TIR domains of plant immune receptors are 2',3'-cAMP/cGMP synthetases mediating cell death.

2',3'-cAMP is a positional isomer of the well-established second messenger 3',5'-cAMP, but little is known about the biology of this noncanonical cyclic nucleotide monophosphate (cNMP). Toll/interleukin-1 receptor (TIR) domains of nucleotide-binding leucine-rich repeat (NLR) immune receptors have the NADase function necessary but insufficient to activate plant immune responses. Here, we show that plant TIR proteins, besides being NADases, act as 2',3'-cAMP/cGMP synthetases by hydrolyzing RNA/DNA. Structural data show that a TIR domain adopts distinct oligomers with mutually exclusive NADase and synthetase activity. Mutations specifically disrupting the synthetase activity abrogate TIR-mediated cell death in Nicotiana benthamiana (Nb), supporting an important role for these cNMPs in TIR signaling. Furthermore, the Arabidopsis negative regulator of TIR-NLR signaling, NUDT7, displays 2',3'-cAMP/cGMP but not 3',5'-cAMP/cGMP phosphodiesterase activity and suppresses cell death activity of TIRs in Nb. Our study identifies a family of 2',3'-cAMP/cGMP synthetases and establishes a critical role for them in plant immune responses.

An Ancient Harappan Genome Lacks Ancestry from Steppe Pastoralists or Iranian Farmers.

We report an ancient genome from the Indus Valley Civilization (IVC). The individual we sequenced fits as a mixture of people related to ancient Iranians (the largest component) and Southeast Asian hunter-gatherers, a unique profile that matches ancient DNA from 11 genetic outliers from sites in Iran and Turkmenistan in cultural communication with the IVC. These individuals had little if any Steppe pastoralist-derived ancestry, showing that it was not ubiquitous in northwest South Asia during the IVC as it is today. The Iranian-related ancestry in the IVC derives from a lineage leading to early Iranian farmers, herders, and hunter-gatherers before their ancestors separated, contradicting the hypothesis that the shared ancestry between early Iranians and South Asians reflects a large-scale spread of western Iranian farmers east. Instead, sampled ancient genomes from the Iranian plateau and IVC descend from different groups of hunter-gatherers who began farming without being connected by substantial movement of people.

Publisher Correction: The discovery of human TSLP as a critical epithelial cytokine in type 2 immunity and allergic disease.

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

Personalized Immunomonitoring Uncovers Molecular Networks that Stratify Lupus Patients.

Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by loss of tolerance to nucleic acids and highly diverse clinical manifestations. To assess its molecular heterogeneity, we longitudinally profiled the blood transcriptome of 158 pediatric patients. Using mixed models accounting for repeated measurements, demographics, treatment, disease activity (DA), and nephritis class, we confirmed a prevalent IFN signature and identified a plasmablast signature as the most robust biomarker of DA. We detected gradual enrichment of neutrophil transcripts during progression to active nephritis and distinct signatures in response to treatment in different nephritis subclasses. Importantly, personalized immunomonitoring uncovered individual correlates of disease activity that enabled patient stratification into seven groups, supported by patient genotypes. Our study uncovers the molecular heterogeneity of SLE and provides an explanation for the failure of clinical trials. This approach may improve trial design and implementation of tailored therapies in genetically and clinically complex autoimmune diseases. PAPERCLIP.

Ordered assembly of the cytosolic RNA-sensing MDA5-MAVS signaling complex via binding to unanchored K63-linked poly-ubiquitin chains.

