Segregation of morphogenetic regulatory function of Shox2 from its cell fate guardian role in sinoatrial node development.

Shox2 plays a vital role in the morphogenesis and physiological function of the sinoatrial node (SAN), the primary cardiac pacemaker, manifested by the formation of a hypoplastic SAN and failed differentiation of pacemaker cells in Shox2 mutants. Shox2 and Nkx2-5 are co-expressed in the developing SAN and regulate the fate of the pacemaker cells through a Shox2-Nkx2-5 antagonistic mechanism. Here we show that simultaneous inactivation of Nkx2-5 in the SAN of Shox2 mutants (dKO) rescued the pacemaking cell fate but not the hypoplastic defects, indicating uncoupling of SAN cell fate determination and morphogenesis. Single-cell RNA-seq revealed that the presumptive SAN cells of Shox2-/- mutants failed to activate pacemaking program but remained in a progenitor state preceding working myocardium, while both wildtype and dKO SAN cells displayed normal pacemaking cell fate with similar cellular state. Shox2 thus acts as a safeguard but not a determinant to ensure the pacemaking cell fate through the Shox2-Nkx2-5 antagonistic mechanism, which is segregated from its morphogenetic regulatory function in SAN development.

Structural design of FeCo alloy implanted into N,S co-doped carbon nanotubes via self-catalyzed growth for advanced liquid and flexible all-state-state Zn-air battery.

The introduction of transition bimetallic alloys can effectively improve oxygen reduction reaction (ORR) activity. However, the alloy particles are inclined to dissolve under harsher conditions, resulting in a serious decrease in catalytic activity and stability. Herein, an efficient ORR catalyst, FeCo alloy nanoparticles (NPs) encapsulated in N,S co-doped carbon nanotubes (FeCo10-NSCNTs), was developed through a self-catalyzed growth strategy. Due to the delicate structural design, the N,S co-doped structure can effectively improve the ORR performance by modulating the electronic properties and surface polarity of the carbon substrate, and the randomly connected carbon nanotube structure with large specific surface area can further enhance the adsorption and dissociation of gas molecules, accelerating the kinetics of gas participation in the reaction. Carbon-encapsulated FeCo alloys are beneficial for improving catalytic activity and durability. The FeCo10-NSCNTs displayed excellent ORR activity with a half-wave potential of E1/2 = 0.84 V and robust stability of 13 k cycles. More impressively, the assembled liquid-state Zn-air battery (ZAB) with FeCo10-NSCNTs as the air-electrode delivers an output power density of 146.68 mW cm-2 along with excellent operation durability. The assembled all-solid ZAB has good cyclic stability under 0-180° bending conditions. The synthesized N,S co-doping, carbon nanotubes and FeCo alloys provide important guidance for the construction of cheap non-noble metal-carbon hybrid nanomaterials.

A molecular atlas reveals the tri-sectional spinning mechanism of spider dragline silk.

The process of natural silk production in the spider major ampullate (Ma) gland endows dragline silk with extraordinary mechanical properties and the potential for biomimetic applications. However, the precise genetic roles of the Ma gland during this process remain unknown. Here, we performed a systematic molecular atlas of dragline silk production through a high-quality genome assembly for the golden orb-weaving spider Trichonephila clavata and a multiomics approach to defining the Ma gland tri-sectional architecture: Tail, Sac, and Duct. We uncovered a hierarchical biosynthesis of spidroins, organic acids, lipids, and chitin in the sectionalized Ma gland dedicated to fine silk constitution. The ordered secretion of spidroins was achieved by the synergetic regulation of epigenetic and ceRNA signatures for genomic group-distributed spidroin genes. Single-cellular and spatial RNA profiling identified ten cell types with partitioned functional division determining the tri-sectional organization of the Ma gland. Convergence analysis and genetic manipulation further validated that this tri-sectional architecture of the silk gland was analogous across Arthropoda and inextricably linked with silk formation. Collectively, our study provides multidimensional data that significantly expand the knowledge of spider dragline silk generation and ultimately benefit innovation in spider-inspired fibers.

Two birds with one stone: large catalytic areas and abundant nitrogen sites inspired by fluorine doping contributing to CO2RR activity and selectivity.

