The malate shuttle detoxifies ammonia in exhausted T cells by producing 2-ketoglutarate.

The malate shuttle is traditionally understood to maintain NAD+/NADH balance between the cytosol and mitochondria. Whether the malate shuttle has additional functions is unclear. Here we show that chronic viral infections induce CD8+ T cell expression of GOT1, a central enzyme in the malate shuttle. Got1 deficiency decreased the NAD+/NADH ratio and limited antiviral CD8+ T cell responses to chronic infection; however, increasing the NAD+/NADH ratio did not restore T cell responses. Got1 deficiency reduced the production of the ammonia scavenger 2-ketoglutarate (2-KG) from glutaminolysis and led to a toxic accumulation of ammonia in CD8+ T cells. Supplementation with 2-KG assimilated and detoxified ammonia in Got1-deficient T cells and restored antiviral responses. These data indicate that the major function of the malate shuttle in CD8+ T cells is not to maintain the NAD+/NADH balance but rather to detoxify ammonia and enable sustainable ammonia-neutral glutamine catabolism in CD8+ T cells during chronic infection.

Pathogenesis of SARS-CoV-2 in Transgenic Mice Expressing Human Angiotensin-Converting Enzyme 2.

COVID-19 has spread worldwide since 2019 and is now a severe threat to public health. We previously identified the causative agent as a novel SARS-related coronavirus (SARS-CoV-2) that uses human angiotensin-converting enzyme 2 (hACE2) as the entry receptor. Here, we successfully developed a SARS-CoV-2 hACE2 transgenic mouse (HFH4-hACE2 in C3B6 mice) infection model. The infected mice generated typical interstitial pneumonia and pathology that were similar to those of COVID-19 patients. Viral quantification revealed the lungs as the major site of infection, although viral RNA could also be found in the eye, heart, and brain in some mice. Virus identical to SARS-CoV-2 in full-genome sequences was isolated from the infected lung and brain tissues. Last, we showed that pre-exposure to SARS-CoV-2 could protect mice from severe pneumonia. Our results show that the hACE2 mouse would be a valuable tool for testing potential vaccines and therapeutics.

Graft-versus-host disease is locally maintained in target tissues by resident progenitor-like T cells.

In allogeneic hematopoietic stem cell transplantation, donor αß T cells attack recipient tissues, causing graft-versus-host disease (GVHD), a major cause of morbidity and mortality. A central question has been how GVHD is sustained despite T cell exhaustion from chronic antigen stimulation. The current model for GVHD holds that disease is maintained through the continued recruitment of alloreactive effectors from blood into affected tissues. Here, we show, using multiple approaches including parabiosis of mice with GVHD, that GVHD is instead primarily maintained locally within diseased tissues. By tracking 1,203 alloreactive T cell clones, we fitted a mathematical model predicting that within each tissue a small number of progenitor T cells maintain a larger effector pool. Consistent with this, we identified a tissue-resident TCF-1+ subpopulation that preferentially engrafted, expanded, and differentiated into effectors upon adoptive transfer. These results suggest that therapies targeting affected tissues and progenitor T cells within them would be effective.

Kinetic Analysis of Protein Stability Reveals Age-Dependent Degradation.

Do young and old protein molecules have the same probability to be degraded? We addressed this question using metabolic pulse-chase labeling and quantitative mass spectrometry to obtain degradation profiles for thousands of proteins. We find that >10% of proteins are degraded non-exponentially. Specifically, proteins are less stable in the first few hours of their life and stabilize with age. Degradation profiles are conserved and similar in two cell types. Many non-exponentially degraded (NED) proteins are subunits of complexes that are produced in super-stoichiometric amounts relative to their exponentially degraded (ED) counterparts. Within complexes, NED proteins have larger interaction interfaces and assemble earlier than ED subunits. Amplifying genes encoding NED proteins increases their initial degradation. Consistently, decay profiles can predict protein level attenuation in aneuploid cells. Together, our data show that non-exponential degradation is common, conserved, and has important consequences for complex formation and regulation of protein abundance.

TMCO1 Is an ER Ca(2+) Load-Activated Ca(2+) Channel.

