ABSTRACT
Our understanding of sex and gender evolves. We asked scientists about their work and the future of sex and gender research. They discuss, among other things, interdisciplinary collaboration, moving beyond binary conceptualizations, accounting for intersecting factors, reproductive strategies, expanding research on sex-related differences, and sex's dynamic nature.
Subject(s)
Biomedical Research , Gender Identity , Sex , Female , Humans , Male , Sex CharacteristicsABSTRACT
All living things experience an increase in entropy, manifested as a loss of genetic and epigenetic information. In yeast, epigenetic information is lost over time due to the relocalization of chromatin-modifying proteins to DNA breaks, causing cells to lose their identity, a hallmark of yeast aging. Using a system called "ICE" (inducible changes to the epigenome), we find that the act of faithful DNA repair advances aging at physiological, cognitive, and molecular levels, including erosion of the epigenetic landscape, cellular exdifferentiation, senescence, and advancement of the DNA methylation clock, which can be reversed by OSK-mediated rejuvenation. These data are consistent with the information theory of aging, which states that a loss of epigenetic information is a reversible cause of aging.
Subject(s)
Aging , Epigenesis, Genetic , Animals , Aging/genetics , DNA Methylation , Epigenome , Mammals/genetics , Nucleoproteins , Saccharomyces cerevisiae/geneticsABSTRACT
Pediatric SARS-CoV-2 vaccines are needed that elicit immunity directly in the airways as well as systemically. Building on pediatric parainfluenza virus vaccines in clinical development, we generated a live-attenuated parainfluenza-virus-vectored vaccine candidate expressing SARS-CoV-2 prefusion-stabilized spike (S) protein (B/HPIV3/S-6P) and evaluated its immunogenicity and protective efficacy in rhesus macaques. A single intranasal/intratracheal dose of B/HPIV3/S-6P induced strong S-specific airway mucosal immunoglobulin A (IgA) and IgG responses. High levels of S-specific antibodies were also induced in serum, which efficiently neutralized SARS-CoV-2 variants of concern of alpha, beta, and delta lineages, while their ability to neutralize Omicron sub-lineages was lower. Furthermore, B/HPIV3/S-6P induced robust systemic and pulmonary S-specific CD4+ and CD8+ T cell responses, including tissue-resident memory cells in the lungs. Following challenge, SARS-CoV-2 replication was undetectable in airways and lung tissues of immunized macaques. B/HPIV3/S-6P will be evaluated clinically as pediatric intranasal SARS-CoV-2/parainfluenza virus type 3 vaccine.
Subject(s)
COVID-19 Vaccines , COVID-19 , Animals , Humans , Antibodies, Neutralizing , Antibodies, Viral , Macaca mulatta , COVID-19/prevention & control , SARS-CoV-2/geneticsABSTRACT
The cholesterol-sensing protein Scap induces cholesterol synthesis by transporting membrane-bound transcription factors called sterol regulatory element-binding proteins (SREBPs) from the endoplasmic reticulum (ER) to the Golgi apparatus for proteolytic activation. Transport requires interaction between Scap's two ER luminal loops (L1 and L7), which flank an intramembrane sterol-sensing domain (SSD). Cholesterol inhibits Scap transport by binding to L1, which triggers Scap's binding to Insig, an ER retention protein. Here we used cryoelectron microscopy (cryo-EM) to elucidate two structures of full-length chicken Scap: (1) a wild-type free of Insigs and (2) mutant Scap bound to chicken Insig without cholesterol. Strikingly, L1 and L7 intertwine tightly to form a globular domain that acts as a luminal platform connecting the SSD to the rest of Scap. In the presence of Insig, this platform undergoes a large rotation accompanied by rearrangement of Scap's transmembrane helices. We postulate that this conformational change halts Scap transport of SREBPs and inhibits cholesterol synthesis.