Electroreduction of CO2 based on metal-free carbon catalysts is an attractive approach for useful products. However, it remains a great chemical challenge due to its unsatisfactory activity and poor selectivity. Here, we report a successful case to greatly improve CO2-to-CO conversion on carbon black (CB) and nitrogen-doped carbon black (N-CB). By introducing fluorine, the faradaic efficiency of CO was increased from 12.8% (CB) and 50.8% (N-CB) to 93.1% (nitrogen and fluorine co-doped carbon black, N,F-CB) at -0.7 V. A partial current density of 4.19 mA cm-2 remained durable for about 23 h. The superiority of N,F-CB can be attributed to its large catalytic areas and abundant N active sites inspired by fluorine doping. Specifically, the fluorine precursor of polyvinylidene fluoride (PVDF) firstly performs as a nitrogen fixator, protecting the catalyst from more nitrogen escaping during the carbonization treatment. The number of nitrogen sites is about 4.4 times higher than it is for the N-CB. Meanwhile, PVDF as the area extender significantly improves the catalytic area; the specific surface area and the ECSA of N,F-CB are 8.7 and 6.9 times higher than that of CB. This work provides an insight into how heteroatoms can manipulate catalytic activity and selectivity through the catalytic area of carbon materials with more active sites.

Discovery and functional assessment of a novel adipocyte population driven by intracellular Wnt/ß-catenin signaling in mammals.

Wnt/ß-catenin signaling has been well established as a potent inhibitor of adipogenesis. Here, we identified a population of adipocytes that exhibit persistent activity of Wnt/ß-catenin signaling, as revealed by the Tcf/Lef-GFP reporter allele, in embryonic and adult mouse fat depots, named as Wnt+ adipocytes. We showed that this ß-catenin-mediated signaling activation in these cells is Wnt ligand- and receptor-independent but relies on AKT/mTOR pathway and is essential for cell survival. Such adipocytes are distinct from classical ones in transcriptomic and genomic signatures and can be induced from various sources of mesenchymal stromal cells including human cells. Genetic lineage-tracing and targeted cell ablation studies revealed that these adipocytes convert into beige adipocytes directly and are also required for beige fat recruitment under thermal challenge, demonstrating both cell autonomous and non-cell autonomous roles in adaptive thermogenesis. Furthermore, mice bearing targeted ablation of these adipocytes exhibited glucose intolerance, while mice receiving exogenously supplied such cells manifested enhanced glucose utilization. Our studies uncover a unique adipocyte population in regulating beiging in adipose tissues and systemic glucose homeostasis.

Conjugated activation of myocardial-specific transcription of Gja5 by a pair of Nkx2-5-Shox2 co-responsive elements.

The sinoatrial node (SAN) is the primary pacemaker in the heart. During cardiogenesis, Shox2 and Nkx2-5 are co-expressed in the junction domain of the SAN and regulate pacemaker cell fate through a Shox2-Nkx2-5 antagonism. Cx40 is a marker of working myocardium and an Nkx2-5 transcriptional output antagonized by Shox2, but the underlying regulatory mechanisms remain elusive. Here we characterized a bona fide myocardial-specific Gja5 (coding gene of Cx40) distal enhancer consisting of a pair of Nkx2-5 and Shox2 co-bound elements in the regulatory region of Gja5. Transgenic reporter assays revealed that neither element alone, but the conjugation of both elements together, drives myocardial-specific transcription. Genetic analyses confirmed that the activation of this enhancer depends on Nkx2-5 but is inhibited by Shox2 in vivo, and its presence is essential for Gja5 expression in the myocardium but not the endothelial cells of the heart. Furthermore, chromatin conformation analysis showed an Nkx2-5-dependent loop formation between these two elements and the Gja5 promoter in vivo, indicating that Nkx2-5 bridges the conjugated activation of this enhancer by pairing the two elements to the Gja5 promoter.

The transcriptional regulator MEIS2 sets up the ground state for palatal osteogenesis in mice.

