufficient to correct the prolonged clotting times in a mouse model of haemophilia B, and remained persistent after induced liver regeneration. Thus, ZFN-driven gene correction can be achieved in vivo, raising the possibility of genome editing as a viable strategy for the treatment of genetic disease.     

Grains and grain boundaries in single-layer graphene atomic patchwork quilts

The properties of polycrystalline materials are often dominated by the size of their grains and by the atomic structure of their grain boundaries. These effects should be especially pronounced in two-dimensional materials, where even a line defect can divide and disrupt a crystal. These issues take on practical significance in graphene, which is a hexagonal, two-dimensional crystal of carbon atoms. Single-atom-thick graphene sheets can now be produced by chemical vapour deposition on scales of up to metres, making their polycrystallinity almost unavoidable. Theoretically, graphene grain boundaries are predicted to have distinct electronic, magnetic, chemical and mechanical properties that strongly depend on their atomic arrangement. Yet because of the five-order-of-magnitude size difference between grains and the atoms at grain boundaries, few experiments have fully explored the graphene grain structure. Here we use a combination of old and new transmission electron microscopy techniques to bridge these length scales. Using atomic-resolution imaging, we determine the location and identity of every atom at a grain boundary and find that different grains stitch together predominantly through pentagon-heptagon pairs. Rather than individually imaging the several billion atoms in each grain, we use diffraction-filtered imaging to rapidly map the location, orientation and shape of several hundred grains and boundaries, where only a handful have been previously reported. The resulting images reveal an unexpectedly small and intricate patchwork of grains connected by tilt boundaries. By correlating grain imaging with scanning probe and transport measurements, we show that these grain boundaries severely weaken the mechanical strength of graphene membranes but do not as drastically alter their electrical properties. These techniques open a new window for studies on the structure, properties and control of grains and grain boundaries in graphene and other two-dimensional materials.     

Acute vision in the giant Cambrian predator Anomalocaris and the origin of compound eyes

Until recently, intricate details of the optical design of non-biomineralized arthropod eyes remained elusive in Cambrian Burgess-Shale-type deposits, despite exceptional preservation of soft-part anatomy in such Konservat-Lagerst?tten. The structure and development of ommatidia in arthropod compound eyes support a single origin some time before the latest common ancestor of crown-group arthropods, but the appearance of compound eyes in the arthropod stem group has been poorly constrained in the absence of adequate fossils. Here we report 2-3-cm paired eyes from the early Cambrian (approximately 515 million years old) Emu Bay Shale of South Australia, assigned to the Cambrian apex predator Anomalocaris. Their preserved visual surfaces are composed of at least 16,000 hexagonally packed ommatidial lenses (in a single eye), rivalling the most acute compound eyes in modern arthropods. The specimens show two distinct taphonomic modes, preserved as iron oxide (after pyrite) and calcium phosphate, demonstrating that disparate styles of early diagenetic mineralization can replicate the same type of extracellular tissue (that is, cuticle) within a single Burgess-Shale-type deposit. These fossils also provide compelling evidence for the arthropod affinities of anomalocaridids, push the origin of compound eyes deeper down the arthropod stem lineage, and indicate that the compound eye evolved before such features as a hardened exoskeleton. The inferred acuity of the anomalocaridid eye is consistent with other evidence that these animals were highly mobile visual predators in the water column. The existence of large, macrophagous nektonic predators possessing sharp vision--such as Anomalocaris--within the early Cambrian ecosystem probably helped to accelerate the escalatory 'arms race' that began over half a billion years ago.     

ATP-induced helicase slippage reveals highly coordinated subunits

Helicases are vital enzymes that carry out strand separation of duplex nucleic acids during replication, repair and recombination. Bacteriophage T7 gene product 4 is a model hexameric helicase that has been observed to use dTTP, but not ATP, to unwind double-stranded (ds)DNA as it translocates from 5' to 3' along single-stranded (ss)DNA. Whether and how different subunits of the helicase coordinate their chemo-mechanical activities and DNA binding during translocation is still under debate. Here we address this question using a single-molecule approach to monitor helicase unwinding. We found that T7 helicase does in fact unwind dsDNA in the presence of ATP and that the unwinding rate is even faster than that with dTTP. However, unwinding traces showed a remarkable sawtooth pattern where processive unwinding was repeatedly interrupted by sudden slippage events, ultimately preventing unwinding over a substantial distance. This behaviour was not observed with dTTP alone and was greatly reduced when ATP solution was supplemented with a small amount of dTTP. These findings presented an opportunity to use nucleotide mixtures to investigate helicase subunit coordination. We found that T7 helicase binds and hydrolyses ATP and dTTP by competitive kinetics such that the unwinding rate is dictated simply by their respective maximum rates V(max), Michaelis constants K(M) and concentrations. In contrast, processivity does not follow a simple competitive behaviour and shows a cooperative dependence on nucleotide concentrations. This does not agree with an uncoordinated mechanism where each subunit functions independently, but supports a model where nearly all subunits coordinate their chemo-mechanical activities and DNA binding. Our data indicate that only one subunit at a time can accept a nucleotide while other subunits are nucleotide-ligated and thus they interact with the DNA to ensure processivity. Such subunit coordination may be general to many ring-shaped helicases and reveals a potential mechanism for regulation of DNA unwinding during replication.     

