Long-Lived Binding of Sox2 to DNA Predicts Cell Fate in the Four-Cell Mouse Embryo.

Transcription factor (TF) binding to DNA is fundamental for gene regulation. However, it remains unknown how the dynamics of TF-DNA interactions change during cell-fate determination in vivo. Here, we use photo-activatable FCS to quantify TF-DNA binding in single cells of developing mouse embryos. In blastocysts, the TFs Oct4 and Sox2, which control pluripotency, bind DNA more stably in pluripotent than in extraembryonic cells. By contrast, in the four-cell embryo, Sox2 engages in more long-lived interactions than does Oct4. Sox2 long-lived binding varies between blastomeres and is regulated by H3R26 methylation. Live-cell tracking demonstrates that those blastomeres with more long-lived binding contribute more pluripotent progeny, and reducing H3R26 methylation decreases long-lived binding, Sox2 target expression, and pluripotent cell numbers. Therefore, Sox2-DNA binding predicts mammalian cell fate as early as the four-cell stage. More generally, we reveal the dynamic repartitioning of TFs between DNA sites driven by physiological epigenetic changes. VIDEO ABSTRACT.

Structure of the human ATP synthase.

Biological energy currency ATP is produced by F1Fo-ATP synthase. However, the molecular mechanism for human ATP synthase action remains unknown. Here, we present snapshot images for three main rotational states and one substate of human ATP synthase using cryoelectron microscopy. These structures reveal that the release of ADP occurs when the ß subunit of F1Fo-ATP synthase is in the open conformation, showing how ADP binding is coordinated during synthesis. The accommodation of the symmetry mismatch between F1 and Fo motors is resolved by the torsional flexing of the entire complex, especially the γ subunit, and the rotational substep of the c subunit. Water molecules are identified in the inlet and outlet half-channels, suggesting that the proton transfer in these two half-channels proceed via a Grotthus mechanism. Clinically relevant mutations are mapped to the structure, showing that they are mainly located at the subunit-subunit interfaces, thus causing instability of the complex.

Inhibition of M. tuberculosis and human ATP synthase by BDQ and TBAJ-587.

Bedaquiline (BDQ), a first-in-class diarylquinoline anti-tuberculosis drug, and its analogue, TBAJ-587, prevent the growth and proliferation of Mycobacterium tuberculosis by inhibiting ATP synthase1,2. However, BDQ also inhibits human ATP synthase3. At present, how these compounds interact with either M. tuberculosis ATP synthase or human ATP synthase is unclear. Here we present cryogenic electron microscopy structures of M. tuberculosis ATP synthase with and without BDQ and TBAJ-587 bound, and human ATP synthase bound to BDQ. The two inhibitors interact with subunit a and the c-ring at the leading site, c-only sites and lagging site in M. tuberculosis ATP synthase, showing that BDQ and TBAJ-587 have similar modes of action. The quinolinyl and dimethylamino units of the compounds make extensive contacts with the protein. The structure of human ATP synthase in complex with BDQ reveals that the BDQ-binding site is similar to that observed for the leading site in M. tuberculosis ATP synthase, and that the quinolinyl unit also interacts extensively with the human enzyme. This study will improve researchers' understanding of the similarities and differences between human ATP synthase and M. tuberculosis ATP synthase in terms of the mode of BDQ binding, and will allow the rational design of novel diarylquinolines as anti-tuberculosis drugs.

Author Correction: Highly specific multiplexed RNA imaging in tissues with split-FISH.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Highly specific multiplexed RNA imaging in tissues with split-FISH.

We present split-FISH, a multiplexed fluorescence in situ hybridization method that leverages a split-probe design to achieve enhanced specificity. Split-FISH reduces off-target background fluorescence, decreases false positives and enables accurate RNA profiling in uncleared tissues. We demonstrate the efficacy of split-FISH on various mouse tissues by quantifying the distribution and abundance of 317 genes in single cells and reveal diverse localization patterns for spatial regulation of the transcriptome in complex tissues.

Superresolution imaging reveals spatiotemporal propagation of human replication foci mediated by CTCF-organized chromatin structures.