Maintaining homeostasis of Ca(2+) stores in the endoplasmic reticulum (ER) is crucial for proper Ca(2+) signaling and key cellular functions. The Ca(2+)-release-activated Ca(2+) (CRAC) channel is responsible for Ca(2+) influx and refilling after store depletion, but how cells cope with excess Ca(2+) when ER stores are overloaded is unclear. We show that TMCO1 is an ER transmembrane protein that actively prevents Ca(2+) stores from overfilling, acting as what we term a "Ca(2+) load-activated Ca(2+) channel" or "CLAC" channel. TMCO1 undergoes reversible homotetramerization in response to ER Ca(2+) overloading and disassembly upon Ca(2+) depletion and forms a Ca(2+)-selective ion channel on giant liposomes. TMCO1 knockout mice reproduce the main clinical features of human cerebrofaciothoracic (CFT) dysplasia spectrum, a developmental disorder linked to TMCO1 dysfunction, and exhibit severe mishandling of ER Ca(2+) in cells. Our findings indicate that TMCO1 provides a protective mechanism to prevent overfilling of ER stores with Ca(2+) ions.

Lithium tantalate photonic integrated circuits for volume manufacturing.

Electro-optical photonic integrated circuits (PICs) based on lithium niobate (LiNbO3) have demonstrated the vast capabilities of materials with a high Pockels coefficient1,2. They enable linear and high-speed modulators operating at complementary metal-oxide-semiconductor voltage levels3 to be used in applications including data-centre communications4, high-performance computing and photonic accelerators for AI5. However, industrial use of this technology is hindered by the high cost per wafer and the limited wafer size. The high cost results from the lack of existing high-volume applications in other domains of the sort that accelerated the adoption of silicon-on-insulator (SOI) photonics, which was driven by vast investment in microelectronics. Here we report low-loss PICs made of lithium tantalate (LiTaO3), a material that has already been adopted commercially for 5G radiofrequency filters6 and therefore enables scalable manufacturing at low cost, and it has equal, and in some cases superior, properties to LiNbO3. We show that LiTaO3 can be etched to create low-loss (5.6 dB m-1) PICs using a deep ultraviolet (DUV) stepper-based manufacturing process7. We demonstrate a LiTaO3 Mach-Zehnder modulator (MZM) with a half-wave voltage-length product of 1.9 V cm and an electro-optic bandwidth of up to 40 GHz. In comparison with LiNbO3, LiTaO3 exhibits a much lower birefringence, enabling high-density circuits and broadband operation over all telecommunication bands. Moreover, the platform supports the generation of soliton microcombs. Our work paves the way for the scalable manufacture of low-cost and large-volume next-generation electro-optical PICs.

Triple-junction solar cells with cyanate in ultrawide-bandgap perovskites.

Perovskite bandgap tuning without quality loss makes perovskites unique among solar absorbers, offering promising avenues for tandem solar cells1,2. However, minimizing the voltage loss when their bandgap is increased to above 1.90 eV for triple-junction tandem use is challenging3-5. Here we present a previously unknown pseudohalide, cyanate (OCN-), with a comparable effective ionic radius (1.97 Å) to bromide (1.95 Å) as a bromide substitute. Electron microscopy and X-ray scattering confirm OCN incorporation into the perovskite lattice. This contributes to notable lattice distortion, ranging from 90.5° to 96.6°, a uniform iodide-bromide distribution and consistent microstrain. Owing to these effects, OCN-based perovskite exhibits enhanced defect formation energy and substantially decreased non-radiative recombination. We achieved an inverted perovskite (1.93 eV) single-junction device with an open-circuit voltage (VOC) of 1.422 V, a VOC × FF (fill factor) product exceeding 80% of the Shockley-Queisser limit and stable performance under maximum power point tracking, culminating in a 27.62% efficiency (27.10% certified efficiency) perovskite-perovskite-silicon triple-junction solar cell with 1 cm2 aperture area.

Osmosensor-mediated control of Ca2+ spiking in pollen germination.

