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Coulombic efficiency has been widely verified as a quantitative index of the reversibility of metal-ion batteries. The problem of low initial coulombic efficiency (ICE) has hindered the practical application of biomass-derived carbon as anode active materials. Herein, we propose a simple biomass-derived carbon based on the bark of melaleuca by acidizing-sintering method. The structure-activity relationship between the acidizing-sintering condition and the tortuosity gradient of the pore structure is also established. As an anode material of sodium-ion batteries, the biomass-derived carbon driven by tortuosity gradient regulation achieves an ultra-high ICE of 95.75 % and a high initial reversible specific capacity of 263 mAh/g at 280 mA/g, and also shows up to 92 % long cycle capacity retention. Through analyzing the results of small-angle X-ray scattering, Bruner-Emmett-Teller and HRTEM, the effect of the sintering temperature in regulating the tortuosity of the material can be determined. Besides, by combining in-situXRD, ex-situXPS, and various electrochemical measurement methods, it is confirmed that the surface structure with high tortuosity can be used as the reaction equilibrium layer to restrain the side reaction with the electrolyte, which can effectively inhibit the formation of unstable solid electrolyte interphase; and the low-tortuosity bulk phase pore structure serves as a fast transport layer, providing more internal space and sites for rapid deintercalation of metal ions, thus simultaneously achieving extremely high ICE and high reversible capacity output. At the same time, this study proves that the strategy can be extended to potassium-ion batteries, showing high efficiency and stable potassium storage characteristics.

Better understanding and control of residual stress in the chip build-up layer are becoming more and more important for the assembly process. To estimate the chip warpage and characterize the residual stress, different methods are proposed. However, most of them have high cost or some limitations for the upper build-up material. In this study, an innovative method is proposed to characterize the residual stress and predict the chip warpage behavior of different size chips at different temperatures. The method combines experimental inspection of chip warpage and finite element analysis. By reducing the silicon die thickness, the influence of residual stress in the build-up layer can be amplified. The residual stress can be obtained by inspecting the increased warpage when the silicon dies are reduced to different thicknesses. Correlating the thermal increase warpages of thinner chips can help characterize the effective modulus and coefficient of thermal expansion (CTE) of the build-up layer. This study can help better understand the commonly classified build-up layer information. The results show good agreements between two types of samples under the same upstream process flow.

ABSTRACTPorcine OTX2 was found to be highly activated in porcine iPS cells (piPSCs) that were reported by different laboratories worldwide. To reveal the regulatory function of OTX2 in porcine reprogrammed cells, we screened porcine miRNA-seq databases and found two miRNAs, miR-1343 and miR-545, that could specifically bind to 3ʹUTR of OTX2and suppress endogenous OTX2 expression in piPSCs. Knockdown of OTX2 by miR-1343 and miR-545 could significantly increase the expression of SOX2and ESRRB, but did not alter the expressions of OCT4 and KLF4, and improve the pluripotency of piPSCs. The promoter-based assays showed that OTX2 potentially bound to the promoter region of SOX2and ESRRBand suppressed their expression. On the other hand, SOX2 could interact with OTX2promoter. Ectopic expression of SOX2 could significantly decrease OTX2promoter activity, showing that there is a negative feedback loop between SOX2 and OTX2. Additionally, SOX2 and ESRRB significantly stimulated miR-1343 expression in piPSCs, but OTX2 down regulated the expression of miR-1343 in either direct or indirect manners. In summary, this study demonstrates that there is a regulatory network mediated by miR-1343, in which downregulation of OTX2 by miR-1343 can elevate the expression of pluripotent genes that were then sustain the pluripotency of piPSCs.

