Epigenetic regulation underlies the robust changes in gene expression that occur during development. How precisely epigenetic enzymes contribute to development and differentiation processes is largely unclear. Here we show that one of the enzymes that removes the activating epigenetic mark of trimethylated lysine 4 on histone H3, lysine (K)-specific demethylase 5A (KDM5A), reinforces the effects of the retinoblastoma (RB) family of transcriptional repressors on differentiation. Global location analysis showed that KDM5A cooccupies a substantial portion of target genes with the E2F4 transcription factor. During ES cell differentiation, knockout of KDM5A resulted in derepression of multiple genomic loci that are targets of KDM5A, denoting a direct regulatory function. In terminally differentiated cells, common KDM5A and E2F4 gene targets were bound by the pRB-related protein p130, a DREAM complex component. KDM5A was recruited to the transcription start site regions independently of E2F4; however, it cooperated with E2F4 to promote a state of deepened repression at cell cycle genes during differentiation. These findings reveal a critical role of H3K4 demethylation by KDM5A in the transcriptional silencing of genes that are suppressed by RB family members in differentiated cells.chromatin | histone demethylase | whole-genome sequencing | histone methylation R egulation of gene expression is accomplished by transcription factors, histone-modifying enzymes, and chromatin remodeling machinery. The combined action of these three has been implicated in a number of biological processes, including cell cycle control, development, reprogramming, differentiation, and aging. Deregulation of chromatin-modifying enzymes has been strongly linked to the development of cancer. For example, two enzymes that regulate methylation at the histone H3 lysine 4 (H3K4) residue, mixed lineage leukemia-1 (MLL1) and lysine (K)-specific demethylase 5A (KDM5A), have been identified in translocations associated with human leukemia. H3K4 histone methylation states exhibit a highly distinct distribution pattern in the genome. Specifically, H3K4 trimethylation (H3K4me3) is strongly associated with transcriptional activation, with the highest levels observed near transcriptional start sites (TSS). In vitro studies suggest that the four KDM5 enzymes (KDM5A, KDM5B, KDM5C, and KDM5D) are able to remove methylation at lysine 4 of histone H3, and in vivo KDM5A and KDM5B may be recruited to common gene regions (1). This finding leads to multiple questions: (i) Do KDM5 proteins play a role in gene regulation by transcription factors? (ii) Are KDM5 enzymes nonredundant H3K4 demethylases?During differentiation, cells exhibit two novel properties: repression of cell cycle genes associated with permanent cell cycle exit and activation of cell type-specific genes. Histone modifications are thought to be important epigenetic events intimately linked to initiation and maintenance of transcriptional changes for both of these processes. Cell cycle exit is associat...
Synthesized transparent CQD–PVA composite films performed stable UV-A blocking, even after exposure to UV light for several days and elevated temperature.
In graphene, the extremely fast charge carriers can be controlled by electron-optical elements, such as waveguides, in which the transmissivity is tuned by the wavelength. In this work, charge carriers are guided in a suspended ballistic few-mode graphene channel, defined by electrostatic gating. By depleting the channel, a reduction of mode number and steps in the conductance are observed, until the channel is completely emptied. The measurements are supported by tight-binding transport calculations including the full electrostatics of the sample.
A true biomimetic of the cartilage extracellular matrix (ECM) could greatly contribute to our ability to regenerate this tissue in a mechanically demanding, often inflamed environment. Articular cartilage is a composite tissue made of cells and fibrillar proteins embedded in a hydrophilic polymeric meshwork. Here, a polyanionic functionalized alginate is used to mimic the glycosaminoglycan component of the native ECM. To create the fibrillar component, cryoelectrospinning of poly(ε-caprolactone) on a -78 °C mandrel, subsequently treated by O plasma, is used to create a stable, ultraporous and hydrophillic nanofiber network. In this study, cell-laden, fiber-reinforced composite scaffolds thicker than 1.5 mm can be created by infiltrating a chondrocyte/alginate solution into the fiber mesh, which is then physically cross-linked. The fibrillar component significantly reinforces the chondroinductive, but mechanically weak sulfated alginate hydrogels. This allows the production of a glycosaminoglycan- and collagen type II-rich matrix by the chondrocytes as well as survival of the composite in vivo. To further enhance the system, the electrospun component is loaded with dexamethasone, which protected the cells from an IL-1β-mediated inflammatory insult.
