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While beige adipocytes have been found to associate with dense sympathetic neurites in mouse inguinal subcutaneous white fat (iWAT), little is known about when and how this patterning is established. Here, we applied whole-tissue imaging to examine the development of sympathetic innervation in iWAT. We found that parenchymal neurites actively grow between postnatal day 6 (P6) and P28, overlapping with early postnatal beige adipogenesis. Constitutive deletion of Prdm16 in adipocytes led to a significant reduction in early postnatal beige adipocytes and sympathetic density within this window. Using an inducible, adipocyte-specific Prdm16 knockout model, we found that Prdm16 is required for guiding sympathetic growth during early development. Deleting Prdm16 in adult animals, however, did not affect sympathetic structure in iWAT. Together, these findings highlight that beige adipocyte-sympathetic neurite communication is crucial to establish sympathetic structure during the early postnatal period but may be dispensable for its maintenance in mature animals.

The progression and metastatic capacity of solid tumors are strongly influenced by immune cells in the tumor microenvironment. In non-small cell lung cancer (NSCLC), accumulation of anti-inflammatory tumor-associated macrophages (TAM) is associated with worse clinical outcome and resistance to therapy. Here we investigated the immune landscape of NSCLC in the presence of protumoral TAMs expressing the macrophage receptor with collagenous structure (MARCO). MARCO-expressing TAM numbers correlated with increased occurrence of regulatory T cells and effector T cells and decreased natural killer (NK) cells in these tumors. Furthermore, transcriptomic data from the tumors uncovered a correlation between MARCO expression and the antiinflammatory cytokine IL37. In vitro studies subsequently showed that lung cancer cells polarized macrophages to express MARCO and gain an immune-suppressive phenotype through the release of IL37. MARCO-expressing TAMs blocked cytotoxic T-cell and NK-cell activation, inhibiting their proliferation, cytokine production, and tumor killing capacity. Mechanistically, MARCO(+) macrophages enhanced regulatory T (Treg) cell proliferation and IL10 production and diminished CD8 T-cell activities. Targeting MARCO or IL37 receptor (IL37R) by antibody or CRISPR knockout of IL37 in lung cancer cell lines repolarized TAMs, resulting in recovered cytolytic activity and antitumoral capacity of NK cells and T cells and down-modulated Treg cell activities. In summary, our data demonstrate a novel immune therapeutic approach targeting human TAMs immune suppression of NK- and T-cell antitumor activities. Significance: This study defines tumor-derived IL37 and the macrophage scavenger receptor MARCO as potential therapeutic targets to remodel the immune-suppressive microenvimnment in patients with lung cancer.

The mushroom body (MB) is a well-characterized associative memory structure within the Drosophila brain. Analyzing MB connectivity using multiple approaches is critical for understanding the functional implications of this structure. Using the genetic anterograde transsynaptic tracing tool, trans-Tango, we identified divergent projections across the brain and convergent downstream targets of the MB output neurons (MBONs). Our analysis revealed at least three separate targets that receive convergent input from MBONs: other MBONs, the fan-shaped body (FSB), and the lateral accessory lobe (LAL). We describe, both anatomically and functionally, a multilayer circuit in which inhibitory and excitatory MBONs converge on the same genetic subset of FSB and LAL neurons. This circuit architecture enables the brain to update and integrate information with previous experience before executing appropriate behavioral responses. Our use of trans-Tango provides a genetically accessible anatomical framework for investigating the functional relevance of components within these complex and interconnected circuits.

Stem cells maintain tissues by balancing self-renewal with differentiation. A stem cell's local microenvironment, or niche, informs stem cell behavior and receives inputs at multiple levels. Increasingly, it is becoming clear that the overall metabolic status of an organism or metabolites themselves can function as integral members of the niche to alter stem cell fate. Macroscopic dietary interventions such as caloric restriction, the ketogenic diet, and a high-fat diet systemically alter an organism's metabolic state in different ways. Intriguingly, however, they all converge on a propensity to enhance self-renewal. Here, we highlight our current knowledge on how dietary changes feed into stem cell behavior across a wide variety of tissues and illuminate possible explanations for why diverse interventions can result in similar stem cell phenotypes. In so doing, we hope to inspire new avenues of inquiry into the importance of metabolism in stem cell homeostasis and disease.