Ten-eleven translocation (TET) proteins play key roles in the regulation of DNA-methylation status by oxidizing 5-methylcytosine (5mC) to generate 5-hydroxymethylcytosine (5hmC), which can both serve as a stable epigenetic mark and participate in active demethylation. Unlike the other members of the TET family, TET2 does not contain a DNA-binding domain, and it remains unclear how it is recruited to chromatin. Here we show that TET2 is recruited by the RNA-binding protein Paraspeckle component 1 (PSPC1) through transcriptionally active loci, including endogenous retroviruses (ERVs) whose long terminal repeats (LTRs) have been co-opted by mammalian genomes as stage- and tissue-specific transcriptional regulatory modules. We found that PSPC1 and TET2 contribute to ERVL and ERVL-associated gene regulation by both transcriptional repression via histone deacetylases and post-transcriptional destabilization of RNAs through 5hmC modification. Our findings provide evidence for a functional role of transcriptionally active ERVs as specific docking sites for RNA epigenetic modulation and gene regulation. The authors show that TET2 is recruited to chromatin by the RNA-binding protein PSPC1. PSPC1 and TET2 contribute to ERV and ERV-associated gene regulation by both transcriptional repression via histone deacetylases and post-transcriptional destabilization of ERV RNAs through 5hmC modification.

Sodium-ion batteries (SIB) have attracted widespread attention in large-scale energy storage fields owing to the abundant reserve in the earth and similar properties of sodium to lithium. Biomass-based carbon materials with low-cost, controllable structure, simple processing technology, and environmental friendliness tick almost all the right boxes as one of the promising anode materials for SIB. Herein, we present a simple novel strategy involving tea tomenta biomass-derived carbon anode with enhanced interlayer carbon distance (0.44 nm) and high performance, which is constructed by N,P co-doped hard carbon (Tea-1100-NP) derived from tea tomenta. The prepared Tea-1100-NP composite could deliver a high reversible capacity (326.1 mAh/g at 28 mA/g), high initial coulombic efficiency (ICE = 90% at 28 mA/g), stable cycle life (262.4 mAh/g at 280 mA/g for 100 cycles), and superior rate performance (224.5 mAh/g at 1400 mA/g). Experimental results show that the excellent electrochemical performance of Tea-1100-NP due to the high number of active N,P-containing groups, and disordered amorphous structures provide ample active sites and increase the conductivity, meanwhile, large amounts of microporous shorten the Na+diffusion distance as well as quicken ion transport. This work provides a new type of N,P co-doped high-performance tomenta-derived carbon, which may also greatly promote the commercial application of SIB.

CAMA-syn, a hybrid composed of N-terminal -helical segment of Cecropin A(amino acid 1-8) and Magainin 2 (amino acid 1-12), is a novel small peptide with the potent antibacterial and synergistic activity without cytotoxicity. In order to test the antibacterial function of CAMA-syn produced in mammalian cells, several vectors containing the synthesized CAMA-syn DNA fragment were constructed and transfected into recipient cells. The results showed that CAMAsyn fusion to green fluorescent protein (GFP) or to hemagglutinin epitope (HA) tag was expressed in both bovine embryo fibroblasts (BEF) and mouse macrophage RAW264.7 cells. The antibacterial assays of CAMA-syn were conducted against both Gram positive and negative bacteria, including S. abortusovis, P. anatis, S. hyicus and S. suis. The results of colony-forming efficiency and cell growth curves proved that the in vitro expressed CAMA-syn could have the antibacterial activity, demonstrating that macrophage specific expression of antimicrobial peptide CAMA-syn could inhibit the growth of bacteria.

We investigated the role of cysteine residues in activity and localization of the NHE1 isoform of the Na+/H+exchanger. Each of the nine cysteine residues was mutated to serine or arginine. Mutation of the first serine (amino acid number 9) and serine number six (amino acid number 477) resulted in dramatic decreases in detectable activity of the Na+/H+exchanger when transfected into AP-1 cells. Some other mutations resulted in minor decreases in activity of the protein. Confocal and light microscopy of mutant cells with decreased activity showed that the antiporter protein was mostly retained in an intracellular compartment which colocalized with the medial-Golgi cisternae. Smaller amounts of active protein still remained targeted to the plasma membrane in these mutants. Treatment of wild-type cells with DTT also caused the retention of the Na+/H+exchanger to the same intracellular compartment. The results suggest that cysteines play an important role in intracellular folding and trafficking of the Na+/H+exchanger.