Higher plants survive terrestrial water deficiency and fluctuation by arresting cellular activities (dehydration) and resuscitating processes (rehydration). However, how plants monitor water availability during rehydration is unknown. Although increases in hypo-osmolarity-induced cytosolic Ca2+ concentration (HOSCA) have long been postulated to be the mechanism for sensing hypo-osmolarity in rehydration1,2, the molecular basis remains unknown. Because osmolarity triggers membrane tension and the osmosensing specificity of osmosensing channels can only be determined in vivo3-5, these channels have been classified as a subtype of mechanosensors. Here we identify bona fide cell surface hypo-osmosensors in Arabidopsis and find that pollen Ca2+ spiking is controlled directly by water through these hypo-osmosensors-that is, Ca2+ spiking is the second messenger for water status. We developed a functional expression screen in Escherichia coli for hypo-osmosensitive channels and identified OSCA2.1, a member of the hyperosmolarity-gated calcium-permeable channel (OSCA) family of proteins6. We screened single and high-order OSCA mutants, and observed that the osca2.1/osca2.2 double-knockout mutant was impaired in pollen germination and HOSCA. OSCA2.1 and OSCA2.2 function as hypo-osmosensitive Ca2+-permeable channels in planta and in HEK293 cells. Decreasing osmolarity of the medium enhanced pollen Ca2+ oscillations, which were mediated by OSCA2.1 and OSCA2.2 and required for germination. OSCA2.1 and OSCA2.2 convert extracellular water status into Ca2+ spiking in pollen and may serve as essential hypo-osmosensors for tracking rehydration in plants.

Human Mast Cell Proteome Reveals Unique Lineage, Putative Functions, and Structural Basis for Cell Ablation.

Mast cells are rare tissue-resident cells of importance to human allergies. To understand the structural basis of principle mast cell functions, we analyzed the proteome of primary human and mouse mast cells by quantitative mass spectrometry. We identified a mast-cell-specific proteome signature, indicative of a unique lineage, only distantly related to other immune cell types, including innate immune cells. Proteome comparison between human and mouse suggested evolutionary conservation of core mast cell functions. In addition to specific proteases and proteins associated with degranulation and proteoglycan biosynthesis, mast cells expressed proteins potentially involved in interactions with neurons and neurotransmitter metabolism, including cell adhesion molecules, ion channels, and G protein coupled receptors. Toward targeted cell ablation in severe allergic diseases, we used MRGPRX2 for mast cell depletion in human skin biopsies. These proteome analyses suggest a unique role of mast cells in the immune system, probably intertwined with the nervous system.

Single-Cell RNA Sequencing of Tumor-Infiltrating NK Cells Reveals that Inhibition of Transcription Factor HIF-1α Unleashes NK Cell Activity.

Enhancing immune cell functions in tumors remains a major challenge in cancer immunotherapy. Hypoxia is a common feature of solid tumors, and cells adapt by upregulating the transcription factor HIF-1α. Here, we defined the transcriptional landscape of mouse tumor-infiltrating natural killer (NK) cells by using single-cell RNA sequencing. Conditional deletion of Hif1a in NK cells resulted in reduced tumor growth, elevated expression of activation markers, effector molecules, and an enriched NF-κB pathway in tumor-infiltrating NK cells. Interleukin-18 (IL-18) from myeloid cells was required for NF-κB activation and the enhanced anti-tumor activity of Hif1a-/- NK cells. Extended culture with an HIF-1α inhibitor increased human NK cell responses. Low HIF1A expression was associated with high expression of IFNG in human tumor-infiltrating NK cells, and an enriched NK-IL18-IFNG signature in solid tumors correlated with increased overall patient survival. Thus, inhibition of HIF-1α unleashes NK cell anti-tumor activity and could be exploited for cancer therapy.

FOXP3 recognizes microsatellites and bridges DNA through multimerization.

