Gasdermin D permeabilization of mitochondrial inner and outer membranes accelerates and enhances pyroptosis.

Gasdermin D (GSDMD)-activated inflammatory cell death (pyroptosis) causes mitochondrial damage, but its underlying mechanism and functional consequences are largely unknown. Here, we show that the N-terminal pore-forming GSDMD fragment (GSDMD-NT) rapidly damaged both inner and outer mitochondrial membranes (OMMs) leading to reduced mitochondrial numbers, mitophagy, ROS, loss of transmembrane potential, attenuated oxidative phosphorylation (OXPHOS), and release of mitochondrial proteins and DNA from the matrix and intermembrane space. Mitochondrial damage occurred as soon as GSDMD was cleaved prior to plasma membrane damage. Mitochondrial damage was independent of the B-cell lymphoma 2 family and depended on GSDMD-NT binding to cardiolipin. Canonical and noncanonical inflammasome activation of mitochondrial damage, pyroptosis, and inflammatory cytokine release were suppressed by genetic ablation of cardiolipin synthase (Crls1) or the scramblase (Plscr3) that transfers cardiolipin to the OMM. Phospholipid scramblase-3 (PLSCR3) deficiency in a tumor compromised pyroptosis-triggered anti-tumor immunity. Thus, mitochondrial damage plays a critical role in pyroptosis.

The NK cell receptor NKp46 recognizes ecto-calreticulin on ER-stressed cells.

Natural killer (NK) cell kill infected, transformed and stressed cells when an activating NK cell receptor is triggered1. Most NK cells and some innate lymphoid cells express the activating receptor NKp46, encoded by NCR1, the most evolutionarily ancient NK cell receptor2,3. Blockage of NKp46 inhibits NK killing of many cancer targets4. Although a few infectious NKp46 ligands have been identified, the endogenous NKp46 cell surface ligand is unknown. Here we show that NKp46 recognizes externalized calreticulin (ecto-CRT), which translocates from the endoplasmic reticulum (ER) to the cell membrane during ER stress. ER stress and ecto-CRT are hallmarks of chemotherapy-induced immunogenic cell death5,6, flavivirus infection and senescence. NKp46 recognition of the P domain of ecto-CRT triggers NK cell signalling and NKp46 caps with ecto-CRT in NK immune synapses. NKp46-mediated killing is inhibited by knockout or knockdown of CALR, the gene encoding CRT, or CRT antibodies, and is enhanced by ectopic expression of glycosylphosphatidylinositol-anchored CRT. NCR1)-deficient human (and Nrc1-deficient mouse) NK cells are impaired in the killing of ZIKV-infected, ER-stressed and senescent cells and ecto-CRT-expressing cancer cells. Importantly, NKp46 recognition of ecto-CRT controls mouse B16 melanoma and RAS-driven lung cancers and enhances tumour-infiltrating NK cell degranulation and cytokine secretion. Thus, NKp46 recognition of ecto-CRT as a danger-associated molecular pattern eliminates ER-stressed cells.

Excessive apoptosis of Rip1-deficient T cells leads to premature aging.

In mammals, the most remarkable T cell variations with aging are the shrinking of the naïve T cell pool and the enlargement of the memory T cell pool, which are partially caused by thymic involution. However, the mechanism underlying the relationship between T-cell changes and aging remains unclear. In this study, we find that T-cell-specific Rip1 KO mice show similar age-related T cell changes and exhibit signs of accelerated aging-like phenotypes, including inflammation, multiple age-related diseases, and a shorter lifespan. Mechanistically, Rip1-deficient T cells undergo excessive apoptosis and promote chronic inflammation. Consistent with this, blocking apoptosis by co-deletion of Fadd in Rip1-deficient T cells significantly rescues lymphopenia, the imbalance between naïve and memory T cells, and aging-like phenotypes, and prolongs life span in T-cell-specific Rip1 KO mice. These results suggest that the reduction and hyperactivation of T cells can have a significant impact on organismal health and lifespan, underscoring the importance of maintaining T cell homeostasis for healthy aging and prevention or treatment of age-related diseases.