Subject(s)
Cholesterol/metabolism , Membrane Proteins/chemistry , Membrane Proteins/metabolism , Amino Acid Sequence , Animals , Antibodies/metabolism , Chickens , Membrane Proteins/isolation & purification , Membrane Proteins/ultrastructure , Models, Biological , Models, Molecular , Protein Binding , Protein Domains , Protein Structure, Secondary , Structure-Activity RelationshipABSTRACT
TANK binding kinase 1 (TBK1) regulates IFN-I, NF-κB, and TNF-induced RIPK1-dependent cell death (RCD). In mice, biallelic loss of TBK1 is embryonically lethal. We discovered four humans, ages 32, 26, 7, and 8 from three unrelated consanguineous families with homozygous loss-of-function mutations in TBK1. All four patients suffer from chronic and systemic autoinflammation, but not severe viral infections. We demonstrate that TBK1 loss results in hypomorphic but sufficient IFN-I induction via RIG-I/MDA5, while the system retains near intact IL-6 induction through NF-κB. Autoinflammation is driven by TNF-induced RCD as patient-derived fibroblasts experienced higher rates of necroptosis in vitro, and CC3 was elevated in peripheral blood ex vivo. Treatment with anti-TNF dampened the baseline circulating inflammatory profile and ameliorated the clinical condition in vivo. These findings highlight the plasticity of the IFN-I response and underscore a cardinal role for TBK1 in the regulation of RCD.
Subject(s)
Inflammation/enzymology , Protein Serine-Threonine Kinases/deficiency , Tumor Necrosis Factor-alpha/pharmacology , A549 Cells , Adaptor Proteins, Signal Transducing/metabolism , Apoptosis , Autoimmunity/drug effects , Brain/diagnostic imaging , Cell Death/drug effects , Cytokines/metabolism , Deubiquitinating Enzyme CYLD/metabolism , Female , HEK293 Cells , Homozygote , Humans , I-kappa B Kinase/metabolism , Immunophenotyping , Inflammation/pathology , Interferon Type I/metabolism , Interferon-gamma/metabolism , Loss of Function Mutation/genetics , Male , Pedigree , Phosphorylation/drug effects , Protein Serine-Threonine Kinases/genetics , Protein Serine-Threonine Kinases/metabolism , Receptor-Interacting Protein Serine-Threonine Kinases/metabolism , Receptors, Pattern Recognition/metabolism , Toll-Like Receptor 3/metabolism , Transcriptome/genetics , Vesiculovirus/drug effects , Vesiculovirus/physiologyABSTRACT
Inflammation is an important component of fibrosis but immune processes that orchestrate kidney fibrosis are not well understood. Here we apply single-cell sequencing to a mouse model of kidney fibrosis. We identify a subset of kidney tubule cells with a profibrotic-inflammatory phenotype characterized by the expression of cytokines and chemokines associated with immune cell recruitment. Receptor-ligand interaction analysis and experimental validation indicate that CXCL1 secreted by profibrotic tubules recruits CXCR2+ basophils. In mice, these basophils are an important source of interleukin-6 and recruitment of the TH17 subset of helper T cells. Genetic deletion or antibody-based depletion of basophils results in reduced renal fibrosis. Human kidney single-cell, bulk gene expression and immunostaining validate a function for basophils in patients with kidney fibrosis. Collectively, these studies identify basophils as contributors to the development of renal fibrosis and suggest that targeting these cells might be a useful clinical strategy to manage chronic kidney disease.
Subject(s)
Basophils , Renal Insufficiency, Chronic , Animals , Fibrosis , Humans , Kidney/metabolism , Kidney Tubules , Mice , Renal Insufficiency, Chronic/metabolism , Single-Cell AnalysisABSTRACT
Clear-cell renal cell carcinoma (ccRCC) exhibits a broad range of metastatic phenotypes that have not been systematically studied to date. Here, we analyzed 575 primary and 335 metastatic biopsies across 100 patients with metastatic ccRCC, including two cases sampledat post-mortem. Metastatic competence was afforded by chromosome complexity, and we identify 9p loss as a highly selected event driving metastasis and ccRCC-related mortality (p = 0.0014). Distinct patterns of metastatic dissemination were observed, including rapid progression to multiple tissue sites seeded by primary tumors of monoclonal structure. By contrast, we observed attenuated progression in cases characterized by high primary tumor heterogeneity, with metastatic competence acquired gradually and initial progression to solitary metastasis. Finally, we observed early divergence of primitive ancestral clones and protracted latency of up to two decades as a feature of pancreatic metastases.