FOXP3 is a transcription factor that is essential for the development of regulatory T cells, a branch of T cells that suppress excessive inflammation and autoimmunity1-5. However, the molecular mechanisms of FOXP3 remain unclear. Here we here show that FOXP3 uses the forkhead domain-a DNA-binding domain that is commonly thought to function as a monomer or dimer-to form a higher-order multimer after binding to TnG repeat microsatellites. The cryo-electron microscopy structure of FOXP3 in a complex with T3G repeats reveals a ladder-like architecture, whereby two double-stranded DNA molecules form the two 'side rails' bridged by five pairs of FOXP3 molecules, with each pair forming a 'rung'. Each FOXP3 subunit occupies TGTTTGT within the repeats in a manner that is indistinguishable from that of FOXP3 bound to the forkhead consensus motif (TGTTTAC). Mutations in the intra-rung interface impair TnG repeat recognition, DNA bridging and the cellular functions of FOXP3, all without affecting binding to the forkhead consensus motif. FOXP3 can tolerate variable inter-rung spacings, explaining its broad specificity for TnG-repeat-like sequences in vivo and in vitro. Both FOXP3 orthologues and paralogues show similar TnG repeat recognition and DNA bridging. These findings therefore reveal a mode of DNA recognition that involves transcription factor homomultimerization and DNA bridging, and further implicates microsatellites in transcriptional regulation and diseases.

Light-induced ferromagnetism in moiré superlattices.

Many-body interactions between carriers lie at the heart of correlated physics. The ability to tune such interactions would allow the possibility to access and control complex electronic phase diagrams. Recently, two-dimensional moiré superlattices have emerged as a promising platform for quantum engineering such phenomena1-3. The power of the moiré system lies in the high tunability of its physical parameters by adjusting the layer twist angle1-3, electrical field4-6, moiré carrier filling7-11 and interlayer coupling12. Here we report that optical excitation can highly tune the spin-spin interactions between moiré-trapped carriers, resulting in ferromagnetic order in WS2 /WSe2 moiré superlattices. Near the filling factor of -1/3 (that is, one hole per three moiré unit cells), as the excitation power at the exciton resonance increases, a well-developed hysteresis loop emerges in the reflective magnetic circular dichroism signal as a function of magnetic field, a hallmark of ferromagnetism. The hysteresis loop persists down to charge neutrality, and its shape evolves as the moiré superlattice is gradually filled, indicating changes of magnetic ground state properties. The observed phenomenon points to a mechanism in which itinerant photoexcited excitons mediate exchange coupling between moiré-trapped holes. This exciton-mediated interaction can be of longer range than direct coupling between moiré-trapped holes9, and thus magnetic order arises even in the dilute hole regime. This discovery adds a dynamic tuning knob to the rich many-body Hamiltonian of moiré quantum matter13-19.

Strigolactones promote plant freezing tolerance by releasing the WRKY41-mediated inhibition of CBF/DREB1 expression.

Cold stress is a major abiotic stress that adversely affects plant growth and crop productivity. The C-REPEAT BINDING FACTOR/DRE BINDING FACTOR 1 (CBF/DREB1) transcriptional regulatory cascade plays a key role in regulating cold acclimation and freezing tolerance in Arabidopsis (Arabidopsis thaliana). Here, we show that max (more axillary growth) mutants deficient in strigolactone biosynthesis and signaling display hypersensitivity to freezing stress. Exogenous application of GR245DS , a strigolactone analog, enhances freezing tolerance in wild-type plants and strigolactone-deficient mutants and promotes the cold-induced expression of CBF genes. Biochemical analysis showed that the transcription factor WRKY41 serves as a substrate for the F-box E3 ligase MAX2. WRKY41 directly binds to the W-box in the promoters of CBF genes and represses their expression, negatively regulating cold acclimation and freezing tolerance. MAX2 ubiquitinates WRKY41, thus marking it for cold-induced degradation and thereby alleviating the repression of CBF expression. In addition, SL-mediated degradation of SMXLs also contributes to enhanced plant freezing tolerance by promoting anthocyanin biosynthesis. Taken together, our study reveals the molecular mechanism underlying strigolactones promote the cold stress response in Arabidopsis.

N1 versus N2 and PMN-MDSC: A critical appraisal of current concepts on tumor-associated neutrophils and new directions for human oncology.