Modular and predictable assembly of porous organic molecular crystals

Nanoporous molecular frameworks are important in applications such as separation, storage and catalysis. Empirical rules exist for their assembly but it is still challenging to place and segregate functionality in three-dimensional porous solids in a predictable way. Indeed, recent studies of mixed crystalline frameworks suggest a preference for the statistical distribution of functionalities throughout the pores rather than, for example, the functional group localization found in the reactive sites of enzymes. This is a potential limitation for 'one-pot' chemical syntheses of porous frameworks from simple starting materials. An alternative strategy is to prepare porous solids from synthetically preorganized molecular pores. In principle, functional organic pore modules could be covalently prefabricated and then assembled to produce materials with specific properties. However, this vision of mix-and-match assembly is far from being realized, not least because of the challenge in reliably predicting three-dimensional structures for molecular crystals, which lack the strong directional bonding found in networks. Here we show that highly porous crystalline solids can be produced by mixing different organic cage modules that self-assemble by means of chiral recognition. The structures of the resulting materials can be predicted computationally, allowing in silico materials design strategies. The constituent pore modules are synthesized in high yields on gram scales in a one-step reaction. Assembly of the porous co-crystals is as simple as combining the modules in solution and removing the solvent. In some cases, the chiral recognition between modules can be exploited to produce porous organic nanoparticles. We show that the method is valid for four different cage modules and can in principle be generalized in a computationally predictable manner based on a lock-and-key assembly between modules.     

Gene polymorphism in Netherton and common atopic disease.

Atopic dermatitis (AD) and asthma are characterized by IgE-mediated atopic (allergic) responses to common proteins (allergens), many of which are proteinases. Loci influencing atopy have been localized to a number of chromosomal regions, including the chromosome 5q31 cytokine cluster. Netherton disease is a rare recessive skin disorder in which atopy is a universal accompaniment. The gene underlying Netherton disease (SPINK5) encodes a 15-domain serine proteinase inhibitor (LEKTI) which is expressed in epithelial and mucosal surfaces and in the thymus. We have identified six coding polymorphisms in SPINK5 (Table 1) and found that a Glu420-->Lys variant shows significant association with atopy and AD in two independent panels of families. Our results implicate a previously unrecognized pathway for the development of common allergic illnesses.     

基于拉格朗日系统中约束力思想的编队卫星构形非线性控制方法

借鉴经典动力学中约束力的思想，提出了一种编队卫星构形精确保持的非线性控制方法．该方法首先将非线性和摄动条件下编队卫星构形保持问题转换为带有完整约束的拉格朗日动力学系统，然后将问题转换为一组微分代数方程，通过求解微分代数方程，确定编队卫星构形保持的非线性控制律．由于借鉴了约束力的思想，该方法自然地利用了编队卫星动力学系统的力学特性，具有节省能量和高精度的特点．通过对线性和非线性条件下空间圆编队卫星构形保持问题的仿真，验证了提出的非线性控制方法的这些特性．     

卫星低频电磁辐射在轨探测研究

利用地球空间探测双星计划探测一号卫星上的磁场波动分析仪的原始数据, 分析了探测一号卫星在轨电磁辐射的特性. 结果显示卫星的电磁辐射主要集中在30 Hz以下. 在30 Hz以上, 卫星的电磁辐射最多延伸到 190 Hz左右, 而且强度明显减弱. 在 190 Hz以下的卫星电磁辐射具有与卫星姿态相关的长周期变化. 在 190~830 Hz的范围的电磁辐射有不明显的长周期变化特征. 830~3990 Hz范围的电磁辐射没有长周期变化特征. 卫星电磁辐射的长周期变化是由卫星姿态变化造成的. 卫星姿态变化引起卫星太阳方位角变化. 卫星太阳方位角越大, 卫星电磁辐射越大. 卫星太阳方位角从90.6增加到93.6, 低于10 Hz以下的电磁辐射约增大为原来的9倍, 10~190 Hz范围的电磁辐射大约增加到原来的1.6倍. 卫星在＜10和10~190 Hz范围内的电磁辐射强度与卫星太阳方位角的相关系数分别达到0.90和0.91. 卫星在光照情况下的电磁辐射要比卫星在阴影情况下大. 卫星太阳能帆板电流产生的电磁辐射是卫星电磁辐射主要来源, 约占整个卫星电磁辐射的87％(低频段<150 Hz)和94％(高频段>150 Hz). 这些中国首次对卫星电磁辐射的在轨探测结果对于我国未来相关科学和应用卫星的设计方案的优化具有重要的参考价值.     

Beta-catenin/TCF4 transactivates miR-30e during intestinal cell differentiation

The Wnt/beta-catenin/TCF4 pathway plays critical roles in the maintenance of small intestinal epithelium; however, downstream targets of the beta-catenin/TCF4 complex are not extensively characterized. We identified miR-30e as an immediate target activated by the beta-catenin/TCF4 complex. miR-30e was detected in the peri-nuclear region of the intestinal crypt IEC-6 cells. Bioinformatics analysis revealed clustered beta-catenin/TCF4 binding sites within the miR-30e promoter region. This promoter region was cloned into pGL3-control luciferase reporter vector, with the enhancer region removed. Transfection of pCMV-SPORT6-beta-catenin expression vector dose-dependently increased luciferase activity, and co-transfection of pCMV-SPORT6-TCF4 expression vector further enhanced the promoter activity. Dexamethasone-induced IEC-6 cells differentiation caused a 2.5-fold increase in miR-30e expression, and upon beta-catenin siRNA transfection, miR-30e increased 1.3-fold. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay confirmed the binding between beta-catenin/TCF4 complexes from IEC-6 nuclear extracts and the putative sequences in the miR-30e promoter. These results demonstrate that beta-catenin/TCF4 transactivates miR-30e during intestinal cell differentiation.