Protracted Indian monsoon droughts of the past millennium and their societal impacts.

Protracted droughts lasting years to decades constitute severe threats to human welfare across the Indian subcontinent. Such events are, however, rare during the instrumental period (ca. since 1871 CE). In contrast, the historic documentary evidence indicates the repeated occurrences of protracted droughts in the region during the preinstrumental period implying that either the instrumental observations underestimate the full spectrum of monsoon variability or the historic accounts overestimate the severity and duration of the past droughts. Here we present a temporally precise speleothem-based oxygen isotope reconstruction of the Indian summer monsoon precipitation variability from Mawmluh cave located in northeast India. Our data reveal that protracted droughts, embedded within multidecadal intervals of reduced monsoon rainfall, frequently occurred over the past millennium. These extreme events are in striking temporal synchrony with the historically documented droughts, famines, mass mortality events, and geopolitical changes in the Indian subcontinent. Our findings necessitate reconsideration of the region's current water resources, sustainability, and mitigation policies that discount the possibility of protracted droughts in the future.

Lipopolysaccharide and Glycolipoprotein Coordinately Triggered Necroptosis Contributes to the Pathogenesis of Leptospira Infection in Mice.

Leptospirosis is a recurring but neglected zoonotic disease caused by pathogenic Leptospira. The explicit underlying mechanism of necroptosis and its role in Leptospira infection have not yet been elucidated. Here we reported that leptospiral pathogen-associated molecular patterns, lipopolysaccharide, and glycolipoprotein activate the necroptotic RIPK1-RIPK3-MLKL cascade through the TLR4 signaling pathway in mouse macrophages. Using the murine acute leptospirosis model, we reveal that abolition of necroptosis exhibited significantly improved outcomes in acute phases, with enhanced eradication of Leptospira from liver, mild clinical symptoms, and decreased cytokine production. RIPK3 was also found to exert a necroptosis-independent function in CXCL1 production and neutrophil recruitment, with the consequence of improved Leptospira control. These findings improve our understanding of the mechanism of Leptospira-macrophage interactions, indicating potential therapeutic values by targeting necroptosis signaling pathways.

MiR-342 controls Mycobacterium tuberculosis susceptibility by modulating inflammation and cell death.

Tuberculosis (TB) is an infectious disease caused by Mycobacterium tuberculosis (Mtb) that places a heavy strain on public health. Host susceptibility to Mtb is modulated by macrophages, which regulate the balance between cell apoptosis and necrosis. However, the role of molecular switches that modulate apoptosis and necrosis during Mtb infection remains unclear. Here, we show that Mtb-susceptible mice and TB patients have relatively low miR-342-3p expression, while mice with miR-342-3p overexpression are more resistant to Mtb. We demonstrate that the miR-342-3p/SOCS6 axis regulates anti-Mtb immunity by increasing the production of inflammatory cytokines and chemokines. Most importantly, the miR-342-3p/SOCS6 axis participates in the switching between Mtb-induced apoptosis and necrosis through A20-mediated K48-linked ubiquitination and RIPK3 degradation. Our findings reveal several strategies by which the host innate immune system controls intracellular Mtb growth via the miRNA-mRNA network and pave the way for host-directed therapies targeting these pathways.

Structure-Based Discovery of a Novel Allosteric Inhibitor against Human Dopamine Transporter.