Research on tumor-associated neutrophils (TAN) currently surges because of the well-documented strong clinical relevance of tumor-infiltrating neutrophils. This relevance is illustrated by strong correlations between high frequencies of intratumoral neutrophils and poor outcome in the majority of human cancers. Recent high-dimensional analysis of murine neutrophils provides evidence for unexpected plasticity of neutrophils in murine models of cancer and other inflammatory non-malignant diseases. New analysis tools enable deeper insight into the process of neutrophil differentiation and maturation. These technological and scientific developments led to the description of an ever-increasing number of distinct transcriptional states and associated phenotypes in murine models of disease and more recently also in humans. At present, functional validation of these different transcriptional states and potential phenotypes in cancer is lacking. Current functional concepts on neutrophils in cancer rely mainly on the myeloid-derived suppressor cell (MDSC) concept and the dichotomous and simple N1-N2 paradigm. In this manuscript, we review the historic development of those concepts, critically evaluate these concepts against the background of our own work and provide suggestions for a refinement of current concepts in order to facilitate the transition of TAN research from experimental insight to clinical translation.

Chronic active Epstein-Barr virus disease originates from infected hematopoietic stem cells.

ABSTRACT: Chronic active Epstein-Barr virus (EBV) disease (CAEBV) is a lethal syndrome because of persistent EBV infection. When diagnosed as CAEBV, EBV infection was observed in multiple hematopoietic lineages, but the etiology of CAEBV is still elusive. Bone marrow and peripheral cells derived from 5 patients with CAEBV, 1 patient with EBV-associated hemophagocytic lymphohistiocytosis, and 2 healthy controls were analyzed. Multiple assays were applied to identify and characterize EBV-infected cells, including quantitative polymerase chain reaction, PrimeFlow, and single-cell RNA-sequencing (scRNA-seq). Based on scRNA-seq data, alterations in gene expression of particular cell types were analyzed between patients with CAEBV and controls, and between infected and uninfected cells. One patient with CAEBV was treated with allogeneic hematopoietic stem cell transplantation (HSCT), and the samples derived from this patient were analyzed again 6 months after HSCT. EBV infected the full spectrum of the hematopoietic system including both lymphoid and myeloid lineages, as well as the hematopoietic stem cells (HSCs) of the patients with CAEBV. EBV-infected HSCs exhibited a higher differentiation rate toward downstream lineages, and the EBV infection had an impact on both the innate and adaptive immunity, resulting in inflammatory symptoms. EBV-infected cells were thoroughly removed from the hematopoietic system after HSCT. Taken together, multiple lines of evidence presented in this study suggest that CAEBV disease originates from the infected HSCs, which might potentially lead to innovative therapy strategies for CAEBV.

PUB25 and PUB26 dynamically modulate ICE1 stability via differential ubiquitination during cold stress in Arabidopsis.

Ubiquitination modulates protein turnover or activity depending on the number and location of attached ubiquitin (Ub) moieties. Proteins marked by a lysine 48 (K48)-linked polyubiquitin chain are usually targeted to the 26S proteasome for degradation; however, other polyubiquitin chains, such as those attached to K63, usually regulate other protein properties. Here, we show that 2 PLANT U-BOX E3 ligases, PUB25 and PUB26, facilitate both K48- and K63-linked ubiquitination of the transcriptional regulator INDUCER OF C-REPEAT BINDING FACTOR (CBF) EXPRESSION1 (ICE1) during different periods of cold stress in Arabidopsis (Arabidopsis thaliana), thus dynamically modulating ICE1 stability. Moreover, PUB25 and PUB26 attach both K48- and K63-linked Ub chains to MYB15 in response to cold stress. However, the ubiquitination patterns of ICE1 and MYB15 mediated by PUB25 and PUB26 differ, thus modulating their protein stability and abundance during different stages of cold stress. Furthermore, ICE1 interacts with and inhibits the DNA-binding activity of MYB15, resulting in an upregulation of CBF expression. This study unravels a mechanism by which PUB25 and PUB26 add different polyubiquitin chains to ICE1 and MYB15 to modulate their stability, thereby regulating the timing and degree of cold stress responses in plants.

Unconventional IFNω-like Genes Dominate the Type I IFN Locus and the Constitutive Antiviral Responses in Bats.