Haploinsufficiency of Meis homeobox 2 (MEIS2), encoding a transcriptional regulator, is associated with human cleft palate, and Meis2 inactivation leads to abnormal palate development in mice, implicating MEIS2 functions in palate development. However, its functional mechanisms remain unknown. Here we observed widespread MEIS2 expression in the developing palate in mice. Wnt1Cre -mediated Meis2 inactivation in cranial neural crest cells led to a secondary palate cleft. Importantly, about half of the Wnt1Cre ;Meis2f/f mice exhibited a submucous cleft, providing a model for studying palatal bone formation and patterning. Consistent with complete absence of palatal bones, the results from integrative analyses of MEIS2 by ChIP sequencing, RNA-Seq, and an assay for transposase-accessible chromatin sequencing identified key osteogenic genes regulated directly by MEIS2, indicating that it plays a fundamental role in palatal osteogenesis. De novo motif analysis uncovered that the MEIS2-bound regions are highly enriched in binding motifs for several key osteogenic transcription factors, particularly short stature homeobox 2 (SHOX2). Comparative ChIP sequencing analyses revealed genome-wide co-occupancy of MEIS2 and SHOX2 in addition to their colocalization in the developing palate and physical interaction, suggesting that SHOX2 and MEIS2 functionally interact. However, although SHOX2 was required for proper palatal bone formation and was a direct downstream target of MEIS2, Shox2 overexpression failed to rescue the palatal bone defects in a Meis2-mutant background. These results, together with the fact that Meis2 expression is associated with high osteogenic potential and required for chromatin accessibility of osteogenic genes, support a vital function of MEIS2 in setting up a ground state for palatal osteogenesis.

Shox2 regulates osteogenic differentiation and pattern formation during hard palate development in mice.

During mammalian palatogenesis, cranial neural crest-derived mesenchymal cells undergo osteogenic differentiation and form the hard palate, which is divided into palatine process of the maxilla and the palatine. However, it remains unknown whether these bony structures originate from the same cell lineage and how the hard palate is patterned at the molecular level. Using mice, here we report that deficiency in Shox2 (short stature homeobox 2), a transcriptional regulator whose expression is restricted to the anterior palatal mesenchyme, leads to a defective palatine process of the maxilla but does not affect the palatine. Shox2 overexpression in palatal mesenchyme resulted in a hyperplastic palatine process of the maxilla and a hypoplastic palatine. RNA sequencing and assay for transposase-accessible chromatin-sequencing analyses revealed that Shox2 controls the expression of pattern specification and skeletogenic genes associated with accessible chromatin in the anterior palate. This highlighted a lineage-autonomous function of Shox2 in patterning and osteogenesis of the hard palate. H3K27ac ChIP-Seq and transient transgenic enhancer assays revealed that Shox2 binds distal-acting cis-regulatory elements in an anterior palate-specific manner. Our results suggest that the palatine process of the maxilla and palatine arise from different cell lineages and differ in ossification mechanisms. Shox2 evidently controls osteogenesis of a cell lineage and contributes to the palatine process of the maxilla by interacting with distal cis-regulatory elements to regulate skeletogenic gene expression and to pattern the hard palate. Genome-wide Shox2 occupancy in the developing palate may provide a marker for identifying active anterior palate-specific gene enhancers.

Nkx2-5 defines a subpopulation of pacemaker cells and is essential for the physiological function of the sinoatrial node in mice.

The sinoatrial node (SAN), the primary cardiac pacemaker, consists of a head domain and a junction/tail domain that exhibit different functional properties. However, the underlying molecular mechanism defining these two pacemaker domains remains elusive. Nkx2-5 is a key transcription factor essential for the formation of the working myocardium, but it was generally thought to be detrimental to SAN development. However, Nkx2-5 is expressed in the developing SAN junction, suggesting a role for Nkx2-5 in SAN junction development and function. In this study, we present unambiguous evidence that SAN junction cells exhibit unique action potential configurations intermediate to those manifested by the SAN head and the surrounding atrial cells, suggesting a specific role for the junction cells in impulse generation and in SAN-atrial exit conduction. Single-cell RNA-seq analyses support this concept. Although Nkx2-5 inactivation in the SAN junction did not cause a malformed SAN at birth, the mutant mice manifested sinus node dysfunction. Thus, Nkx2-5 defines a population of pacemaker cells in the transitional zone. Despite Nkx2-5 being dispensable for SAN morphogenesis during embryogenesis, its deletion hampers atrial activation by the pacemaker.