The RNA sensor MDA5 recruits the signaling adaptor MAVS to initiate type I interferon signaling and downstream antiviral responses, a process that requires K63-linked polyubiquitin chains. Here, we examined the mechanisms whereby K63-polyUb chain regulate MDA5 activation. Only long unanchored K63-polyUbn (n ≥ 8) could mediate tetramerization of the caspase activation and recruitment domains of MDA5 (MDA5CARDs). Cryoelectron microscopy structures of a polyUb13-bound MDA5CARDs tetramer and a polyUb11-bound MDA5CARDs-MAVSCARD assembly revealed a tower-like formation, wherein eight Ubs tethered along the outer rim of the helical shell, bridging MDA5CARDs and MAVSCARD tetramers into proximity. ATP binding and hydrolysis promoted the stabilization of RNA-bound MDA5 prior to MAVS activation via allosteric effects on CARDs-polyUb complex. Abundant ATP prevented basal activation of apo MDA5. Our findings reveal the ordered assembly of a MDA5 signaling complex competent to recruit and activate MAVS and highlight differences with RIG-I in terms of CARD orientation and Ub sensing that suggest different abilities to induce antiviral responses.

The Transcriptome and DNA Methylome Landscapes of Human Primordial Germ Cells.

Germ cells are vital for transmitting genetic information from one generation to the next and for maintaining the continuation of species. Here, we analyze the transcriptome of human primordial germ cells (PGCs) from the migrating stage to the gonadal stage at single-cell and single-base resolutions. Human PGCs show unique transcription patterns involving the simultaneous expression of both pluripotency genes and germline-specific genes, with a subset of them displaying developmental-stage-specific features. Furthermore, we analyze the DNA methylome of human PGCs and find global demethylation of their genomes. Approximately 10 to 11 weeks after gestation, the PGCs are nearly devoid of any DNA methylation, with only 7.8% and 6.0% of the median methylation levels in male and female PGCs, respectively. Our work paves the way toward deciphering the complex epigenetic reprogramming of the germline with the aim of restoring totipotency in fertilized oocytes.

Oligomerization-mediated autoinhibition and cofactor binding of a plant NLR.

Nucleotide-binding leucine-rich repeat (NLR) proteins have a pivotal role in plant immunity by recognizing pathogen effectors1,2. Maintaining a balanced immune response is crucial, as excessive NLR expression can lead to unintended autoimmunity3,4. Unlike most NLRs, plant NLR required for cell death 2 (NRC2) belongs to a small NLR group characterized by constitutively high expression without self-activation5. The mechanisms underlying NRC2 autoinhibition and activation are not yet understood. Here we show that Solanum lycopersicum (tomato) NRC2 (SlNRC2) forms dimers and tetramers, and higher-order oligomers at elevated concentrations. Cryo-electron microscopy (cryo-EM) reveals an inactive conformation of SlNRC2 within these oligomers. Dimerization and oligomerization not only stabilize the inactive state but also sequester SlNRC2 from assembling into an active form. Mutations at the dimeric or inter-dimeric interfaces enhance pathogen-induced cell death and immunity in Nicotiana (N.) benthamiana. The cryo-EM structures unexpectedly reveal inositol hexakisphosphate (IP6) or pentakisphosphate (IP5) bound to the inner surface of SlNRC2's C-terminal LRR domain as confirmed by mass spectrometry. Mutations at the IP-binding site impair inositol phosphate binding of SlNRC2 and pathogen-induced SlNRC2-mediated cell death in N. benthamiana. Together, our study unveils a novel negative regulatory mechanism of NLR activation and suggests inositol phosphates as cofactors of NRCs.

Identification of a role for TRIM29 in the control of innate immunity in the respiratory tract.

The respiratory tract is heavily populated with innate immune cells, but the mechanisms that control such cells are poorly defined. Here we found that the E3 ubiquitin ligase TRIM29 was a selective regulator of the activation of alveolar macrophages, the expression of type I interferons and the production of proinflammatory cytokines in the lungs. We found that deletion of TRIM29 enhanced macrophage production of type I interferons and protected mice from infection with influenza virus, while challenge of Trim29-/- mice with Haemophilus influenzae resulted in lethal lung inflammation due to massive production of proinflammatory cytokines by macrophages. Mechanistically, we demonstrated that TRIM29 inhibited interferon-regulatory factors and signaling via the transcription factor NF-κB by degrading the adaptor NEMO and that TRIM29 directly bound NEMO and subsequently induced its ubiquitination and proteolytic degradation. These data identify TRIM29 as a key negative regulator of alveolar macrophages and might have important clinical implications for local immunity and immunopathology.