Subject(s)
Carcinoma, Renal Cell/genetics , Carcinoma, Renal Cell/pathology , Kidney Neoplasms/genetics , Kidney Neoplasms/pathology , Mutation , Neoplasm Metastasis , Adult , Aged , Aged, 80 and over , Biomarkers/metabolism , Biopsy , Chromosome Mapping , Chromosomes, Human, Pair 14 , Chromosomes, Human, Pair 9 , Disease Progression , Female , Humans , Longitudinal Studies , Male , Middle Aged , Phenotype , Prospective Studies , Thrombosis , Treatment OutcomeABSTRACT
Foxp3+ regulatory T (Treg) cells expressing the interleukin (IL)-33 receptor ST2 mediate tissue repair in response to IL-33. Whether Treg cells also respond to the alarmin IL-33 to regulate specific aspects of the immune response is not known. Here we describe an unexpected function of ST2+ Treg cells in suppressing the innate immune response in the lung to environmental allergens without altering the adaptive immune response. Following allergen exposure, ST2+ Treg cells were activated by IL-33 to suppress IL-17-producing γδ T cells. ST2 signaling in Treg cells induced Ebi3, a component of the heterodimeric cytokine IL-35 that was required for Treg cell-mediated suppression of γδ T cells. This response resulted in fewer eosinophil-attracting chemokines and reduced eosinophil recruitment into the lung, which was beneficial to the host in reducing allergen-induced inflammation. Thus, we define a fundamental role for ST2+ Treg cells in the lung as a negative regulator of the early innate γδ T cell response to mucosal injury.
Subject(s)
Immunomodulation , Interleukin-33/metabolism , Intraepithelial Lymphocytes/immunology , Intraepithelial Lymphocytes/metabolism , Lung/immunology , Lung/metabolism , T-Lymphocytes, Regulatory/immunology , T-Lymphocytes, Regulatory/metabolism , Allergens/immunology , Animals , Biomarkers , Immunophenotyping , Interleukin-1 Receptor-Like 1 Protein/metabolism , Leukocytes/immunology , Leukocytes/metabolism , MiceABSTRACT
Mutations that impact immune cell migration and result in immune deficiency illustrate the importance of cell movement in host defense. In humans, loss-of-function mutations in DOCK8, a guanine exchange factor involved in hematopoietic cell migration, lead to immunodeficiency and, paradoxically, allergic disease. Here, we demonstrate that, like humans, Dock8-/- mice have a profound type 2 CD4+ helper T (TH2) cell bias upon pulmonary infection with Cryptococcus neoformans and other non-TH2 stimuli. We found that recruited Dock8-/-CX3CR1+ mononuclear phagocytes are exquisitely sensitive to migration-induced cell shattering, releasing interleukin (IL)-1ß that drives granulocyte-macrophage colony-stimulating factor (GM-CSF) production by CD4+ T cells. Blocking IL-1ß, GM-CSF or caspase activation eliminated the type-2 skew in mice lacking Dock8. Notably, treatment of infected wild-type mice with apoptotic cells significantly increased GM-CSF production and TH2 cell differentiation. This reveals an important role for cell death in driving type 2 signals during infection, which may have implications for understanding the etiology of type 2 CD4+ T cell responses in allergic disease.