Human dopamine transporter (hDAT) regulates the reuptake of extracellular dopamine (DA) and is an essential therapeutic target for central nervous system (CNS) diseases. The allosteric modulation of hDAT has been identified for decades. However, the molecular mechanism underlying the transportation is still elusive, which hinders the rational design of allosteric modulators against hDAT. Here, a systematic structure-based method was performed to explore allosteric sites on hDAT in inward-open (IO) conformation and to screen compounds with allosteric affinity. First, the model of the hDAT structure was constructed based on the recently reported Cryo-EM structure of the human serotonin transporter (hSERT) and Gaussian-accelerated molecular dynamics (GaMD) simulation was further utilized for the identification of intermediate energetic stable states of the transporter. Then, with the potential druggable allosteric site on hDAT in IO conformation, virtual screening of seven enamine chemical libraries (∼440,000 compounds) was processed, resulting in 10 compounds being purchased for in vitro assay and with Z1078601926 discovered to allosterically inhibit hDAT (IC50 = 0.527 [0.284; 0.988] µM) when nomifensine was introduced as an orthosteric ligand. Finally, the synergistic effect underlying the allosteric inhibition of hDAT by Z1078601926 and nomifensine was explored using additional GaMD simulation and postbinding free energy analysis. The hit compound discovered in this work not only provides a good starting point for lead optimization but also demonstrates the usability of the method for the structure-based discovery of novel allosteric modulators of other therapeutic targets.

Timing and structure of the Younger Dryas event and its underlying climate dynamics.

The Younger Dryas (YD), arguably the most widely studied millennial-scale extreme climate event, was characterized by diverse hydroclimate shifts globally and severe cooling at high northern latitudes that abruptly punctuated the warming trend from the last glacial to the present interglacial. To date, a precise understanding of its trigger, propagation, and termination remains elusive. Here, we present speleothem oxygen-isotope data that, in concert with other proxy records, allow us to quantify the timing of the YD onset and termination at an unprecedented subcentennial temporal precision across the North Atlantic, Asian Monsoon-Westerlies, and South American Monsoon regions. Our analysis suggests that the onsets of YD in the North Atlantic (12,870 ± 30 B.P.) and the Asian Monsoon-Westerlies region are essentially synchronous within a few decades and lead the onset in Antarctica, implying a north-to-south climate signal propagation via both atmospheric (decadal-time scale) and oceanic (centennial-time scale) processes, similar to the Dansgaard-Oeschger events during the last glacial period. In contrast, the YD termination may have started first in Antarctica at ∼11,900 B.P., or perhaps even earlier in the western tropical Pacific, followed by the North Atlantic between ∼11,700 ± 40 and 11,610 ± 40 B.P. These observations suggest that the initial YD termination might have originated in the Southern Hemisphere and/or the tropical Pacific, indicating a Southern Hemisphere/tropics to North Atlantic-Asian Monsoon-Westerlies directionality of climatic recovery.

Influencing Factors and Evaluation of the Self-Healing Behavior of Asphalt Binder Using Molecular Dynamics Simulation Method.

In order to investigate the self-healing behavior of asphalt binder at the molecule scale, the self-healing models of neat and aged asphalt binder with different damage degrees were established by introducing a vacuum pad between two layers filled with asphalt molecules. With this model, the self-healing process was simulated at various healing conditions to reveal the effects of oxidative aging, damage degree and healing temperature on the self-healing property. In addition, self-healing efficiency was evaluated using the indexes representative of the characteristics of different self-healing stages. Our results show that the oxidative aging weakened the stacked structure of the asphalt binder and increased the healing activation energy barrier. The increasing damage degree extended the distance for particles to travel, thus prolonging the time required for the crack interfaces contacting with each other. The elevated temperature improved the molecular mobility by supplying more energy to the molecular system. Furthermore, the self-healing process was evaluated quantitatively by the density variation at the crack closing stage and the diffusion coefficient at the intrinsic healing stage. The duration of each stage was influenced by the oxidative aging, damage degree and healing temperature. The findings in this paper are helpful to reveal and evaluate the self-healing property of asphalt binder.

Potent Human Single-Domain Antibodies Specific for a Novel Prefusion Epitope of Respiratory Syncytial Virus F Glycoprotein.