IL-1 is a critical regulator of group 2 innate lymphoid cell function and plasticity.

Group 2 innate lymphoid cells (ILC2 cells) are important for type 2 immune responses and are activated by the epithelial cytokines interleukin 33 (IL-33), IL-25 and thymic stromal lymphopoietin (TSLP). Here we demonstrated that IL-1ß was a critical activator of ILC2 cells, inducing proliferation and cytokine production and regulating the expression of epithelial cytokine receptors. IL-1ß also governed ILC2 plasticity by inducing low expression of the transcription factor T-bet and the cytokine receptor chain IL-12Rß2, which enabled the conversion of these cells into an ILC1 phenotype in response to IL-12. This transition was marked by an atypical chromatin landscape characterized by the simultaneous transcriptional accessibility of the locus encoding interferon-γ (IFN-γ) and the loci encoding IL-5 and IL-13. Finally, IL-1ß potentiated ILC2 activation and plasticity in vivo, and IL-12 acted as the switch that determined an ILC2-versus-ILC1 response. Thus, we have identified a previously unknown role for IL-1ß in facilitating ILC2 maturation and plasticity.

Splicing factor YBX1 regulates bone marrow stromal cell fate during aging.

Senescence and altered differentiation potential of bone marrow stromal cells (BMSCs) lead to age-related bone loss. As an important posttranscriptional regulatory pathway, alternative splicing (AS) regulates the diversity of gene expression and has been linked to induction of cellular senescence. However, the role of splicing factors in BMSCs during aging remains poorly defined. Herein, we found that the expression of the splicing factor Y-box binding protein 1 (YBX1) in BMSCs decreased with aging in mice and humans. YBX1 deficiency resulted in mis-splicing in genes linked to BMSC osteogenic differentiation and senescence, such as Fn1, Nrp2, Sirt2, Sp7, and Spp1, thus contributing to BMSC senescence and differentiation shift during aging. Deletion of Ybx1 in BMSCs accelerated bone loss in mice, while its overexpression stimulated bone formation. Finally, we identified a small compound, sciadopitysin, which attenuated the degradation of YBX1 and bone loss in old mice. Our study demonstrated that YBX1 governs cell fate of BMSCs via fine control of RNA splicing and provides a potential therapeutic target for age-related osteoporosis.

T(H)17 cells promote microbial killing and innate immune sensing of DNA via interleukin 26.

Interleukin 17-producing helper T cells (T(H)17 cells) have a major role in protection against infections and in mediating autoimmune diseases, yet the mechanisms involved are incompletely understood. We found that interleukin 26 (IL-26), a human T(H)17 cell-derived cytokine, is a cationic amphipathic protein that kills extracellular bacteria via membrane-pore formation. Furthermore, T(H)17 cell-derived IL-26 formed complexes with bacterial DNA and self-DNA released by dying bacteria and host cells. The resulting IL-26-DNA complexes triggered the production of type I interferon by plasmacytoid dendritic cells via activation of Toll-like receptor 9, but independently of the IL-26 receptor. These findings provide insights into the potent antimicrobial and proinflammatory function of T(H)17 cells by showing that IL-26 is a natural human antimicrobial that promotes immune sensing of bacterial and host cell death.

Trained immunity in recurrent Staphylococcus aureus infection promotes bacterial persistence.

Bacterial persister cells, a sub-population of dormant phenotypic variants highly tolerant to antibiotics, present a significant challenge for infection control. Investigating the mechanisms of antibiotic persistence is crucial for developing effective treatment strategies. Here, we found a significant association between tolerance frequency and previous infection history in bovine mastitis. Previous S. aureus infection led to S. aureus tolerance to killing by rifampicin in subsequent infection in vivo and in vitro. Actually, the activation of trained immunity contributed to rifampicin persistence of S. aureus in secondary infection, where it reduced the effectiveness of antibiotic treatment and increased disease severity. Mechanically, we found that S. aureus persistence was mediated by the accumulation of fumarate provoked by trained immunity. Combination therapy with metformin and rifampicin promoted eradication of persisters and improved the severity of recurrent S. aureus infection. These findings provide mechanistic insight into the relationship between trained immunity and S. aureus persistence, while providing proof of concept that trained immunity is a therapeutic target in recurrent bacterial infections involving persistent pathogens.