No abstract

A number of nuclear-encoded proteins are known to bind to the mitochondrial genome and may be involved in mitochondrial replication and in mitochondrial diseases. Mitochondrial diseases such as Kearns–Sayre syndrome (KSS) are flanked by common direct repeats. To study protein binding to these mitochondria DNA regions we used gel mobility shift binding assays. Proteins present in nuclear or in mitochondrial extracts from normal or KSS-derived fibroblasts bound to a 280-bp mitochondrial DNA fragment encompassing a deletion breakpoint in the mitochondrial genome. In addition, nuclear and mitochondrial protein bound to a nearby surrounding fragment and to a synthetic oligonucleotide with a related consensus DNA sequence. Southwestern blot analysis showed that the DNA binding ability resided in a 35-kDa protein. The amino terminal sequence of the 35-kDa protein was very similar to human high mobility group proteins. These results suggest that this protein may be involved in mitochondrial DNA deletions.

Abstract:  We examined the expression and activity of the Na+/H+ exchanger in the human choriocarcinoma BeWo cell line. When treated with methotrexate, these cells differentiated from cytotrophoblast-like cells to enlarged multinucleate syncytiotrophoblast-like cells. There was no change in the apparent K m for Na+ between undifferentiated and differentiated cells. However, differentiated cells could transport more than five times the proton flux of undifferentiated cells. There was no difference in the Hill coefficient between undifferentiated and differentiated cells. However, the maximal flux (J max) for undifferentiated cells was higher than that for differentiated cells. Inhibition of Na+/H+ exchange activity by an amiloride analog and Hoe694 revealed a sensitive and a resistant component in both differentiated and undifferentiated cells. Northern blot analysis and immunocytochemistry suggested that the sensitive component was due to the NHE1 isoform of the protein while the resistant component was due to the NHE3 isoform. The NHE1 isoform was localized to the brush border membrane of BeWo cells and Western blot analysis showed that the NHE1 protein was more abundant in brush border membranes from differentiated BeWo cells compared to undifferentiated cells. The results show that BeWo cells contain the NHE1 and NHE3 isoforms of the Na+/H+ exchanger and that the NHE1 isoform is primarily localized to the brush border membrane.

In this study we characterized regulation of the Na+/H+exchanger promoter in several tissue types. A conserved poly (dA:dT) region was important in regulation of the promoter. Nuclear extracts from rat myocardium and from mouse proximal tubule cells protected the poly (dA:dT) region of the NHE1 promoter. A protein from nuclear extracts also bound to the poly (dA:dT) element in gel mobility shift binding assays. The binding was specific and was removed by mutations in the poly (dA:dT) region. Characterization of the binding to the poly (dA:dT) region in gel mobility shift assays showed that it was reduced by high concentrations of the divalent cations Mg++and Mn++. The inhibition by divalent cations was reduced by decreasing the pH of the binding assay. N-terminal sequencing of the poly (dA:dT) binding protein showed that it was a member of the HMG (high mobility group) family of nuclear proteins which are important in cell growth and proliferation. The results are the first direct detection of a protein that regulates the NHE1 promoter.

The Na+/H+exchanger is a ubiquitous plasma membrane protein that is responsible for pH regulation and is activated by growth factors. We examined the role of the Na+/H+exchanger in cell growth and differentiation. Treatment of P19 cells with the Na+/H+exchanger inhibitor Hoe 694 eliminated retinoic acid-induced differentiation in this cell line. We developed a P19 embryonal carcinoma cell line that was deficient in the Na+/H+antiporter. Na+/H+exchanger-deficient cells were reduced in the rate of cell growth and this effect was enhanced by the removal of added HCO3−and by reducing extracellular pH. The antiporter-deficient cells were also markedly deficient in their ability to differentiate to neuronal-like cells and recovered this ability when the Na+/H+antiporter was reintroduced. The results show that the absence of Na+/H+antiport as a pH regulatory mechanism can result in deficiencies in both cell growth and differentiation in embryonal carcinoma cells.

