Comparative multi-omic analyses of cardiac mitochondrial stress in three mouse models of frataxin deficiency.

Cardiomyopathy is often fatal in Friedreich ataxia (FA). However, FA hearts maintain adequate function until advanced disease stages, suggesting initial adaptation to the loss of frataxin (FXN). Conditional cardiac knockout mouse models of FXN show transcriptional and metabolic profiles of the mitochondrial integrated stress response (ISRmt), which could play an adaptive role. However, the ISRmt has not been investigated in models with disease-relevant, partial decrease in FXN. We characterized the heart transcriptomes and metabolomes of three mouse models with varying degrees of FXN depletion: YG8-800, KIKO-700 and FXNG127V. Few metabolites were changed in YG8-800 mice, which did not provide a signature of cardiomyopathy or ISRmt; several metabolites were altered in FXNG127V and KIKO-700 hearts. Transcriptional changes were found in all models, but differentially expressed genes consistent with cardiomyopathy and ISRmt were only identified in FXNG127V hearts. However, these changes were surprisingly mild even at advanced age (18 months), despite a severe decrease in FXN levels to 1% of those of wild type. These findings indicate that the mouse heart has low reliance on FXN, highlighting the difficulty in modeling genetically relevant FA cardiomyopathy.

Premature transcription termination at the expanded GAA repeats and aberrant alternative polyadenylation contributes to the Frataxin transcriptional deficit in Friedreich's ataxia.

Frataxin deficiency in Friedreich's ataxia results from transcriptional downregulation of the FXN gene caused by expansion of the intronic trinucleotide guanine-adenine-adenine (GAA) repeats. We used multiple transcriptomic approaches to determine the molecular mechanism of transcription inhibition caused by long GAAs. We uncovered that transcription of FXN in patient cells is prematurely terminated upstream of the expanded repeats leading to the formation of a novel, truncated and stable RNA. This FXN early terminated transcript (FXN-ett) undergoes alternative, non-productive splicing and does not contribute to the synthesis of functional frataxin. The level the FXN-ett RNA directly correlates with the length of the longer of the two expanded GAA tracts. Targeting GAAs with antisense oligonucleotides or excision of the repeats eliminates the transcription impediment, diminishes expression of the aberrant FXN-ett, while increasing levels of FXN mRNA and frataxin. Non-productive transcription may represent a common phenomenon and attractive therapeutic target in diseases caused by repeat-mediated transcription aberrations.

Twinning-assisted dynamic adjustment of grain boundary mobility.

Grain boundary (GB) plasticity dominates the mechanical behaviours of nanocrystalline materials. Under mechanical loading, GB configuration and its local deformation geometry change dynamically with the deformation; the dynamic variation of GB deformability, however, remains largely elusive, especially regarding its relation with the frequently-observed GB-associated deformation twins in nanocrystalline materials. Attention here is focused on the GB dynamics in metallic nanocrystals, by means of well-designed in situ nanomechanical testing integrated with molecular dynamics simulations. GBs with low mobility are found to dynamically adjust their configurations and local deformation geometries via crystallographic twinning, which instantly changes the GB dynamics and enhances the GB mobility. This self-adjust twin-assisted GB dynamics is found common in a wide range of face-centred cubic nanocrystalline metals under different deformation conditions. These findings enrich our understanding of GB-mediated plasticity, especially the dynamic behaviour of GBs, and bear practical implication for developing high performance nanocrystalline materials through interface engineering.

Targeting 3' and 5' untranslated regions with antisense oligonucleotides to stabilize frataxin mRNA and increase protein expression.

Friedreich's ataxia (FRDA) is a severe multisystem disease caused by transcriptional repression induced by expanded GAA repeats located in intron 1 of the Frataxin (FXN) gene encoding frataxin. FRDA results from decreased levels of frataxin; thus, stabilization of the FXN mRNA already present in patient cells represents an attractive and unexplored therapeutic avenue. In this work, we pursued a novel approach based on oligonucleotide-mediated targeting of FXN mRNA ends to extend its half-life and availability as a template for translation. We demonstrated that oligonucleotides designed to bind to FXN 5' or 3' noncoding regions can increase FXN mRNA and protein levels. Simultaneous delivery of oligonucleotides targeting both ends increases efficacy of the treatment. The approach was confirmed in several FRDA fibroblast and induced pluripotent stem cell-derived neuronal progenitor lines. RNA sequencing and single-cell expression analyses confirmed oligonucleotide-mediated FXN mRNA upregulation. Mechanistically, a significant elongation of the FXN mRNA half-life without any changes in chromatin status at the FXN gene was observed upon treatment with end-targeting oligonucleotides, indicating that transcript stabilization is responsible for frataxin upregulation. These results identify a novel approach toward upregulation of steady-state mRNA levels via oligonucleotide-mediated end targeting that may be of significance to any condition resulting from transcription downregulation.