Respiratory syncytial virus (RSV) poses great health threats to humans. However, there are no licensed vaccines or therapeutic drugs to date. Only one humanized monoclonal antibody, palivizumab, is available on the market, but it is used prophylactically and is limited to infants with high risk. With advances in antibody engineering, it has been found that a single-domain antibody (sdAb) can be therapeutically administered by inhalation, which would be more efficient for respiratory diseases. Here, we identified two human sdAbs, m17 and m35, by phage display technology. They specifically bind to RSV fusion glycoprotein (F protein) in the prefusion state with subnanomolar affinity and potently neutralize both RSV subtypes A and B with 50% inhibitory concentration (IC50) values ranging from pM to nM. Interestingly, these sdAbs recognize a novel epitope, termed VI, that is unique to the prefusion state. This epitope is located at the C terminus of the F1 subunit, close to the viral membrane, and might be sterically restricted. We further find that m17 and m35 neutralize RSV by preventing the prefusion F conformational arrangement, thus inhibiting membrane fusion. These two sdAbs have the potential to be further developed as therapeutic candidates and may also provide novel insight for developing other antiviral reagents against RSV. IMPORTANCE Because respiratory syncytial virus (RSV) can cause serious respiratory disease in immunodeficient groups, including infants and seniors, the development of vaccines and therapeutic drugs, such as neutralizing antibodies, is urgently needed. Compared to the conventional full-length antibody, a single-domain antibody (sdAb) has been demonstrated to be efficient for respiratory diseases when administered by inhalation, thereby potentially introducing a kind of novel therapeutic agent in the market. Here, we discovered two potent neutralizing human sdAbs against RSV that recognized a novel prefusion epitope, termed VI, and prevented conformational arrangement during the fusion process. Our work provides not only therapeutic candidates but also novel targets for new drug and vaccine development.

The Asian monsoon over the past 640,000 years and ice age terminations.

Oxygen isotope records from Chinese caves characterize changes in both the Asian monsoon and global climate. Here, using our new speleothem data, we extend the Chinese record to cover the full uranium/thorium dating range, that is, the past 640,000 years. The record's length and temporal precision allow us to test the idea that insolation changes caused by the Earth's precession drove the terminations of each of the last seven ice ages as well as the millennia-long intervals of reduced monsoon rainfall associated with each of the terminations. On the basis of our record's timing, the terminations are separated by four or five precession cycles, supporting the idea that the '100,000-year' ice age cycle is an average of discrete numbers of precession cycles. Furthermore, the suborbital component of monsoon rainfall variability exhibits power in both the precession and obliquity bands, and is nearly in anti-phase with summer boreal insolation. These observations indicate that insolation, in part, sets the pace of the occurrence of millennial-scale events, including those associated with terminations and 'unfinished terminations'.

Rotation angle measurement method based on self-mixing interference of a fiber laser.

A rotation angle measurement method based on self-mixing interference (SMI) of a fiber laser is proposed. The rotation angle can be calculated indirectly by the displacement measured by SMI. In the experiment, a linear cavity fiber laser with simple structure and high flexibility is used as the optical source for measuring the deflection angle. To improve the measurement accuracy, the SMI signal is filtered by the variational mode decomposition (VMD) algorithm. The filtered SMI signal is normalized by Hilbert transform. The even-power algorithm is used to subdivide the interference fringes, so as to improve the measurement resolution. The experimental result shows that the measurement error of angular shift is less than 1% in the range of 10°.

Identification of new antiviral agents against Kaposi's sarcoma-associated herpesvirus (KSHV) by high-throughput drug screening reveals the role of histamine-related signaling in promoting viral lytic reactivation.