Contamination characteristics of trace metals in dust from different levels of roads of a heavily air-polluted city in north China.

Concentrations of eight trace metals (TMs) in road dust (RD) (particles < 25 µm) from urban areas of Xinxiang, China, were determined by inductively coupled plasma mass spectrometry. The geometric mean concentrations of Zn, Mn, Pb, As, Cu, Cr, Ni and Cd were 489, 350, 114, 101, 60.0, 39.7, 31.6, and 5.1 mg kg-1, respectively. When compared with TM levels in background soil, the samples generally display elevated TM concentrations, except for Cr and Mn, and for Cd the enrichment value was 69.6. Spatial variations indicated TMs in RD from park path would have similar sources with main roads, collector streets and bypasses. Average daily exposure doses of the studied TMs were about three orders of magnitude higher for hand-to-mouth ingestion than dermal contact, and the exposure doses for children were 9.33 times higher than that for adults. The decreasing trend of calculated hazard indexes (HI) for the eight elements was As > Pb > Cr > Mn > Cd > Zn > Ni > Cu for both children and adults.

Pten regulates neural crest proliferation and differentiation during mouse craniofacial development.

BACKGROUND: The phosphatase and tensin homolog deleted on chromosome TEN (Pten) is implicated in a broad range of developmental events and diseases. However, its role in neural crest and craniofacial development has not been well illustrated. RESULTS: Using genetically engineered mouse models, we showed that inactivating Pten specifically in neural crest cells causes malformation of craniofacial structures. Pten conditional knockout mice exhibit perinatal lethality with overgrowth of craniofacial structures. At the cellular level, Pten deficiency increases cell proliferation rate and enhances osteoblast differentiation. Our data further revealed that inactivating Pten elevates PI3K/Akt signaling activity in neural crest derivatives, and confirmed that attenuation of PI3K/Akt activity led to decreased neural crest cell proliferation and differentiation both in vitro and in vivo. CONCLUSIONS: Our study revealed that Pten is essential for craniofacial morphogenesis in mice. Inactivating Pten in neural crest cells increases proliferation rate and promotes their differentiation toward osteoblasts. Our data further indicate that Pten acts via modulating PI3K/Akt activity during these processes. Developmental Dynamics 247:304-314, 2018. © 2017 Wiley Periodicals, Inc.

Preliminary assessment on exposure of four typical populations to potentially toxic metals by means of skin wipes under the influence of haze pollution.

To investigate the exposure risk of human beings to nine potentially toxic metals (PTMs), namely, Cu, Cr, Zn, As, Cd, Pb, Ni, Mn, and Co, skin wipe samples were collected from four types of populations, namely, children, undergraduates, security guards, and professional drivers, under different haze pollution levels in Xinxiang, China by using Ghost wipes. The Ghost wipes were quantitatively analyzed by inductively coupled plasma mass spectrometry (ICP-MS) after microwave digestion. Generally, Zn (ND-1350µg/m2 for undergraduates, ND-2660µg/m2 for security guards, ND-2460µg/m2 for children, and ND-2530µg/m2 for professional drivers) showed the highest concentration among the four populations, followed by Cu (0.02-83.4µg/m2 for undergraduates, ND-70.2µg/m2 for security guards, 23.2-487µg/m2 for children, and ND-116µg/m2 for professional drivers). As (ND-5.7µg/m2 for undergraduates, ND-2.3µg/m2 for security guards, ND-21.1µg/m2 for children, and ND-11.0µg/m2 for professional drivers) and Co (ND-6.0µg/m2 for undergraduates, ND-7.9µg/m2 for security guards, ND-13.4µg/m2 for children, and ND-2.1µg/m2 for professional drivers) showed the lowest concentrations in all populations. Remarkable differences were found among the four populations and PTM levels decreased in the following order: children, professional drivers, security guards, and undergraduates. Gender variation was discovered for undergraduates and children. Generally, PTM contamination in skin wipes collected during a light haze pollution level was generally higher than that during a heavy haze pollution level, but PTM contamination was comparable between the two haze pollution levels for children. Non-carcinogenic exposure risks to As, Cd, and Pb for all populations were higher than those for the other six elements but all of them were within the acceptable safety threshold, indicating no apparent non-carcinogenic risk.