In the type III CRISPR-Cas immune response of prokaryotes, infection triggers the production of cyclic oligoadenylates that bind and activate proteins that contain a CARF domain(1,2). Many type III loci are associated with proteins in which the CRISPR-associated Rossman fold (CARF) domain is fused to a restriction endonuclease-like domain(3,4). However, with the exception of the well-characterized Csm6 and Csx1 ribonucleases(5,6), whether and how these inducible effectors provide defence is not known. Here we investigated a type III CRISPR accessory protein, which we name cyclic-oligoadenylate-activated single-stranded ribonuclease and single-stranded deoxyribonuclease 1 (Card1). Card1 forms a symmetrical dimer that has a large central cavity between its CRISPR-associated Rossmann fold and restriction endonuclease domains that binds cyclic tetra-adenylate. The binding of ligand results in a conformational change comprising the rotation of individual monomers relative to each other to form a more compact dimeric scaffold, in which a manganese cation coordinates the catalytic residues and activates the cleavage of single-stranded-but not double-stranded-nucleic acids (both DNA and RNA). In vivo, activation of Card1 induces dormancy of the infected hosts to provide immunity against phage infection and plasmids. Our results highlight the diversity of strategies used in CRISPR systems to provide immunity. Structural analyses of the type III CRISPR accessory protein Card1, which induces dormancy in infected hosts to provide immunity against phage infection, reveal the mechanisms by which it cleaves single-stranded RNA and DNA.

Recent advances in single-particle cryogenic electron microscopy (cryo-EM) have enabled the structural determination of numerous protein assemblies at high resolution, yielding unprecedented insights into their function. However, despite its extraordinary capabilities, cryo-EM remains time-consuming and resource-intensive. It is therefore beneficial to have a means for rapidly assessing and optimizing the quality of samples prior to lengthy cryo-EM analyses. To do this, we have developed a native mass spectrometry (nMS) platform that provides rapid feedback on sample quality and highly streamlined biochemical screening. Because nMS enables accurate mass analysis of protein complexes, it is well suited to routine evaluation of the composition, integrity, and homogeneity of samples prior to their plunge-freezing on EM grids. We demonstrate the utility of our nMS-based platform for facilitating cryo-EM studies using structural characterizations of exemplar bacterial transcription complexes as well as the replication-transcription assembly from the SARS-CoV-2 virus that is responsible for the COVID-19 pandemic.

Notch signaling is critical for controlling a variety of cell fate decisions during metazoan development and homeostasis. This unique, highly conserved signaling pathway relies on cell-to-cell contact, which triggers the proteolytic release of the cytoplasmic domain of the membrane-anchored transcription factor Notch from the membrane. A disintegrin and metalloproteinase (ADAM) proteins are crucial for Notch activation by processing its S2 site. While ADAM10 cleaves Notch1 under physiological, ligand-dependent conditions, ADAM17 mainly cleaves Notch1 under ligand-independent conditions. However, the mechanism(s) that regulate the distinct contributions of these ADAMs in Notch processing remain unclear. Using cell-based assays in mouse embryonic fibroblasts (mEFs) lacking ADAM10 and/or ADAM17, we aimed to clarify what determines the relative contributions of ADAM10 and ADAM17 to ligand-dependent or ligand-independent Notch processing. We found that EDTA-stimulated ADAM17-dependent Notch1 processing is rapid and requires the ADAM17-regulators iRhom1 and iRhom2, whereas the Delta-like 4-induced ligand-dependent Notch1 processing is slower and requires ADAM10. The selectivity of ADAM17 for EDTA-induced Notch1 processing can most likely be explained by a preference for ADAM17 over ADAM10 for the Notch1 cleavage site and by the stronger inhibition of ADAM10 by EDTA. The physiological ADAM10-dependent processing of Notch1 cannot be compensated for by ADAM17 in Adam10-/- mEFs, or by other ADAMs shown here to be able to cleave the Notch1 cleavage site, such as ADAMs9, 12, and 19. Collectively, these results provide new insights into the mechanisms underlying the substrate selectivity of ADAM10 and ADAM17 towards Notch1.