Mammalian DNA replication is initiated at numerous replication origins, which are clustered into thousands of replication domains (RDs) across the genome. However, it remains unclear whether the replication origins within each RD are activated stochastically or preferentially near certain chromatin features. To understand how DNA replication in single human cells is regulated at the sub-RD level, we directly visualized and quantitatively characterized the spatiotemporal organization, morphology, and in situ epigenetic signatures of individual replication foci (RFi) across S-phase at superresolution using stochastic optical reconstruction microscopy. Importantly, we revealed a hierarchical radial pattern of RFi propagation dynamics that reverses directionality from early to late S-phase and is diminished upon caffeine treatment or CTCF knockdown. Together with simulation and bioinformatic analyses, our findings point to a "CTCF-organized REplication Propagation" (CoREP) model, which suggests a nonrandom selection mechanism for replication activation at the sub-RD level during early S-phase, mediated by CTCF-organized chromatin structures. Collectively, these findings offer critical insights into the key involvement of local epigenetic environment in coordinating DNA replication across the genome and have broad implications for our conceptualization of the role of multiscale chromatin architecture in regulating diverse cell nuclear dynamics in space and time.

Nitrogen Vacancy-Modulated Peroxymonosulfate Nonradical Activation for Organic Contaminant Removal via High-Valent Cobalt-Oxo Species.

Rapid generation of high-valent cobalt-oxo species (Co(IV)═O) for the removal of organic contaminants has been challenging because of the low conversion efficiency of Co(III)/Co(II) and the high activation energy barrier of the Co(II)-oxidant complex. Herein, we introduced nitrogen (N) vacancies into graphite carbon nitride imbedded with cobalt carbonate (CCH/CN-Vn) in a peroxymonosulfate (PMS)/visible light system to break the limitations of a conventional two-electron transfer path. These N vacancies enhanced the electron distribution of the Co 3d orbital and lowered the energy barrier to cleave the O-O bond of PMS in the Co(II)-PMS complex, achieving the modulation of major active species from 1O2 to Co(IV)═O. The developed synergistic system that exhibited adsorption and oxidation showed remarkable selectivity and contaminant removal performance in inorganic (Cl-, NO3-, HCO3-, and HPO4-) organic (HA) and even practical aqueous matrices (tap water and secondary effluent). This study provides a novel mechanistic perspective to modulate the nonradical path for refractory contaminant treatment via defect engineering.

In situ preparation of BiOCl0.75I0.25/g-C3N4-Cl in reduced graphene hydrogel photoanode for simultaneous removal of tetracycline hydrochloride and hexavalent chromium with efficient electricity generation.

A novel three-dimensional porous photoanode of BiOCl0.75I0.25/g-C3N4-Cl/reduced graphene hydrogel (BOCI/CNCl/rGH) was successfully fabricated by a combined in-situ growth and re-dispersion strategy. It was verified that BOCI/CNCl composite exhibited photocatalytic efficiency, and the introduced rGH not only provided superior conductivity which was favorable for charge transfer, but also increased the specific surface area and reactive sites than the fluorine-doped tin oxide (FTO) coated glass. On the basis of these advantages, the short-circuit current and maximum power density were increased by 5.1 and 1.2 times, and the respective removal efficiency of tetracycline hydrochloride (TCH) and hexavalent chromium (Cr(VI)) was increased by 29% and 32% in BOCI/CNCl/rGH, comparing with BOCI/CNCl/FTO. Notably, the removal efficiencies could reach 87% and 85% in TCH and Cr(VI) coexistence system, which were higher than those in TCH or Cr(VI) alone system. This study provides a novel strategy for designing highly efficient photoanode for multiple pollutants removal and electricity generation.

Phase Separation-Mediated Chromatin Organization and Dynamics: From Imaging-Based Quantitative Characterizations to Functional Implications.

As an effective and versatile strategy to compartmentalize cellular components without the need for lipid membranes, phase separation has been found to underpin a wide range of intranuclear processes, particularly those involving chromatin. Many of the unique physico-chemical properties of chromatin-based phase condensates are harnessed by the cell to accomplish complex regulatory functions in a spatially and temporally controlled manner. Here, we survey key recent findings on the mechanistic roles of phase separation in regulating the organization and dynamics of chromatin-based molecular processes across length scales, packing states and intranuclear functions, with a particular emphasis on quantitative characterizations of these condensates enabled by advanced imaging-based approaches. By illuminating the complex interplay between chromatin and various chromatin-interacting molecular species mediated by phase separation, this review sheds light on an emerging multi-scale, multi-modal and multi-faceted landscape that hierarchically regulates the genome within the highly crowded and dynamic nuclear space. Moreover, deficiencies in existing studies also highlight the need for mechanism-specific criteria and multi-parametric approaches for the characterization of chromatin-based phase separation using complementary techniques and call for greater efforts to correlate the quantitative features of these condensates with their functional consequences in close-to-native cellular contexts.