Interlayer Coupling Dependent Discrete H → T' Phase Transition in Lithium Intercalated Bilayer Molybdenum Disulfide.

In this work, the interlayer coupling dependent lithium intercalation induced phase transition in bilayer MoS2 (BL-MoS2) was investigated using an atomic-resolution annual dark-field scanning transmission electron microscope (ADF-STEM). It was revealed that the lithiation induced H → T' phase transition in BL-MoS2 strongly depended on the interlayer twist angle; i.e., the H → T' phase transition occurred in well-stacked H phase BL-MoS2 (with a twist angle of θt = 0°) but not for θt ≠ 0° BL-MoS2. The lithiated BL-MoS2 appeared in homophase stacking, either T'/T' or H/H (locally, no phase transformation) stacking, without any heterophase stacking such as H/T' or T'/H observed. This finding indicated the H → T' phase transition occurred via a domain-by-domain mode rather than layer-by-layer. Up to 15 types of stacking orders were experimentally identified locally in lithiated bilayer T'-MoS2, and the formation mechanism was attributed to the discrete interlayer translation with a unit step of (m/6a, n/6b) (m, n = 0, 1, 2, 3), where a and b were the primitive lattice vectors of T'-MoS2. Our experimental results were further corroborated by ab initio density functional theory (DFT) calculations, where the occurrence of different stacking orders can be quantitatively correlated with the variation of intercalated lithium contents into the BL-MoS2. The present study aids in the understanding of the phase transition mechanisms in atomically thin 2D transition metal dichalcogenides (TMDCs) and will also shed light on the precisely controlled phase engineering of 2D materials for memory applications.

Pattern-Potential-Guided Growth of Textured Macromolecular Films on Graphene/High-Index Copper.

Macromolecular films are crucial functional materials widely used in the fields of mechanics, electronics, optoelectronics, and biology, due to their superior properties of chemical stability, small density, high flexibility, and solution-processing ability. Their electronic and mechanical properties, however, are typically much lower than those of crystalline materials, as the macromolecular films have no long-range structural ordering. The state-of-the-art for producing highly ordered macromolecular films is still facing a great challenge due to the complex interactions between adjacent macromolecules. Here, the growth of textured macromolecular films on a designed graphene/high-index copper (Cu) surface is demonstrated. This successful growth is driven by a patterned potential that originates from the different amounts of charge transfer between the graphene and Cu surfaces with, alternately, terraces and step edges. The textured films exhibit a remarkable improvement in remnant ferroelectric polarization and fracture strength. It is also demonstrated that this growth mechanism is universal for different macromolecules. As meter-scale graphene/high-index Cu substrates have recently become available, the results open a new regime for the production and applications of highly ordered macromolecular films with obvious merits of high production and low cost.

A CRISPR-Cas9, Cre-lox, and Flp-FRT Cascade Strategy for the Precise and Efficient Integration of Exogenous DNA into Cellular Genomes.

We describe a protocol for the precise integration of exogenous DNA into user-defined genomic loci in cultured cells. This strategy first introduces a promoter and a lox site to a specific location via a Cas9-induced double-strand break. Second, a gene of interest (GOI) is inserted into the lox site via Cre-lox recombination. Upon correct insertion, a cis-linked antibiotic resistance gene will be expressed from a promoter introduced into the genome in the first step assuring selection for correct integrants. Last, the selection cassette is excised via a Flp-FRT recombination event, leaving a precisely targeted GOI. This method is broadly applicable to any exogenous DNA to be integrated, choice of integration site, and choice of cell type. The most remarkable aspect of this versatile approach, termed "CasPi" (cascaded precise integration), is that it allows for precise genome targeting with large, frequently complex, and repetitive DNA sequences that do not integrate efficiently or at all with current genome targeting methods.

Defining Transcription Regulatory Elements in the Human Frataxin Gene: Implications for Gene Therapy.