Bats are the natural reservoir hosts of some viruses, some of which may spill over to humans and cause global-scale pandemics. Different from humans, bats may coexist with high pathogenic viruses without showing symptoms of diseases. As one of the most important first defenses, bat type I IFNs (IFN-Is) were thought to play a role during this virus coexistence and thus were studied in recent years. However, there are arguments about whether bats have a contracted genome locus or constitutively expressed IFNs, mainly due to species-specific findings. We hypothesized that because of the lack of pan-bat analysis, the common characteristics of bat IFN-Is have not been revealed yet. In this study, we characterized the IFN-I locus for nine Yangochiroptera bats and three Yinpterochiroptera bats on the basis of their high-quality bat genomes. We also compared the basal expression in six bats and compared the antiviral and antiproliferative activity and the thermostability of representative Rhinolophus bat IFNs. We found a dominance of unconventional IFNω-like responses in the IFN-I system, which is unique to bats. In contrast to IFNα-dominated IFN-I loci in the majority of other mammals, bats generally have shorter IFN-I loci with more unconventional IFNω-like genes (IFNω or related IFNαω), but with fewer or even no IFNα genes. In addition, bats generally have constitutively expressed IFNs, the highest expressed of which is more likely an IFNω-like gene. Likewise, the highly expressed IFNω-like protein also demonstrated the best antiviral activity, antiproliferative activity, or thermostability, as shown in a representative Rhinolophus bat species. Overall, we revealed pan-bat unique, to our knowledge, characteristics in the IFN-I system, which provide insights into our understanding of the innate immunity that contributes to a special coexistence between bats and viruses.

H2A.Z facilitates licensing and activation of early replication origins.

DNA replication is a tightly regulated process that ensures the precise duplication of the genome during the cell cycle1. In eukaryotes, the licensing and activation of replication origins are regulated by both DNA sequence and chromatin features2. However, the chromatin-based regulatory mechanisms remain largely uncharacterized. Here we show that, in HeLa cells, nucleosomes containing the histone variant H2A.Z are enriched with histone H4 that is dimethylated on its lysine 20 residue (H4K20me2) and with bound origin-recognition complex (ORC). In vitro studies show that H2A.Z-containing nucleosomes bind directly to the histone lysine methyltransferase enzyme SUV420H1, promoting H4K20me2 deposition, which is in turn required for ORC1 binding. Genome-wide studies show that signals from H4K20me2, ORC1 and nascent DNA strands co-localize with H2A.Z, and that depletion of H2A.Z results in decreased H4K20me2, ORC1 and nascent-strand signals throughout the genome. H2A.Z-regulated replication origins have a higher firing efficiency and early replication timing compared with other origins. Our results suggest that the histone variant H2A.Z epigenetically regulates the licensing and activation of early replication origins and maintains replication timing through the SUV420H1-H4K20me2-ORC1 axis.

Author Correction: H2A.Z facilitates licensing and activation of early replication origins.

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

A pneumonia outbreak associated with a new coronavirus of probable bat origin.

Since the outbreak of severe acute respiratory syndrome (SARS) 18 years ago, a large number of SARS-related coronaviruses (SARSr-CoVs) have been discovered in their natural reservoir host, bats1-4. Previous studies have shown that some bat SARSr-CoVs have the potential to infect humans5-7. Here we report the identification and characterization of a new coronavirus (2019-nCoV), which caused an epidemic of acute respiratory syndrome in humans in Wuhan, China. The epidemic, which started on 12 December 2019, had caused 2,794 laboratory-confirmed infections including 80 deaths by 26 January 2020. Full-length genome sequences were obtained from five patients at an early stage of the outbreak. The sequences are almost identical and share 79.6% sequence identity to SARS-CoV. Furthermore, we show that 2019-nCoV is 96% identical at the whole-genome level to a bat coronavirus. Pairwise protein sequence analysis of seven conserved non-structural proteins domains show that this virus belongs to the species of SARSr-CoV. In addition, 2019-nCoV virus isolated from the bronchoalveolar lavage fluid of a critically ill patient could be neutralized by sera from several patients. Notably, we confirmed that 2019-nCoV uses the same cell entry receptor-angiotensin converting enzyme II (ACE2)-as SARS-CoV.