Exogenous hydrogen sulfide inhibits neutrophils extracellular traps formation via the HMGB1/TLR4/p-38 MAPK/ROS axis in hyperhomocysteinemia rats.

Hydrogen sulfide (H2S) prevents platelet activation and neutrophils extracellular traps (NETs) formation. However, the mechanism of sodium hydrosulfide (NaHS, a donor that produces H2S) inhibits the formation of NETs in hyperhomocysteinemia (HHcy) rats has not been previously investigated. In the experiment, the expressions of HMGB1 of platelets, the expressions of TLR4, PAD4 and the phosphor-p38 of neutrophils were measured. The NETs formations, the concentration of DNA in the serum and the culture solution of cultured neutrophils which was stimulated by platelet-rich plasma (PRP) were tested. Additionally, the cellular ROS level and SOD activity were detected. The platelets were activated and the expression of HMGB1 of platelets and NETs formation, the concentration of DNA, and the expressions of TLR4, phosphor-p38 and PAD4, the ROS level were all increased while the activity of SOD decreased in the HHcy group compared to the control group. NaHS significantly inhibited the activation of platelets, the production of ROS and the formation of NETs in neutrophils, reversed the expressions of HMGB1, TLR4, phosphor-p38, PAD4 and decreased concentration of DNA which was caused by high homocysteine. Our results demonstrate that the donor of H2S inhibits NETs formation of neutrophils via the HMGB1/TLR4/p38 MAPK/ROS pathway in hyperhomocysteinemia.

Exogenous hydrogen sulfide protects from endothelial cell damage, platelet activation, and neutrophils extracellular traps formation in hyperhomocysteinemia rats.

Hydrogen sulfide (H2S) prevents endothelial cells damage and P-selectin of platelets promotes neutrophils extracellular traps (NETs) formation. However, how sodium hydrosulfide (NaHS), a donor that produces H2S regulates the activation of platelets and whether H2S inhibits the formation of neutrophils extracellular traps in hyperhomocysteinemia rats have not been previously investigated. The morphological and ultrastructural alterations of endothelial cells (ECs) and platelets were tested by transmission electron microscopy. The expressions of P-selectin of platelets were determined by flow cytometry. Additionally, the cellular ROS and the H2S level were detected by DCFH-DA staining and H2S probe, the expressions of Bax and Bcl-2 in arteries and cultured ECs from rat thoracic aortas and the phosphor-p38 mitogen-activated protein kinase (MAPK), CSE and CBS of platelets were measured by western blotting. The NETs formations, the concentration of DNA in serum and supernatant of cultured neutrophils stimulated with platelet-rich plasma (PRP) were tested by Sytox Green and PicoGreen commercial Kits. The vascular ECs damaged, the expression of P-selectin of platelets and NETs formation increased; the concentration of DNA in serum and supernatant of cultured neutrophils stimulated with PRP also increased; the expression of Bax increased while Bcl-2 decreased in arteries, the phosphor-p38 MAPK of platelets increased while CSE and CBS have no statistically significant changes in the HHcy group compared to the control group. In the cultured ECs, the ROS level increased while the H2S level decreased after 48 and 72 h treatment by HHcy; the expression of Bcl-2 decreased while Bax increased after 72 h treatment by HHcy. NaHS significantly inhibited the ECs injured, cellular ROS production, platelet activation and NETs formation, reversed the expressions of Bax, Bcl-2, phosphor-p38 MAPK, P-selectin and the increased concentration of DNA in serum and supernatant of cultured neutrophils which caused by high homocysteine. Our results demonstrate that the donor of H2S inhibits the platelets activation and NETs formation, which concerts the protection of ECs in hyperhomocysteinemia.

Calcium sensing receptor initiating cystathionine-gamma-lyase/hydrogen sulfide pathway to inhibit platelet activation in hyperhomocysteinemia rat.