Friedreich's ataxia (FRDA) is the most common inherited form of ataxia in humans. It is caused by severe downregulation of frataxin (FXN) expression instigated by hyperexpansion of the GAA repeats located in intron 1 of the FXN gene. Despite numerous studies focused on identifying compounds capable of stimulating FXN expression, current knowledge regarding cis-regulatory elements involved in FXN gene expression is lacking. Using a combination of episomal and genome-integrated constructs, we defined a minimal endogenous promoter sequence required to efficiently drive FXN expression in human cells. We generated 19 constructs varying in length of the DNA sequences upstream and downstream of the ATG start codon. Using transient transfection, we evaluated the capability of these constructs to drive FXN expression. These analyses allowed us to identify a region of the gene indispensable for FXN expression. Subsequently, selected constructs containing the FXN expression control regions of varying lengths were site specifically integrated into the genome of HEK293T and human-induced pluripotent stem cells (iPSCs). FXN expression was detected in iPSCs and persisted after differentiation to neuronal and cardiac cells, indicating lineage independent function of defined regulatory DNA sequences. Finally, based on these results, we generated AAV encoding miniFXN genes and demonstrated in vivo FXN expression in mice. Results of these studies identified FXN sequences necessary to express FXN in human and mouse cells and provided rationale for potential use of endogenous FXN sequence in gene therapy strategies for FRDA.

Tuning element distribution, structure and properties by composition in high-entropy alloys.

High-entropy alloys are a class of materials that contain five or more elements in near-equiatomic proportions1,2. Their unconventional compositions and chemical structures hold promise for achieving unprecedented combinations of mechanical properties3-8. Rational design of such alloys hinges on an understanding of the composition-structure-property relationships in a near-infinite compositional space9,10. Here we use atomic-resolution chemical mapping to reveal the element distribution of the widely studied face-centred cubic CrMnFeCoNi Cantor alloy2 and of a new face-centred cubic alloy, CrFeCoNiPd. In the Cantor alloy, the distribution of the five constituent elements is relatively random and uniform. By contrast, in the CrFeCoNiPd alloy, in which the palladium atoms have a markedly different atomic size and electronegativity from the other elements, the homogeneity decreases considerably; all five elements tend to show greater aggregation, with a wavelength of incipient concentration waves11,12 as small as 1 to 3 nanometres. The resulting nanoscale alternating tensile and compressive strain fields lead to considerable resistance to dislocation glide. In situ transmission electron microscopy during straining experiments reveals massive dislocation cross-slip from the early stage of plastic deformation, resulting in strong dislocation interactions between multiple slip systems. These deformation mechanisms in the CrFeCoNiPd alloy, which differ markedly from those in the Cantor alloy and other face-centred cubic high-entropy alloys, are promoted by pronounced fluctuations in composition and an increase in stacking-fault energy, leading to higher yield strength without compromising strain hardening and tensile ductility. Mapping atomic-scale element distributions opens opportunities for understanding chemical structures and thus providing a basis for tuning composition and atomic configurations to obtain outstanding mechanical properties.

Excision of the expanded GAA repeats corrects cardiomyopathy phenotypes of iPSC-derived Friedreich's ataxia cardiomyocytes.

Friedreich's ataxia is caused by large homozygous, intronic expansions of GAA repeats in the frataxin (FXN) gene, resulting in severe downregulation of its expression. Pathogenic repeats are located in intron one, hence patients express unaffected FXN protein, albeit in low quantities. Although FRDA symptoms typically afflict the nervous system, hypertrophic cardiomyopathy is the predominant cause of death. Our studies were conducted using cardiomyocytes differentiated from induced pluripotent stem cells derived from control individuals, FRDA patients, and isogenic cells corrected by zinc finger nucleases-mediated excision of pathogenic expanded GAA repeats. This correction of the FXN gene removed the primary trigger of the transcription defect, upregulated frataxin expression, reduced pathological lipid accumulation observed in patient cardiomyocytes, and reversed gene expression signatures of FRDA cardiomyocytes. Transcriptome analyses revealed hypertrophy-specific expression signatures unique to FRDA cardiomyocytes, and emphasized similarities between unaffected and ZFN-corrected FRDA cardiomyocytes. Thus, the iPSC-derived FRDA cardiomyocytes exhibit various molecular defects characteristic for cellular models of cardiomyopathy that can be corrected by genome editing of the expanded GAA repeats. These results underscore the utility of genome editing in generating isogenic cellular models of FRDA and the potential of this approach as a future therapy for this disease.