Theoretical investigation of the Fe+-catalyzed oxidation of acetylene by N2O.

The gas-phase Fe(+)-mediated oxidation of acetylene by N2O on both sextet and quartet potential energy surfaces (PESs) is theoretically investigated using density functional theory. Geometries and energies of all the stationary points involved in the catalytic reaction are located. For the catalytic cycles, the crucial step is the initial N2O reduction by Fe(+) to form FeO(+), in which a direct O-abstraction mechanism is located on the sextet PES, whereas the quartet pathway favors a N-O insertion mechanism. Spin inversion moves the energy barrier for this process downward to a position below the ground-state entrance channel. The second step of the catalytic cycles involves two mechanisms corresponding to direct hydrogen abstraction and cyclization. The former mechanism accounts for the ethynol formation with the upmost activation barrier below the entrance channel by about 5 kcal/mol. The other mechanism involves a "metallaoxacyclobutene" structure, followed by four possible pathways, i.e., direct dissociation, C-C insertion, C-to-O hydrogen shift, and/or C-to-C hydrogen shift. Among these pathways, strong exothermicities as well as energetically low location of the intermediates suggest oxidation to ketene and carbon monoxide along the C-to-C hydrogen shift pathway is the most favorable. Reduction of the CO loss partner FeCH2(+) by another N2O molecule constitutes the third step of the catalytic cycles, which contains direct abstraction of O from N2O giving OFeCH2(+), intramolecular rearrangement to form Fe(+)-OCH2, and nonreactive dissociation. This reaction is also energetically favored considering the energy acquired from the initial reactants.

Gas-phase reactions of Co+ with ethylamine: a theoretical approach to the reaction mechanisms of transition metal ions with primary amines.

We report herein a comprehensive study of gas-phase reactions of Co(+) with ethylamine using density functional theory. Geometries and energies for all the stationary points involved in the reactions are investigated at the B3LYP/6-311++G(2df,2pd) level. Six different "classical" N and "nonclassical" ethyl-H attached isomers are found for the Co(+)-ethylamine complexes. The classical complexes are much more stable than the nonclassical ones, which have the complexation energies close to Co(+) complexes with small alkanes. Extensive conversions could occur readily between these encounter complexes. All conceivable reaction pathways from each encounter complex to the experimentally observed products are carefully surveyed, and the most likely reaction mechanisms are derived. Activation of the C(alpha)-H bond of ethylamine by Co(+) through both the classical and nonclassical complexes leads to not only the H2 loss but also the hydride abstraction. The loss of ethylene arises from Co(+) insertion into the polar C-N bond in the classical complexes as well as from C(beta)-H activation through the nonclassical methyl-H attached complex of Co(+)- gauche-ethylamine. CH4 only forms via C-C activation from the nonclassical complex with the metal bound to two Hs from the different carbons. Initial N-H insertion is unlikely to be important. It is the reactions of the nonclassical complexes that closely parallel with the Co(+) + alkane reactions. The theoretical work sheds new light on the title reactions and can serve as a theoretical approach to the reaction mechanisms of transition metal ions with primary amines.

Theoretical survey of the potential energy surface of methyl nitrite + Cu+ reaction.

The gas-phase reaction of methyl nitrite with Cu+ has been investigated using density functional theory. The geometries and energies of all the stationary points involved in the reaction have been investigated at the B3LYP/6-311+G(2df,2pd) level. Seven different structures of the encounter complexes could be formed when Cu+ attacking at different electronegative heteroatoms of trans and cis conformational isomers of methyl nitrite, in which the inner oxygen attacks account for the most stable complexes. Extensive conversions could take place for these complexes converting into each other. Various mechanisms leading to the loss of NO and HNO are analyzed in terms of the topology of the potential energy surface. The reaction proceeds exclusively from the inner oxygen attachments, followed by four different mechanisms, i.e., direct dissociation, direct H abstraction, N-O activation, and C-H activation, where the former two provide direct channels for the respective losses of NO and HNO, the third one accounts for both of the losses, and C-H activation is unlikely to be important due to the energetics.