Subject(s)
Guanine Nucleotide Exchange Factors/deficiency , Lymphocyte Activation/genetics , Lymphocyte Activation/immunology , Th2 Cells/immunology , Th2 Cells/metabolism , Animals , Biomarkers , Caspases/metabolism , Cell Movement/genetics , Cell Movement/immunology , Cytokines/genetics , Cytokines/metabolism , Disease Susceptibility , Gene Expression , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Humans , Immunophenotyping , Leukocytes, Mononuclear/immunology , Leukocytes, Mononuclear/metabolism , Mice , Mice, Knockout , Myeloid Cells/immunology , Myeloid Cells/metabolism , Phagocytes/immunology , Phagocytes/metabolism , Signal TransductionABSTRACT
Familial Mediterranean fever (FMF) is an autoinflammatory disease caused by homozygous or compound heterozygous gain-of-function mutations in MEFV, which encodes pyrin, an inflammasome protein. Heterozygous carrier frequencies for multiple MEFV mutations are high in several Mediterranean populations, suggesting that they confer selective advantage. Among 2,313 Turkish people, we found extended haplotype homozygosity flanking FMF-associated mutations, indicating evolutionarily recent positive selection of FMF-associated mutations. Two pathogenic pyrin variants independently arose >1,800 years ago. Mutant pyrin interacts less avidly with Yersinia pestis virulence factor YopM than with wild-type human pyrin, thereby attenuating YopM-induced interleukin (IL)-1ß suppression. Relative to healthy controls, leukocytes from patients with FMF harboring homozygous or compound heterozygous mutations and from asymptomatic heterozygous carriers released heightened IL-1ß specifically in response to Y. pestis. Y. pestis-infected MefvM680I/M680I FMF knock-in mice exhibited IL-1-dependent increased survival relative to wild-type knock-in mice. Thus, FMF mutations that were positively selected in Mediterranean populations confer heightened resistance to Y. pestis.
Subject(s)
Disease Resistance/genetics , Familial Mediterranean Fever/genetics , Plague , Pyrin/genetics , Selection, Genetic/genetics , Animals , Bacterial Outer Membrane Proteins/immunology , Bacterial Outer Membrane Proteins/metabolism , Disease Resistance/immunology , Haplotypes , Humans , Inflammasomes/immunology , Inflammasomes/metabolism , Mice , Mice, Inbred C57BL , Mutation , Plague/immunology , Plague/metabolism , Pyrin/immunology , Pyrin/metabolism , Turkey , Virulence Factors/immunology , Virulence Factors/metabolism , Yersinia pestisABSTRACT
Friedreich's ataxia (FA) is a debilitating, multisystemic disease caused by the depletion of frataxin (FXN), a mitochondrial iron-sulfur (Fe-S) cluster biogenesis factor. To understand the cellular pathogenesis of FA, we performed quantitative proteomics in FXN-deficient human cells. Nearly every annotated Fe-S cluster-containing protein was depleted, indicating that as a rule, cluster binding confers stability to Fe-S proteins. We also observed depletion of a small mitoribosomal assembly factor METTL17 and evidence of impaired mitochondrial translation. Using comparative sequence analysis, mutagenesis, biochemistry, and cryoelectron microscopy, we show that METTL17 binds to the mitoribosomal small subunit during late assembly and harbors a previously unrecognized [Fe4S4]2+ cluster required for its stability. METTL17 overexpression rescued the mitochondrial translation and bioenergetic defects, but not the cellular growth, of FXN-depleted cells. These findings suggest that METTL17 acts as an Fe-S cluster checkpoint, promoting translation of Fe-S cluster-rich oxidative phosphorylation (OXPHOS) proteins only when Fe-S cofactors are replete.
Subject(s)
Friedreich Ataxia , Iron-Sulfur Proteins , Humans , Iron-Sulfur Proteins/genetics , Iron-Sulfur Proteins/metabolism , Cryoelectron Microscopy , Frataxin , Protein Biosynthesis , Mitochondria/genetics , Mitochondria/metabolism , Friedreich Ataxia/metabolism , Methyltransferases/genetics , Methyltransferases/metabolismABSTRACT
Ligand binding usually moves the target protein from an ensemble of inactive states to a well-defined active conformation. Matthies et al. flip this scheme around, finding that, for the magnesium channel CorA, loss of ligand binding induces an ensemble of conformations that turn the channel on.