We present an improved synthesis route to hollow silica particles starting from tetramethyl orthosilicate (TMOS) instead of the traditionally used ethyl ester. The silica was first deposited onto polystyrene (PS) particles that were later removed. The here introduced, apparently minor modification in synthesis, however, allowed for a very high purity material. The improved, low density hollow silica particles were successfully implemented into polymer films and permitted maintaining optical transparency while significantly improving the heat barrier properties of the composite. Mechanistic investigations revealed the dominant role of here used methanol as a cosolvent and its role in controlling the hydrolysis rate of the silicic ester, and subsequent formation of hollow silica particles. Systematic experiments using various reaction parameters revealed a transition between regions of inhomogeneous material production at fast hydrolysis rate and reliable silica deposition on the surface of PS as a core-shell structured particle. The shell-thickness was controlled from 6.2 to 17.4 nm by increasing TMOS concentration and the diameter from 95 to 430 nm through use of the different sizes of PS particles. Hollow silica particle with the shell-thickness about 6.2 nm displayed a high light transmittance intensity up to 95% at 680 nm (length of light path ∼ 1 cm). Polyethersulfone (PES)/hollow silica composite films (35 ± 5 μm thick) exhibited a much lower thermal conductivity (0.03 ± 0.005 W m·K(-1)) than pure polymer films. This indicates that the prepared hollow silica is able to be used for cost and energy effective optical devices requiring thermal insulation.
Polymer -metal organic framework (MOF) composite membranes are promising materials for gas separation and could be potentially applied within many industrial applications. However, key limitations of currently reported layered MOF-polymer composites are their lack of scalability and mechanical stability. A big challenge for synthesis of such composites is directing the growth of homogeneous, defect-free MOF crystal layers. Here, a membrane synthesis method allowing the formation of flexible, noncontinuous zeolitic imidazolate framework 8 (ZIF-8) − poly(ether sulfone) (PES) composite membranes is presented. The ZIF-8 growth is restricted to the PES pores by exploiting directed ZnO seed-nanoparticles. The seeding process is part of the membrane formation process itself and allows for specifically integrating ZnO within polymeric membranes enabling easy and scalable control of the MOF-crystal formation. During solvent casting and membrane formation, a phase separation process allows trapping ZnO seed-nanoparticles within bicontinuous PES pores. Because of ZnO serving in parallel as seed and zinc source, ZIF formation can be induced and controlled by adding only one solution containing the organic linker. This ZnO nanoparticle seeding technique enables a pore-specific in situ growth of small (<5 μm in diameter) ZIF-8 islands via solvothermal synthesis. This leads to mechanically flexible self-supporting ZIF-8 membranes exhibiting gas selectivities of 9.3 ± 3.1 (H 2 /CO 2 ) and 11.5 ± 2.1 (H 2 /N 2 ).
In reverse genetics, a gene’s function is elucidated through targeted modifications in the coding region or associated DNA cis-regulatory elements. To this purpose, recently developed customizable transcription activator-like effector nucleases (TALENs) have proven an invaluable tool, allowing introduction of double-strand breaks at predetermined sites in the genome. Here we describe a practical and efficient method for the targeted genome engineering in Drosophila. We demonstrate TALEN-mediated targeted gene integration and efficient identification of mutant flies using a traceable marker phenotype. Furthermore, we developed an easy TALEN assembly (easyT) method relying on simultaneous reactions of DNA Bae I digestion and ligation, enabling construction of complete TALENs from a monomer unit library in a single day. Taken together, our strategy with easyT and TALEN-plasmid microinjection simplifies mutant generation and enables isolation of desired mutant fly lines in the F1 generation.
Mechanistic and kinetic insights into the removal of soluble nanoparticles as templates for mesopores now permit scale-up of mesoporous polymer membranes. Investigations on the effect of pH and nanoparticle dissolution time showed that pH levels of 0 or 1 are necessary to enable fast pore template dissolution. Approximately 5 wt % of the originally applied nanoparticles remained in the membrane due to complete encapsulation by the polymer matrix but did not contaminate the permeate during prolonged use. We demonstrated continuous production of 17 m2 (100 m length) of membrane using a commercial roll-to-roll coating unit. These membranes were successfully applied in gravitation driven water filtration. The rates of rejection of bacteria from heavily contaminated natural pond water were higher than 99.99%.
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