In this study we examine regulation of expression of the Na+/H+exchanger promoter in L6 and NIH 3T3 cells. We have identified a highly conserved poly(dA·dT)-rich region that appears to be important in regulation of expression of the NHE1 gene. Deletion or mutation of this region results in dramatic decreases in promoter activity in both L6 and NIH 3T3 cells. In addition, DNase I footprinting experiments demonstrated that this region is protected by nuclear extracts from both cell types, and gel mobility shift assays showed that a protein or proteins specifically binds to the poly(dA·dT)-rich element. Using Southwestern blotting, we determined that a 33-kDa protein binds to the poly(dA·dT)-containing region. Mutations that abolished protein binding to this element diminished activity of the promoter. Insertion of the poly(dA·dT)-rich element into a plasmid containing the SV40 promoter demonstrated that this element can also enhance the activity of a foreign promoter. Together, the results we have presented here show that the poly(dA·dT)-rich region is important in regulation of NHE1 expression in different cell types.

Mouse embryonic stem cells (ESCs) sporadically express preimplantation two-cell-stage (2C) transcripts, including MERVL endogenous retrovirus and Zscan4 cluster genes. Such 2C-like cells (2CLCs) can contribute to both embryonic and extraembryonic tissues when reintroduced into early embryos, although the molecular mechanism underlying such an expanded 2CLC potency remains elusive. We examine global nucleosome occupancy and gene expression in 2CLCs and identified miR-344 as the noncoding molecule that positively controls 2CLC potency. We find that activation of endogenous MERVL or miR-344-2 alone is sufficient to induce 2CLCs with activation of 2C genes and an expanded potency. Mechanistically, miR-344 is activated by DUX and post-transcriptionally represses ZMYM2 and its partner LSD1, and ZMYM2 recruits LSD1/HDAC corepressor complex to MERVL LTR for transcriptional repression. Consistently, zygotic depletion of Zmym2 compromises the totipotency-to-pluripotency transition during early development. Our studies establish the previously unappreciated DUX-miR-344-Zmym2/Lsd1 axis that controls MERVL for expanded stem cell potency.

Previous reports have demonstrated that embryonic stem cells were capable of differentiating into primordial germ cells through the formation of embryoid bodies that subsequently generated oocyte-like cells (OLCs). Such a process could facilitate studies of primordial follicle oocyte development in vitro and regenerative medicine. To investigate the pluripotency of human amniotic fluid stem cells (hAFSCs) and their ability to differentiate into germ cells, we isolated a CD117+/CD44+hAFSC line that showed fibroblastoid morphology and intrinsically expressed both stem cell markers (OCT4, NANOG, SOX2) and germ cell markers (DAZL, STELLA). To encourage differentiation into OLCs, the hAFSCs were first cultured in a medium supplemented with 5% porcine follicular fluid for 10 days. During the induction period, cell aggregates formed and syntheses of steroid hormones were detected; some OLCs and granulosa cell-like cells could be loosened from the surface of the culture dish. Cell aggregates were collected and replated in oocyte culture medium for an additional 7–10 days. OLCs ranging from 50 to 120 μm presenting zona pellucida were observed in cumulus-oocyte complexes; some OLCs developed spontaneously into multicell structures similar to preimplantation embryos. Approximately 2% of the hAFSCs differentiated to meiotic germ cells that expressed folliculogenesis- and oogenesis-associated markers. Although the in vitro maturation and fertilization potentials are as yet unproven, short-term (<25 days) and high-efficiency (>2%) derivation of OLCs from hAFSCs might provide a new approach to the study of human germ cell development in vitro.