USP36 inhibits apoptosis by deubiquitinating cIAP1 and survivin in colorectal cancer cells.

Chemotherapeutic agents for treating colorectal cancer (CRC) primarily induce apoptosis in tumor cells. The ubiquitin-proteasome system is critical for apoptosis regulation. Deubiquitinating enzymes (DUBs) remove ubiquitin from substrates to reverse ubiquitination. Although over 100 DUB members have been discovered, the biological functions of only a small proportion of DUBs have been characterized. Here, we aimed to systematically identify the DUBs that contribute to the development of CRC. Among the DUBs, ubiquitin-specific protease 36 (USP36) is upregulated in CRC. We showed that the knockdown of USP36 induces intrinsic and extrinsic apoptosis. Through gene silencing and coimmunoprecipitation techniques, we identified survivin and cIAP1 as USP36 targets. Mechanistically, USP36 binds and removes lysine-11-linked ubiquitin chains from cIAP1 and lysine-48-linked ubiquitin chains from survivin to abolish protein degradation. Overexpression of USP36 disrupts the formation of the XIAP-second mitochondria-derived activator of caspase complex and promotes receptor-interacting protein kinase 1 ubiquitination, validating USP36 as an inhibitor to intrinsic and extrinsic apoptosis through deubiquitinating survivin and cIAP1. Therefore, our results suggest that USP36 is involved in CRC progression and is a potential therapeutic target.

The E3 ubiquitin ligase TRIM21 negatively regulates the innate immune response to intracellular double-stranded DNA.

DDX41 is a sensor of intracellular double-stranded DNA (dsDNA) in myeloid dendritic cells (mDCs) that triggers a type I interferon response via the signaling adaptor STING. We identified the E3 ligase TRIM21 as a DDX41-interacting protein and found that knockdown of or deficiency in TRIM21 resulted in enhanced type I interferon responses to intracellular dsDNA and DNA viruses. Overexpression of TRIM21 resulted in more degradation of DDX41 and less production of interferon-ß (IFN-ß) in response to intracellular dsDNA. The SPRY-PRY domain of TRIM21 interacted with the DEADc domain of DDX41. Lys9 and Lys115 of DDX41 were the targets of TRIM21-mediated ubiquitination. TRIM21 is therefore an interferon-inducible E3 ligase that induces the Lys48 (K48)-linked ubiquitination and degradation of DDX41 and negatively regulates the innate immune response to intracellular dsDNA.

Rethinking coagulation: from enzymatic cascade and cell-based reactions to a convergent model involving innate immune activation.

ABSTRACT: Advancements in the conceptual thinking of hemostasis and thrombosis have been catalyzed by major developments within health research over several decades. The cascade model of coagulation was first described in the 1960s, when biochemistry gained prominence through innovative experimentation and technical developments. This was followed by the cell-based model, which integrated cellular coordination to the enzymology of clot formation and was conceptualized during the growth period in cell biology at the turn of the millennium. Each step forward has heralded a revolution in clinical therapeutics, both in procoagulant and anticoagulant treatments to improve patient care. In current times, the COVID-19 pandemic may also prove to be a catalyst: thrombotic challenges including the mixed responses to anticoagulant treatment and the vaccine-induced immune thrombotic thrombocytopenia have exposed limitations in our preexisting concepts while simultaneously demanding novel therapeutic approaches. It is increasingly clear that innate immune activation as part of the host response to injury is not separate but integrated into adaptive clot formation. Our review summarizes current understanding of the major molecules facilitating such a cross talk between immunity, inflammation and coagulation. We demonstrate how such effects can be layered upon the cascade and cell-based models to evolve conceptual understanding of the physiology of immunohemostasis and the pathology of immunothrombosis.

