ABSTRACT
Apelin is a key hormone in cardiovascular homeostasis that activates the apelin receptor (APLNR), which is regarded as a promising therapeutic target for cardiovascular disease. However, adverse effects through the ß-arrestin pathway limit its pharmacological use. Here, we report cryoelectron microscopy (cryo-EM) structures of APLNR-Gi1 complexes bound to three agonists with divergent signaling profiles. Combined with functional assays, we have identified "twin hotspots" in APLNR as key determinants for signaling bias, guiding the rational design of two exclusive G-protein-biased agonists WN353 and WN561. Cryo-EM structures of WN353- and WN561-stimulated APLNR-G protein complexes further confirm that the designed ligands adopt the desired poses. Pathophysiological experiments have provided evidence that WN561 demonstrates superior therapeutic effects against cardiac hypertrophy and reduced adverse effects compared with the established APLNR agonists. In summary, our designed APLNR modulator may facilitate the development of next-generation cardiovascular medications.
Subject(s)
Apelin Receptors , Cardiovascular Agents , Drug Design , Apelin Receptors/agonists , Apelin Receptors/chemistry , Apelin Receptors/ultrastructure , Cryoelectron Microscopy , GTP-Binding Proteins/metabolism , Receptors, G-Protein-Coupled/metabolism , Signal Transduction , Humans , Cardiovascular Agents/chemistryABSTRACT
Cannabis activates the cannabinoid receptor 1 (CB1), which elicits analgesic and emotion regulation benefits, along with adverse effects, via Gi and ß-arrestin signaling pathways. However, the lack of understanding of the mechanism of ß-arrestin-1 (ßarr1) coupling and signaling bias has hindered drug development targeting CB1. Here, we present the high-resolution cryo-electron microscopy structure of CB1-ßarr1 complex bound to the synthetic cannabinoid MDMB-Fubinaca (FUB), revealing notable differences in the transducer pocket and ligand-binding site compared with the Gi protein complex. ßarr1 occupies a wider transducer pocket promoting substantial outward movement of the TM6 and distinctive twin toggle switch rearrangements, whereas FUB adopts a different pose, inserting more deeply than the Gi-coupled state, suggesting the allosteric correlation between the orthosteric binding pocket and the partner protein site. Taken together, our findings unravel the molecular mechanism of signaling bias toward CB1, facilitating the development of CB1 agonists.
Subject(s)
Arrestin , Receptor, Cannabinoid, CB1 , Signal Transduction , Arrestin/metabolism , beta-Arrestin 1/metabolism , beta-Arrestins/metabolism , Cryoelectron Microscopy , Receptor, Cannabinoid, CB1/metabolism , Humans , Animals , Cell LineABSTRACT
The D1- and D2-dopamine receptors (D1R and D2R), which signal through Gs and Gi, respectively, represent the principal stimulatory and inhibitory dopamine receptors in the central nervous system. D1R and D2R also represent the main therapeutic targets for Parkinson's disease, schizophrenia, and many other neuropsychiatric disorders, and insight into their signaling is essential for understanding both therapeutic and side effects of dopaminergic drugs. Here, we report four cryoelectron microscopy (cryo-EM) structures of D1R-Gs and D2R-Gi signaling complexes with selective and non-selective dopamine agonists, including two currently used anti-Parkinson's disease drugs, apomorphine and bromocriptine. These structures, together with mutagenesis studies, reveal the conserved binding mode of dopamine agonists, the unique pocket topology underlying ligand selectivity, the conformational changes in receptor activation, and potential structural determinants for G protein-coupling selectivity. These results provide both a molecular understanding of dopamine signaling and multiple structural templates for drug design targeting the dopaminergic system.