Kaposi's sarcoma-associated herpesvirus (KSHV) causes several human cancers, such as Kaposi's sarcoma (KS) and primary effusion lymphoma (PEL). Current treatment options for KSHV infection and virus associated diseases are sometimes ineffective, therefore, more effectively antiviral agents are urgently needed. As a herpesvirus, lytic replication is critical for KSHV pathogenesis and oncogenesis. In this study, we have established a high-throughput screening assay by using an inducible KSHV+ cell-line, iSLK.219. After screening a compound library that consisted of 1280 Food and Drug Administration (FDA)-approved drugs, 15 hit compounds that effectively inhibited KSHV virion production were identified, most of which have never been reported with anti-KSHV activities. Interestingly, 3 of these drugs target histamine receptors or signaling. Our data further confirmed that antagonists targeting different histamine receptors (HxRs) displayed excellent inhibitory effects on KSHV lytic replication from induced iSLK.219 or BCBL-1 cells. In contrast, histamine and specific agonists of HxRs promoted viral lytic replication from induced iSLK.219 or KSHV-infected primary cells. Mechanistic studies indicated that downstream MAPK and PI3K/Akt signaling pathways were required for histamine/receptors mediated promotion of KSHV lytic replication. Direct knockdown of HxRs in iSLK.219 cells effectively blocked viral lytic gene expression during induction. Using samples from a cohort of HIV+ patients, we found that the KSHV+ group has much higher levels of histamine in their plasma and saliva than the KSHV- group. Taken together, our data have identified new anti-KSHV agents and provided novel insights into the molecular bases of host factors that contribute to lytic replication and reactivation of this oncogenic herpesvirus.

Structure-Based Discovery of Novel Nonpeptide Inhibitors Targeting SARS-CoV-2 Mpro.

The continual spread of novel coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), posing a severe threat to the health worldwide. The main protease (Mpro, alias 3CLpro) of SARS-CoV-2 is a crucial enzyme for the maturation of viral particles and is a very attractive target for designing drugs to treat COVID-19. Here, we propose a multiple conformation-based virtual screening strategy to discover inhibitors that can target SARS-CoV-2 Mpro. Based on this strategy, nine Mpro structures and a protein mimetics library with 8960 commercially available compounds were prepared to carry out ensemble docking for the first time. Five of the nine structures are apo forms presented in different conformations, whereas the other four structures are holo forms complexed with different ligands. The surface plasmon resonance assay revealed that 6 out of 49 compounds had the ability to bind to SARS-CoV-2 Mpro. The fluorescence resonance energy transfer experiment showed that the biochemical half-maximal inhibitory concentration (IC50) values of the six compounds could hamper Mpro activities ranged from 0.69 ± 0.05 to 2.05 ± 0.92 µM. Evaluation of antiviral activity using the cell-based assay indicated that two compounds (Z1244904919 and Z1759961356) could strongly inhibit the cytopathic effect and reduce replication of the living virus in Vero E6 cells with the half-maximal effective concentrations (EC50) of 4.98 ± 1.83 and 8.52 ± 0.92 µM, respectively. The mechanism of the action for the two inhibitors were further elucidated at the molecular level by molecular dynamics simulation and subsequent binding free energy analysis. As a result, the discovered noncovalent reversible inhibitors with novel scaffolds are promising antiviral drug candidates, which may be used to develop the treatment of COVID-19.

Pollutant source, ecological and human health risks assessment of heavy metals in soils from coal mining areas in Xinjiang, China.

This study aims to analyze the pollution characteristics and sources of heavy metal elements for the first time in the Zhundong mining area in Xinjiang using the linear regression model. Additionaly, the health risks with their probability and infleuencing factors on different groups of people's were also evaluated using Monte Carlo (MC) simulation approach. The results shows that 89.28% of Hg was from coal combustion, 40.28% of Pb was from transportation, and 19.54% of As was from atmospheric dust. The main source of Cu and Cr was coal dust, Hg has the greatest impact on potential ecological risks. which accounted for 60.2% and 81.46% of the Cu and Cr content in soil, respectively. The all samples taken from Pb have been Extremely polluted (100%). 93.3% samples taken from As have been Extremely polluted. The overall potential ecological risk was moderate. Adults experienced higher non-carcinogenic risks of heavy metals from their diets than children. Interestingly, body weight was the main factor affecting the adult's health risks. This research provides more comprehensive information for better soil management, soil remediation, and soil pollution control in the Xinjiang mining areas.

South American monsoon response to iceberg discharge in the North Atlantic.