Dihydroartemisinin is an inhibitor of trained immunity through Akt/mTOR/HIF1α signaling pathway.

Trained immunity is mechanistically defined as the metabolically and epigenetically mediated long-term functional adaptation of the innate immune system, characterized by a heightened response to a secondary stimulation. Given appropriate activation, trained immunity represents an attractive anti-infective therapeutic target. Nevertheless, excessive immune response and subsequent inflammatory cascades may contribute to pathological tissue damage, indicating that the negative impacts of trained immunity appear to be significant. In this study, we show that innate immune responses such as the production of extracellular traps, pro-inflammatory cytokines, and autophagy-related proteins were markedly augmented in trained BMDMs. Furthermore, heat-killed C. albicans priming promotes the activation of the AIM2 inflammasome, and AIM2-/- mice exhibit impaired memory response induced by heat-killed C. albicans. Therefore, we establish that the AIM2 inflammasome is involved in trained immunity and emerges as a promising therapeutic target for potentially deleterious effects. Dihydroartemisinin can inhibit the memory response induced by heat-killed C. albicans through modulation of mTOR signaling and the AIM2 inflammasome. The findings suggest that dihydroartemisinin can reduce the induction of trained immunity by heat-killed C. albicans in C57BL/6 mice. Dihydroartemisinin is one such therapeutic intervention that has the potential to treat of diseases characterized by excessive trained immunity.

Genomic and epigenetic landscapes drive CRISPR-based genome editing in Bifidobacterium.

Bifidobacterium is a commensal bacterial genus ubiquitous in the human gastrointestinal tract, which is associated with a range of health benefits. The advent of CRISPR-based genome editing technologies provides opportunities to investigate the genetics of important bacteria and transcend the lack of genetic tools in bifidobacteria to study the basis for their health-promoting attributes. Here, we repurpose the endogenous type I-G CRISPR-Cas system and adopt an exogenous CRISPR base editor for genome engineering in B. animalis subsp. lactis, demonstrating that both genomic and epigenetic contexts drive editing outcomes across strains. We reprogrammed the endogenous type I-G system to screen for naturally occurring large deletions up to 27 kb and to generate a 500-bp deletion in tetW to abolish tetracycline resistance. A CRISPR-cytosine base editor was optimized to install C•G-to-T•A amber mutations to resensitize multiple B. lactis strains to tetracycline. Remarkably, we uncovered epigenetic patterns that are distributed unevenly among B. lactis strains, despite their genomic homogeneity, that may contribute to editing efficiency variability. Insights were also expanded to Bifidobacterium longum subsp. infantis to emphasize the broad relevance of these findings. This study highlights the need to develop individualized CRISPR-based genome engineering approaches for distinct bacterial strains and opens avenues for engineering of next generation probiotics.

Regulating Lewis Acidic Sites of 1T-2H MoS2 Catalysts for Solar-Driven Photothermal Catalytic H2 Production from Lignocellulosic Biomass.

Solar-driven photothermal catalytic H2 production from lignocellulosic biomass was achieved by using 1T-2H MoS2 with tunable Lewis acidic sites as catalysts in an alkaline aqueous solution, in which the number of Lewis acidic sites derived from the exposed Mo edges of MoS2 was successfully regulated by both the formation of an edge-terminated 1T-2H phase structure and tunable layer number. Owing to the abundant Lewis acidic sites for the oxygenolysis of lignocellulosic biomass, the 1T-2H MoS2 catalyst shows high photothermal catalytic lignocellulosic biomass-to-H2 transformation performance in polar wood chips, bamboo, rice straw corncobs, and rice hull aqueous solutions, and the highest H2 generation rate and solar-to-H2 (STH) efficiency respectively achieves 3661 µmol·h-1·g-1 and 0.18% in the polar wood chip system under 300 W Xe lamp illumination. This study provides a sustainable and cost-effective method for the direct transformation of renewable lignocellulosic biomass to H2 fuel driven by solar energy.