Subject(s)
Receptors, Dopamine D1/chemistry , Receptors, Dopamine D1/metabolism , Receptors, Dopamine D2/chemistry , Receptors, Dopamine D2/metabolism , Signal Transduction , 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine/analogs & derivatives , 2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine/pharmacology , Amino Acid Sequence , Conserved Sequence , Cryoelectron Microscopy , Cyclic AMP/metabolism , GTP-Binding Proteins/metabolism , HEK293 Cells , Humans , Ligands , Models, Molecular , Mutant Proteins/chemistry , Mutant Proteins/metabolism , Receptors, Adrenergic, beta-2/metabolism , Receptors, Dopamine D1/ultrastructure , Receptors, Dopamine D2/ultrastructure , Structural Homology, ProteinABSTRACT
Physical or mental stress leads to neuroplasticity in the brain and increases the risk of depression and anxiety. Stress exposure causes the dysfunction of peripheral T lymphocytes. However, the pathological role and underlying regulatory mechanism of peripheral T lymphocytes in mood disorders have not been well established. Here, we show that the lack of CD4+ T cells protects mice from stress-induced anxiety-like behavior. Physical stress-induced leukotriene B4 triggers severe mitochondrial fission in CD4+ T cells, which further leads to a variety of behavioral abnormalities including anxiety, depression, and social disorders. Metabolomic profiles and single-cell transcriptome reveal that CD4+ T cell-derived xanthine acts on oligodendrocytes in the left amygdala via adenosine receptor A1. Mitochondrial fission promotes the de novo synthesis of purine via interferon regulatory factor 1 accumulation in CD4+ T cells. Our study implicates a critical link between a purine metabolic disorder in CD4+ T cells and stress-driven anxiety-like behavior.
Subject(s)
Anxiety/metabolism , Behavior, Animal/physiology , Brain Diseases, Metabolic/metabolism , Stress, Psychological/metabolism , Amygdala/metabolism , Amygdala/pathology , Animals , Anxiety/genetics , Anxiety/immunology , Anxiety/physiopathology , Brain Diseases, Metabolic/genetics , Brain Diseases, Metabolic/physiopathology , CD4-Positive T-Lymphocytes/metabolism , CD4-Positive T-Lymphocytes/pathology , Disease Models, Animal , Humans , Mice , Mitochondrial Dynamics/genetics , Oligodendroglia/metabolism , Oligodendroglia/pathology , Single-Cell Analysis , Stress, Psychological/genetics , Stress, Psychological/physiopathology , Transcriptome/genetics , Xanthine/metabolismABSTRACT
Triggering receptor expressed on myeloid cells 2 (TREM2) is strongly linked to Alzheimer's disease (AD) risk, but its functions are not fully understood. Here, we found that TREM2 specifically attenuated the activation of classical complement cascade via high-affinity binding to its initiator C1q. In the human AD brains, the formation of TREM2-C1q complexes was detected, and the increased density of the complexes was associated with lower deposition of C3 but higher amounts of synaptic proteins. In mice expressing mutant human tau, Trem2 haploinsufficiency increased complement-mediated microglial engulfment of synapses and accelerated synaptic loss. Administration of a 41-amino-acid TREM2 peptide, which we identified to be responsible for TREM2 binding to C1q, rescued synaptic impairments in AD mouse models. We thus demonstrate a critical role for microglial TREM2 in restricting complement-mediated synaptic elimination during neurodegeneration, providing mechanistic insights into the protective roles of TREM2 against AD pathogenesis.
Subject(s)
Alzheimer Disease , Complement C1q , Mice , Animals , Humans , Complement C1q/genetics , Complement C1q/metabolism , Brain/metabolism , Synapses/metabolism , Complement Activation , Microglia/metabolism , Alzheimer Disease/complications , Alzheimer Disease/metabolism , Alzheimer Disease/pathology , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Receptors, Immunologic/genetics , Receptors, Immunologic/metabolismABSTRACT
Adhesion G protein-coupled receptors (aGPCRs) are evolutionarily ancient receptors involved in a variety of physiological and pathophysiological processes. Modulators of aGPCR, particularly antagonists, hold therapeutic promise for diseases like cancer and immune and neurological disorders. Hindered by the inactive state structural information, our understanding of antagonist development and aGPCR activation faces challenges. Here, we report the cryo-electron microscopy structures of human CD97, a prototypical aGPCR that plays crucial roles in immune system, in its inactive apo and G13-bound fully active states. Compared with other family GPCRs, CD97 adopts a compact inactive conformation with a constrained ligand pocket. Activation induces significant conformational changes for both extracellular and intracellular sides, creating larger cavities for Stachel sequence binding and G13 engagement. Integrated with functional and metadynamics analyses, our study provides significant mechanistic insights into the activation and signaling of aGPCRs, paving the way for future drug discovery efforts.