Subject(s)
Bacterial Proteins/ultrastructure , Cation Transport Proteins/ultrastructure , Magnesium/metabolism , Thermotoga maritima/chemistryABSTRACT
Genes encode components of coevolved and interconnected networks. The effect of genotype on phenotype therefore depends on genotypic context through gene interactions known as epistasis. Epistasis is important in predicting phenotype from genotype for an individual. It is also examined in population studies to identify genetic risk factors in complex traits and to predict evolution under selection. Paradoxically, the effects of genotypic context in individuals and populations are distinct and sometimes contradictory. We argue that predicting genotype from phenotype for individuals based on population studies is difficult and, especially in human genetics, likely to result in underestimating the effects of genotypic context.
Subject(s)
Epistasis, Genetic , Genotype , Animals , Genetics, Medical , Genetics, Population , Humans , Quantitative Trait, HeritableABSTRACT
Voltage-gated ion channels (VGICs) are outfitted with diverse cytoplasmic domains that impact function. To examine how such elements may affect VGIC behavior, we addressed how the bacterial voltage-gated sodium channel (BacNa(V)) C-terminal cytoplasmic domain (CTD) affects function. Our studies show that the BacNa(V) CTD exerts a profound influence on gating through a temperature-dependent unfolding transition in a discrete cytoplasmic domain, the neck domain, proximal to the pore. Structural and functional studies establish that the BacNa(V) CTD comprises a bi-partite four-helix bundle that bears an unusual hydrophilic core whose integrity is central to the unfolding mechanism and that couples directly to the channel activation gate. Together, our findings define a general principle for how the widespread four-helix bundle cytoplasmic domain architecture can control VGIC responses, uncover a mechanism underlying the diverse BacNa(V) voltage dependencies, and demonstrate that a discrete domain can encode the temperature-dependent response of a channel.
Subject(s)
Bacterial Proteins/chemistry , Gammaproteobacteria/metabolism , Voltage-Gated Sodium Channels/chemistry , Amino Acid Sequence , Electron Spin Resonance Spectroscopy , Models, Molecular , Molecular Sequence Data , Protein Structure, Tertiary , Protein Unfolding , Sequence AlignmentABSTRACT
Type VI secretion systems (T6SSs) are newly identified contractile nanomachines that translocate effector proteins across bacterial membranes. The Francisella pathogenicity island, required for bacterial phagosome escape, intracellular replication, and virulence, was presumed to encode a T6SS-like apparatus. Here, we experimentally confirm the identity of this T6SS and, by cryo electron microscopy (cryoEM), show the structure of its post-contraction sheath at 3.7 Å resolution. We demonstrate the assembly of this T6SS by IglA/IglB and secretion of its putative effector proteins in response to environmental stimuli. The sheath has a quaternary structure with handedness opposite that of contracted sheath of T4 phage tail and is organized in an interlaced two-dimensional array by means of ß sheet augmentation. By structure-based mutagenesis, we show that this interlacing is essential to secretion, phagosomal escape, and intracellular replication. Our atomic model of the T6SS will facilitate design of drugs targeting this highly prevalent secretion apparatus.
Subject(s)
Bacterial Proteins/chemistry , Bacterial Secretion Systems , Francisella/ultrastructure , Bacterial Proteins/ultrastructure , Bacteriophage T4/chemistry , Bacteriophages/chemistry , Cryoelectron Microscopy , Models, Molecular , Protein Structure, SecondaryABSTRACT
The stem cells that maintain and repair the postnatal skeleton remain undefined. One model suggests that perisinusoidal mesenchymal stem cells (MSCs) give rise to osteoblasts, chondrocytes, marrow stromal cells, and adipocytes, although the existence of these cells has not been proven through fate-mapping experiments. We demonstrate here that expression of the bone morphogenetic protein (BMP) antagonist gremlin 1 defines a population of osteochondroreticular (OCR) stem cells in the bone marrow. OCR stem cells self-renew and generate osteoblasts, chondrocytes, and reticular marrow stromal cells, but not adipocytes. OCR stem cells are concentrated within the metaphysis of long bones not in the perisinusoidal space and are needed for bone development, bone remodeling, and fracture repair. Grem1 expression also identifies intestinal reticular stem cells (iRSCs) that are cells of origin for the periepithelial intestinal mesenchymal sheath. Grem1 expression identifies distinct connective tissue stem cells in both the bone (OCR stem cells) and the intestine (iRSCs).