Background Bimekizumab is a monoclonal IgG1 antibody that selectively inhibits interleukin (IL)-17F in addition to IL-17A. This study investigated the efficacy and safety of bimekizumab in patients with moderate to severe plaque psoriasis, the effects of treatment withdrawal, and two maintenance dosing schedules over 56 weeks. Methods BE READY was a phase 3, multicentre, randomised, double-blind, placebo-controlled trial done at 77 sites (hospitals, clinics, private doctor's practices, and dedicated clinical research centres) in nine countries across Asia, Australia, Europe, and North America. Adult patients aged 18 years or older with moderate to severe plaque psoriasis were stratified by region and previous biologic exposure, and randomly assigned (4:1) to receive bimekizumab 320 mg every 4 weeks or placebo every 4 weeks by use of interactive response technology. Coprimary endpoints were the proportion of patients achieving 90% or greater improvement from baseline in the Psoriasis Area Severity Index (PASI90) and the proportion of patients achieving a score of 0 (clear) or 1 (almost clear) on the five-point Investigator's Global Assessment (IGA) scale at week 16 (non-responder imputation). Bimekizumab-treated patients achieving PASI90 at week 16 were re-allocated (1:1:1) to receive bimekizumab 320 mg every 4 weeks, every 8 weeks, or placebo for weeks 16-56. Efficacy analyses were done in the intention-to-treat population; the safety analysis set comprised all patients who received at least one dose of study treatment. This trial is registered with ClinicalTrials.gov (NCT03410992), and is now completed. Findings Between Feb 5, 2018, and Jan 7, 2020, 435 patients were randomly assigned to receive either bimekizumab 320 mg every 4 weeks (n=349) or placebo every 4 weeks (n=86). Coprimary endpoints were met: at week 16, 317 (91%) of 349 patients receiving bimekizumab 320 mg every 4 weeks achieved PASI90, compared with one (1%) of 86 patients receiving placebo (risk difference 89.8 [95% CI 86.1-93.4]; p<0.0001); and 323 (93%) of 349 patients receiving bimekizumab 320 mg every 4 weeks achieved an IGA score of 0 or 1 versus one (1%) of 86 patients receiving placebo (risk difference 91.5 [95% CI 88.0-94.9]; p<0.0001). Responses were maintained through to week 56 with bimekizumab 320 mg every 8 weeks and every 4 weeks. Treatment-emergent adverse events in the initial treatment period (up to week 16) were reported in 213 (61%) of 349 patients receiving bimekizumab 320 mg every 4 weeks and 35 (41%) of 86 patients receiving placebo every 4 weeks. From week 16 to week 56, treatment-emergent adverse events were reported in 78 (74%) of 106 patients receiving bimekizumab 320 mg every 4 weeks, 77 (77%) of 100 patients receiving bimekizumab 320 mg every 8 weeks, and 72 (69%) of 105 patients receiving placebo. Interpretation Bimekizumab showed high levels of response, which were durable over 56 weeks, with both maintenance dosing schedules (every 4 weeks and every 8 weeks). Moreover, bimekizumab was well tolerated, with no unexpected safety findings. Data presented here further support the therapeutic value of bimekizumab and inhibition of IL-17F in addition to IL-17A for patients with moderate to severe plaque psoriasis.

Mendelian Susceptibility to Mycobacterial diseases (MSMD) are a group of innate immune defects with more than 17 genes and 32 clinical phenotypes identified. Defects in the IFN-gamma mediated immunity lead to an increased susceptibility to intracellular pathogens like mycobacteria including attenuated Mycobacterium bovis-Bacillus Calmette-Guerin (BCG) vaccine strains and non-tuberculous environmental mycobacteria (NTM), Salmonella, fungi, parasites like Leishmania and some viruses, in otherwise healthy individuals. Mutations in the IL12R beta 1 gene are the commonest genetic defects identified. This retrospective study reports the clinical, immunological, and molecular characteristics of a cohort of 55 MSMD patients from 10 centers across India. Mycobacterial infection was confirmed by GeneXpert, Histopathology, and acid fast bacilli staining. Immunological workup included lymphocyte subset analysis, Nitro blue tetrazolium (NBT) test, immunoglobulin levels, and flow-cytometric evaluation of the IFN-gamma mediated immunity. Genetic analysis was done by next generation sequencing (NGS). Disseminated BCG-osis was the commonest presenting manifestation (82%) with a median age of presentation of 6 months due to the practice of BCG vaccination at birth. This was followed by infection with Salmonella and non-typhi Salmonella (13%), Cytomegalovirus (CMO (11%), Candida (7%), NTM (4%), and Histoplasma (2%). Thirty-six percent of patients in cohort were infected by more than one organism. This study is the largest cohort of MSMD patients reported from India to the best of our knowledge and we highlight the importance of work up for IL-12/IL-23/ISG15/IFN-gamma circuit in all patients with BCG-osis and suspected MSMD irrespective of age.

Neural network computations are usually assumed to emerge from patterns of fast electrical activity. Challenging this view, we show that a male fly's decision to persist in mating hinges on a biochemical computation that enables processing over minutes to hours. Each neuron in a recurrent network contains slightly different internal molecular estimates of mating progress. Protein kinase A (PKA) activity contrasts this internal measurement with input from the other neurons to represent accumulated evidence that the goal of the network has been achieved. When consensus is reached, PKA pushes the network toward a large-scale and synchronized burst of calcium influx that we call an eruption. Eruptions transform continuous deliberation within the network into an all-or-nothing output, after which the male will no longer sacrifice his life to continue mating. Here, biochemical activity, invisible to most large-scale recording techniques, is the key computational currency directing behavior and motivational state.