Hyperhomocysteinemia (HHcy, high homocysteine) induces the injury of endothelial cells (ECs). Hydrogen sulfide (H2S) protects ECs and inhibits the activation of platelets. Calcium-sensing receptor (CaSR) regulates the production of endogenous H2S. However, whether CaSR inhibits the injury of ECs and the activation of platelets by regulating the endogenous cystathionine-gamma-lyase (CSE, a major enzyme that produces H2S)/H2S pathway in hyperhomocysteinemia has not been previously investigated. Here, we tested the ultrastructure alterations of ECs and platelets, the changes in the concentration of serum homocysteine and the parameters of blood of hyperhomocysteinemia rats were measured. The aggregation rate and expression of P-selectin of platelets were assessed. Additionally, the expression levels of CaSR and CSE in the aorta of rats were examined by western blotting. The mitochondrial membrane potential and the production of reactive oxygen species (ROS) were measured; the expression of phospho-calmodulin kinases II (p-CaMK II) and Von Willebrand Factor (vWF) of cultured ECs from rat thoracic aortas were measured. We found that the aggregation rate and the expression of P-selectin of platelets increased, and the expression of CaSR and CSE decreased in HHcy rats. In the ECs of HHcy group, the ROS production increased and the mitochondrial membrane potential decreased markedly, the expression of CSE and the p-CaMK II increased after treatment with CaSR agonist while decreased upon administration of U73122 (PLC-specific inhibitor) and 2-APB (IP3 Receptor inhibitor). CaSR agonist or NaHS significantly reversed the ECs injured and platelet aggregation caused by hyperhomocysteinemia. Our results demonstrate that CaSR regulates the endogenous CSE/H2S pathway to inhibit the activation of platelets which concerts the protection of ECs in hyperhomocysteinemia.

Spatial organization of RNA polymerase II inside a mammalian cell nucleus revealed by reflected light-sheet superresolution microscopy.

Superresolution microscopy based on single-molecule centroid determination has been widely applied to cellular imaging in recent years. However, quantitative imaging of the mammalian nucleus has been challenging due to the lack of 3D optical sectioning methods for normal-sized cells, as well as the inability to accurately count the absolute copy numbers of biomolecules in highly dense structures. Here we report a reflected light-sheet superresolution microscopy method capable of imaging inside the mammalian nucleus with superior signal-to-background ratio as well as molecular counting with single-copy accuracy. Using reflected light-sheet superresolution microscopy, we probed the spatial organization of transcription by RNA polymerase II (RNAP II) molecules and quantified their global extent of clustering inside the mammalian nucleus. Spatiotemporal clustering analysis that leverages on the blinking photophysics of specific organic dyes showed that the majority (>70%) of the transcription foci originate from single RNAP II molecules, and no significant clustering between RNAP II molecules was detected within the length scale of the reported diameter of "transcription factories." Colocalization measurements of RNAP II molecules equally labeled by two spectrally distinct dyes confirmed the primarily unclustered distribution, arguing against a prevalent existence of transcription factories in the mammalian nucleus as previously proposed. The methods developed in our study pave the way for quantitative mapping and stoichiometric characterization of key biomolecular species deep inside mammalian cells.

Quantitative imaging of mammalian transcriptional dynamics: from single cells to whole embryos.

Probing dynamic processes occurring within the cell nucleus at the quantitative level has long been a challenge in mammalian biology. Advances in bio-imaging techniques over the past decade have enabled us to directly visualize nuclear processes in situ with unprecedented spatial and temporal resolution and single-molecule sensitivity. Here, using transcription as our primary focus, we survey recent imaging studies that specifically emphasize the quantitative understanding of nuclear dynamics in both time and space. These analyses not only inform on previously hidden physical parameters and mechanistic details, but also reveal a hierarchical organizational landscape for coordinating a wide range of transcriptional processes shared by mammalian systems of varying complexity, from single cells to whole embryos.

Single-molecule imaging of transcription factor binding to DNA in live mammalian cells.

Imaging single fluorescent proteins in living mammalian cells is challenged by out-of-focus fluorescence excitation. To reduce out-of-focus fluorescence we developed reflected light-sheet microscopy (RLSM), a fluorescence microscopy method allowing selective plane illumination throughout the nuclei of living mammalian cells. A thin light sheet parallel to the imaging plane and close to the sample surface is generated by reflecting an elliptical laser beam incident from the top by 90° with a small mirror. The thin light sheet allows for an increased signal-to-background ratio superior to that in previous illumination schemes and enables imaging of single fluorescent proteins with up to 100-Hz time resolution. We demonstrated the single-molecule sensitivity of RLSM by measuring the DNA-bound fraction of glucocorticoid receptor (GR) and determining the residence times on DNA of various oligomerization states and mutants of GR and estrogen receptor-α (ER), which permitted us to resolve different modes of DNA binding of GR. We demonstrated two-color single-molecule imaging by observing the spatiotemporal colocalization of two different protein pairs. Our single-molecule measurements and statistical analysis revealed dynamic properties of transcription factors.

Microplastics release from face masks: Characteristics, influential factors, and potential risks.