Engineering Graphene Wrinkles for Large Enhancement of Interlaminar Friction Enabled Damping Capability.

Graphene nanoplates are hoped-for solid lubricants to reduce friction and energy dissipation in micro and nanoscale devices benefiting from their interface slips to reach an expected superlubricity. On the contrary, we propose here by introducing engineered wrinkles of graphene nanoplates to exploit and optimize the interfacial energy dissipation mechanisms between the nanoplates in graphene-based composites for enhanced vibration damping performance. Polyurethane (PU) beams with designed sandwich structures have been successfully fabricated to activate the interlaminar slips of wrinkled graphene-graphene, which significantly contribute to the dissipation of vibration energy. These engineered composite materials with extremely low graphene content (∼0.08 wt %) yield a significant increase in quasi-static and dynamic damping compared to the baseline PU beams (by 71% and 94%, respectively). Friction force images of wrinkled graphene oxide (GO) nanoplates detected via an atomic force microscope (AFM) indicate that wrinkles with large coefficients of friction (COFs) indeed play a dominant role in delaying slip occurrences. Reduction of GO further enhances the COFs of the interacting wrinkles by 7.8%, owing to the increased effective contact area and adhesive force. This work provides a new insight into how to design graphene-based composites with optimized damping properties from the microstructure perspective.

In situ atomistic observation of disconnection-mediated grain boundary migration.

Shear-coupled grain boundary (GB) migration is of general significance in the deformation of nanocrystalline and polycrystalline materials, but comprehensive understanding of the migration mechanism at the atomic scale remains largely lacking. Here, we systematically investigate the atomistic migration of Σ11(113) coherent GBs in gold bicrystals using a state-of-art in situ shear testing technique combined with molecular dynamic simulations. We show that shear-coupled GB migration can be realised by the lateral motion of layer-by-layer nucleated GB disconnections, where both single-layer and double-layer disconnections have important contributions to the GB migration through their frequent composition and decomposition. We further demonstrate that the disconnection-mediated GB migration is fully reversible in shear loading cycles. Such disconnection-mediated GB migration should represent a general deformation phenomenon in GBs with different structures in polycrystalline and nanocrystalline materials, where the triple junctions can act as effective nucleation sites of GB disconnections.

Template-free hydrothermal synthesis of MgWO4 nanoplates and their application as photocatalysts.

As a semiconductor, MgWO4 has a potential in photocatalytic applications; however, it has been overlooked in previous studies; in this study, it has been demonstrated that MgWO4 exhibits the ability to drive photocatalytic hydrogen evolution. Compared to nanoparticle structures, MgWO4 nanoplates show an increased photocatalytic ability due to their higher specific surface area. Moreover, the formation mechanism of MgWO4 nanoplates has been discussed. An oriented attachment and ripening process is proposed for the formation of the MgWO4 nanoplates. This study demonstrates that MgWO4 can be considered a valuable photocatalytic material, and future studies should be focused on how to promote its photocatalytic conversion efficiency.

Towards quantitative mapping of the charge distribution along a nanowire by in-line electron holography.

Mapping the charge distribution in nano scale systems still is a difficult task, but is important to provide fundamental insights into the properties of materials. We demonstrate how in-line holography in transmission electron microscopy can be used to extract the charge distribution in the nanowire in a quantitative way. This technique can realize a fast acquisition of delicate charge variations. By taking advantage of the possibilities of in-situ electron microscopy, variations of the external field can be used to modulate the charge distribution. Because of the fast response to charge variations, this method provides an efficient probing tool for detecting dynamic charge redistribution.

Consecutive crystallographic reorientations and superplasticity in body-centered cubic niobium nanowires.

Plasticity of metallic nanowires is often controlled by the activities of single deformation mode. It remains largely unclear whether multiple deformation modes can be activated in an individual metallic nanowire and how much plasticity they can contribute. In situ nanomechanical testing reveals a superior plastic deformation ability of body-centered cubic (BCC) niobium nanowires, in which a remarkable elongation of more than 269% is achieved before fracture. This superplastic deformation originates from a synergy of consecutively nucleated multiple reorientation processes that occur for more than five times via three distinct mechanisms, that is, stress-activated phase transformation, deformation twinning, and slip-induced crystal rotation. These three coupled mechanisms work concurrently, resulting in sequential reorientations and therefore superplastic deformation of Nb nanowires. Our findings reveal a superior mechanical property of BCC Nb nanowires through the close coordination of multiple deformation modes, which may have some implications in other metallic nanowire systems.