Subject(s)
Bone and Bones/cytology , Intercellular Signaling Peptides and Proteins/metabolism , Intestine, Small/cytology , Mesenchymal Stem Cells/cytology , Animals , Cartilage/metabolism , Intestine, Small/metabolism , Mesenchymal Stem Cells/metabolism , Mice , Mice, Inbred C57BLABSTRACT
The genetic code of living cells has been reprogrammed to enable the site-specific incorporation of hundreds of non-canonical amino acids into proteins, and the encoded synthesis of non-canonical polymers and macrocyclic peptides and depsipeptides1-3. Current methods for engineering orthogonal aminoacyl-tRNA synthetases to acylate new monomers, as required for the expansion and reprogramming of the genetic code, rely on translational readouts and therefore require the monomers to be ribosomal substrates4-6. Orthogonal synthetases cannot be evolved to acylate orthogonal tRNAs with non-canonical monomers (ncMs) that are poor ribosomal substrates, and ribosomes cannot be evolved to polymerize ncMs that cannot be acylated onto orthogonal tRNAs-this co-dependence creates an evolutionary deadlock that has essentially restricted the scope of translation in living cells to α-L-amino acids and closely related hydroxy acids. Here we break this deadlock by developing tRNA display, which enables direct, rapid and scalable selection for orthogonal synthetases that selectively acylate their cognate orthogonal tRNAs with ncMs in Escherichia coli, independent of whether the ncMs are ribosomal substrates. Using tRNA display, we directly select orthogonal synthetases that specifically acylate their cognate orthogonal tRNA with eight non-canonical amino acids and eight ncMs, including several ß-amino acids, α,α-disubstituted-amino acids and ß-hydroxy acids. We build on these advances to demonstrate the genetically encoded, site-specific cellular incorporation of ß-amino acids and α,α-disubstituted amino acids into a protein, and thereby expand the chemical scope of the genetic code to new classes of monomers.
Subject(s)
Amino Acids , Amino Acyl-tRNA Synthetases , Escherichia coli , Genetic Code , RNA, Transfer , Acylation , Amino Acids/chemistry , Amino Acids/metabolism , Amino Acyl-tRNA Synthetases/chemistry , Amino Acyl-tRNA Synthetases/genetics , Amino Acyl-tRNA Synthetases/metabolism , Genetic Code/genetics , Hydroxy Acids/chemistry , Hydroxy Acids/metabolism , RNA, Transfer/chemistry , RNA, Transfer/genetics , RNA, Transfer/metabolism , Substrate Specificity , Ribosomes/metabolism , Escherichia coli/enzymology , Escherichia coli/genetics , Escherichia coli/metabolismABSTRACT
Humans have the remarkable cognitive capacity to rapidly adapt to changing environments. Central to this capacity is the ability to form high-level, abstract representations that take advantage of regularities in the world to support generalization1. However, little is known about how these representations are encoded in populations of neurons, how they emerge through learning and how they relate to behaviour2,3. Here we characterized the representational geometry of populations of neurons (single units) recorded in the hippocampus, amygdala, medial frontal cortex and ventral temporal cortex of neurosurgical patients performing an inferential reasoning task. We found that only the neural representations formed in the hippocampus simultaneously encode several task variables in an abstract, or disentangled, format. This representational geometry is uniquely observed after patients learn to perform inference, and consists of disentangled directly observable and discovered latent task variables. Learning to perform inference by trial and error or through verbal instructions led to the formation of hippocampal representations with similar geometric properties. The observed relation between representational format and inference behaviour suggests that abstract and disentangled representational geometries are important for complex cognition.