Since the COVID-19 pandemic, face masks have been used popularly and disposed of improperly, leading to the generation of a large amount of microplastics. The objective of this review is to provide a comprehensive insight into the characteristics of mask-derived microplastics, the influential factors of microplastics release, and the potential risks of these microplastics to the environment and organisms. Mask-derived microplastics were predominantly transparent fibers, with a length of <1 mm. The release of microplastics from masks is mainly influenced by mask types, use habits, and weathering conditions. Under the same conditions, surgical masks release more microplastics than other types of masks. Long-term wearing of masks and the disinfection for reuse can promote the release of microplastics. Environmental media, UV irradiation, temperature, pH value, and mechanical shear can also influence the microplastics release. The risks of mask-derived microplastics to human health via inhalation cannot be neglected. Future studies should pay more attention to the release of microplastics from the masks with alternative materials and under more weathering conditions.

Social support and posttraumatic growth: A meta-analysis.

BACKGROUND: The beneficial role of social support on posttraumatic growth (PTG) has been assumed by theoretical models and established in some studies. However, there are inconsistent findings and little knowledge on moderators. The present study aims to investigate the overall effect size of the relationship and identify factors affecting the association. METHODS: Six electronic databases were searched. Newcastle-Ottawa Quality Assessment Scale (NOS) were used to evaluate the quality of studies. Study quality, study design, trauma type, PTG measure, social support measure, continent, publishing language, sample size, gender, religion, and age were analyzed as moderators. Meta-regression was conducted with the significant differential predictors in moderator analysis. RESULTS: The meta-analysis included 217 samples and a total of 47,940 participants from both longitudinal and cross-sectional studies. There was a medium positive effect size between social support and PTG in random effect model, r = 0.418, p < .001. The meta-regression analysis indicated that the association between social support and PTG was stronger among caregivers (vs. other traumatized samples), Chinese, older individuals and studies with smaller sample size. LIMITATIONS: Only survey results were included in the analysis. The retrospective self-report may limit a more objective assessment of the relations. In addition, 87 % of the studies were cross-sectional, which may influence the estimation of a valid effect size. CONCLUSIONS: Regarding the medium positive association between social support and PTG, it is important to enhance social support for trauma survivors. It will be especially effective for caregivers, Chinese, and older people.

Solid pseudopapillary neoplasm: Report of a case of primary ovarian origin and review of the literature.

Background: Solid pseudopapillary neoplasms (SPNs) are uncommon tumors of low malignancy with a generally favorable prognosis, mostly originating from the pancreas. To date, 12 cases of SPNs with a primary ovarian origin (SPN-Os) have been reported globally, and their detailed characteristics have not been fully elucidated. Case description: We reported the 13th SPN-O case, which occurred in a 52-year-old woman with an 18.5 cm left ovarian mass. Four imaging methods, including ultrasound, computed tomography, magnetic resonance imaging and positron emission tomography, were utilized before surgery. An elevated level of serum cancer antigen 125 was detected and a total hysterectomy plus bilateral salpingo-oophorectomy was performed. Microscopic examination revealed a typical solid pseudopapillary structure. The tumor cells were stained focally for pan-cytokeratin, synaptophysin, CD99 and CD10, while ß-catenin, vimentin and CD56 were diffusely expressed. The Ki-67 proliferation index was 3%, and immunohistochemical (IHC) staining for chromogranin-A, inhibin-a, and E-cadherin was negative. No evidence of recurrence or metastasis was observed by clinical and imaging data during a 5-month postoperative follow-up. Conclusion: This is a report of an unusual case of a primary ovarian SPN with an up-to-date review of SPN-Os. A minimum combination of imaging methods and IHC stains was proposed for SPN-Os, which may prove beneficial in clinical practice.

Technology challenges among deaf and hard of hearing elders in China during COVID-19 pandemic emergency isolation: A qualitative study.

Digital technology can be an effective tool to facilitate emergency assistance in a pandemic, but many deaf and hard-of-hearing elders may experience challenges in using and adopting these technologies. In the context of the second wave of the COVID-19 outbreak, this study employs a qualitative research method based on in-depth interviews to explore technology challenges among deaf and hard-of-hearing elders in China. The results showed that this group's technology challenges arose mainly from barriers to the mastery of digital technology tools, among which barriers to the use of smartphones, to the accessibility of online medical consultations, and to the presentation of health codes, were most noteworthy. For the informants, these barriers led to social isolation and technology avoidance. What's more, the expectation of individuals to adopt certain types of digital intelligence technologies can inadvertently create inequities for disadvantaged groups and exacerbate the "digital divide." This study highlights the need for emergency management systems to be inclusive of elders with hearing loss in times of public health crises, by providing effective technology support and training to facilitate individuals' access to services and to safeguard their health, interests, and livelihood.