Discrete shear band plasticity through dislocation activities in body-centered cubic tungsten nanowires.

Shear band in metallic crystals is localized deformation with high dislocation density, which is often observed in nanopillar deformation experiments. The shear band dynamics coupled with dislocation activities, however, remains unclear. Here, we investigate the dynamic processes of dislocation and shear band in body-centered cubic (BCC) tungsten nanowires via an integrated approach of in situ nanomechanical testing and atomistic simulation. We find a strong effect of surface orientation on dislocation nucleation in tungsten nanowires, in which {111} surfaces act as favorite sites under high strain. While dislocation activities in a localized region give rise to an initially thin shear band, self-catalyzed stress concentration and dislocation nucleation at shear band interfaces cause a discrete thickening of shear band. Our findings not only advance the current understanding of defect activities and deformation morphology of BCC nanowires, but also shed light on the deformation dynamics in other microscopic crystals where jerky motion of deformation band is observed.

Growth and structural characterisation of Sr-doped Bi2Se3 thin films.

We grew Sr-doped Bi2Se3 thin films using molecular beam epitaxy, and their high quality was verified using transmission electron microscopy. The thin films exhibited weak antilocalisation behaviours in magneto-resistance measurements, a typical transport signature of topological insulators, but were not superconducting. In addition, the carrier densities of the non-superconducting thin-film samples were similar to those of their superconducting bulk counterparts. Atom-by-atom energy-dispersive X-ray mapping also revealed similar Sr doping structures in the bulk and thin-film samples. Because no qualitative distinction between non-superconducting thin-film and superconducting bulk samples had been found, we turned to a quantitative statistical analysis, which uncovered a key structural difference between the bulk and thin-film samples. The separation between Bi layers in the same quintuple layer was compressed whereas that between the closest Bi layers in two neighbouring quintuple layers was expanded in the thin-film samples compared with the separations in pristine bulk Bi2Se3. In marked contrast, the corresponding changes in the bulk doped samples showed opposite trends. These differences may provide insight into the absence of superconductivity in doped topological insulator thin films.

Single-Crystal BiFeO3 Nanoplates with Robust Antiferromagnetism.

Freestanding and single-crystal BiFeO3 (BFO) nanoplates have been successfully synthesized by a fluoride ion-assisted hydrothermal method, and the thickness of the nanoplates can be effectively tailored from 80 to 380 nm by the concentration of fluoride ions. It is revealed that BFO nanoplates grew via an oriented attachment of layer by layer, giving rise to the formation of the inner interface within the nanoplates. In particular, antiferromagnetic (AFM) phase-transition temperature (Néel temperature, TN) of the BFO nanoplates is significantly enhanced from typical 370 to ∼512 °C, whereas the Curie temperature (TC) of the BFO nanoplates is determined to be ∼830 °C, in good agreement with a bulk value. The combination of scanning transmission electron microscopy, electron energy loss spectroscopy, and the first-principle calculations reveals that the interfacial tensile strain remarkably improves the stability of AFM ordering, accounting for the significant enhancement in TN of BFO plates. Correspondingly, the tensile strain induced the polarization and oxygen octahedral tilting has been observed near the interface. The findings presented here suggest that single-crystal BFO nanoplate is an ideal system for exploring an intrinsic magnetoelectric property, where a tensile strain can be a very promising approach to tailor AFM ordering and polarization rotation for an enhanced coupling effect.

Oxygen vacancy regulation on tungsten oxides with specific exposed facets for enhanced visible-light-driven photocatalytic oxidation.

Introduced oxygen vacancy on WO3 with specific exposed facets was prepared through facile solvothermal treatment and different cooling methods. We demonstrated that the density of oxygen defects could be regulated by different cooling methods and speculated that oxygen vacancy with appropriate concentration range could promote photocatalytic activity through suppressing the recombination of photo-induced carriers. The specific exposed facets with higher oxidation efficiency were prepared by solvothermal reaction. WO3-A treated by air cooling exhibits the best photocatalytic oxygen evolution rate at 500 µmol g-1 h-1 using AgNO3 as sacrifice agent under visible light (λ > 400 nm) without any co-catalysts, which is about 2 times higher than WO3-N without oxygen defects. This strategy, using different cooling methods to regulate oxygen vacancy concentration on tungsten oxides, could contribute to the design of other high efficiency photocatalysts.