Heinrich Stadials significantly affected tropical precipitation through changes in the interhemispheric temperature gradient as a result of abrupt cooling in the North Atlantic. Here, we focus on changes in South American monsoon precipitation during Heinrich Stadials using a suite of speleothem records covering the last 85 ky B.P. from eastern South America. We document the response of South American monsoon precipitation to episodes of extensive iceberg discharge, which is distinct from the response to the cooling episodes that precede the main phase of ice-rafted detritus deposition. Our results demonstrate that iceberg discharge in the western subtropical North Atlantic led to an abrupt increase in monsoon precipitation over eastern South America. Our findings of an enhanced Southern Hemisphere monsoon, coeval with the iceberg discharge into the North Atlantic, are consistent with the observed abrupt increase in atmospheric methane concentrations during Heinrich Stadials.

A Comparison of Pinyin Invented Spelling and Oddity Test in Measuring Phonological Awareness in L2 Learners of Chinese.

The importance of phonological awareness (PA) for the acquisition of literacy skills has been widely recognized. Across languages, PA is commonly examined using the Oddity test, however, for Chinese-speaking children, Pinyin invented spelling is recommended as being a more powerful tool to assess PA. However, it is still unclear whether this holds true for learners of Chinese as a second language (CSL). To address this issue, we administered an oddity test and a Pinyin invented spelling task to explore Chinese PA in 43 Arabic and 40 English CSL learners at pre-intermediate and intermediate levels. The results generated two major findings. First, Pinyin invented spelling revealed more significant cross-group differences in Chinese PA between the Arabic and English CSL learners than the oddity test. Second, the participants' performance in Pinyin invented spelling was a stronger predictor of character-reading and character-writing skills than their performance in the oddity test. The overall findings suggest that Pinyin invented spelling may be a more powerful measurement of Chinese PA for CSL learners in comparison to the conventional test, in line with the findings reported for Chinese-speaking children. The theoretical implications for understanding the role of phonological skills in the growth of Chinese literacy skills and practical suggestions for measuring Chinese PA are proposed.

High-power and high-energy Nd:YAG-Nd:YVO4 hybrid gain Raman yellow laser.

We present a simple and reliable method to successfully reconcile the average output power and pulse energy of the solid-state Raman yellow lasers. By virtue of the hybrid laser gain of Nd:YAG and Nd:YVO4 in an intracavity frequency-doubled Raman, much higher pumping is allowed and nearly linear polarized fundamental and Stokes waves can be delivered for efficient non-critical phase matching. 7.6 W of yellow output at 588 nm is obtained under incident pump power of 42.0 W at the pulse repetition frequency (PRF) of 110 kHz and the pulse energy reaches 0.41 mJ under the same incident pump power at the PRF of 10 kHz.

Vicarious traumatization in the general public, members, and non-members of medical teams aiding in COVID-19 control.

Since December 2019, more than 79,000 people have been diagnosed with infection of the Corona Virus Disease 2019 (COVID-19). A large number of medical staff was sent to Wuhan city and Hubei province to aid COVID-19 control. Psychological stress, especially vicarious traumatization caused by the COVID-19 pandemic, should not be ignored. To address this concern, the study employed a total of 214 general public and 526 nurses (i.e., 234 front-line nurses and 292 non-front-line nurses) to evaluate vicarious traumatization scores via a mobile app-based questionnaire. Front-line nurses are engaged in the process of providing care for patients with COVID-19. The results showed that the vicarious traumatization scores for front-line nurses including scores for physiological and psychological responses, were significantly lower than those of non-front-line nurses (P < 0.001). Interestingly, the vicarious traumatization scores of the general public were significantly higher than those of the front-line nurses (P < 0.001); however, no statistical difference was observed compared to the scores of non-front-line nurses (P > 0.05). Therefore, increased attention should be paid to the psychological problems of the medical staff, especially non-front-line nurses, and general public under the situation of the spread and control of COVID-19. Early strategies that aim to prevent and treat vicarious traumatization in medical staff and general public are extremely necessary.