Subject(s)
Antigens, CD , Receptors, G-Protein-Coupled , Signal Transduction , Humans , Cell Adhesion , Cryoelectron Microscopy , Platelet Glycoprotein GPIb-IX Complex , Receptors, G-Protein-Coupled/chemistry , Receptors, G-Protein-Coupled/metabolism , Antigens, CD/chemistry , Antigens, CD/metabolismABSTRACT
The adaptor CARD9 functions downstream of C-type lectin receptors (CLRs) for the sensing of microbial infection, which leads to responses by the TH1 and TH17 subsets of helper T cells. The single-nucleotide polymorphism rs4077515 at CARD9 in the human genome, which results in the substitution S12N (CARD9S12N), is associated with several autoimmune diseases. However, the function of CARD9S12N has remained unknown. Here we generated CARD9S12N knock-in mice and found that CARD9S12N facilitated the induction of type 2 immune responses after engagement of CLRs. Mechanistically, CARD9S12N mediated CLR-induced activation of the non-canonical transcription factor NF-κB subunit RelB, which initiated production of the cytokine IL-5 in alveolar macrophages for the recruitment of eosinophils to drive TH2 cell-mediated allergic responses. We identified the homozygous CARD9 mutation encoding S12N in patients with allergic bronchopulmonary aspergillosis and revealed activation of RelB and production of IL-5 in peripheral blood mononuclear cells from these patients. Our study provides genetic and functional evidence demonstrating that CARD9S12N can turn alveolar macrophages into IL-5-producing cells and facilitates TH2 cell-mediated pathologic responses.
Subject(s)
Aspergillosis, Allergic Bronchopulmonary/immunology , CARD Signaling Adaptor Proteins/immunology , Interleukin-5/biosynthesis , Macrophages, Alveolar/immunology , Th2 Cells/immunology , Animals , Aspergillosis, Allergic Bronchopulmonary/genetics , CARD Signaling Adaptor Proteins/genetics , Humans , Interleukin-5/immunology , Macrophages, Alveolar/metabolism , Mice , Polymorphism, Single Nucleotide , Signal Transduction/immunologyABSTRACT
Inflammatory bowel disease (IBD) mainly includes Crohn's disease (CD) and ulcerative colitis (UC). Immune disorders play an essential role in the pathogenesis of these two IBDs, but the differences in the immune microenvironment of the colon and their underlying mechanisms remain poorly investigated. Here we examined the immunological features and metabolic microenvironment of untreated individuals with IBD by multiomics analyses. Modulation of CD-specific metabolites, particularly reduced selenium, can obviously shape type 1 T helper (Th1) cell differentiation, which is specifically enriched in CD. Selenium supplementation suppressed the symptoms and onset of CD and Th1 cell differentiation via selenoprotein W (SELW)-mediated cellular reactive oxygen species scavenging. SELW promoted purine salvage pathways and inhibited one-carbon metabolism by recruiting an E3 ubiquitin ligase, tripartite motif-containing protein 21, which controlled the stability of serine hydroxymethyltransferase 2. Our work highlights selenium as an essential regulator of T cell responses and potential therapeutic targets in CD.
Subject(s)
Antioxidants/pharmacology , Crohn Disease/drug therapy , Crohn Disease/immunology , Selenium/pharmacology , Selenoprotein W/metabolism , Th1 Cells/cytology , Cell Differentiation/immunology , Cell Polarity , Colon/immunology , Colon/pathology , Glycine Hydroxymethyltransferase/metabolism , Humans , Reactive Oxygen Species/metabolism , Ribonucleoproteins/metabolism , Th1 Cells/immunology , Ubiquitin-Protein Ligases/metabolismABSTRACT
Serotonin (or 5-hydroxytryptamine, 5-HT) is an important neurotransmitter that activates 12 different G protein-coupled receptors (GPCRs) through selective coupling of Gs, Gi, or Gq proteins. The structural basis for G protein subtype selectivity by these GPCRs remains elusive. Here, we report the structures of the serotonin receptors 5-HT4, 5-HT6, and 5-HT7 with Gs, and 5-HT4 with Gi1. The structures reveal that transmembrane helices TM5 and TM6 alternate lengths as a macro-switch to determine receptor's selectivity for Gs and Gi, respectively. We find that the macro-switch by the TM5-TM6 length is shared by class A GPCR-G protein structures. Furthermore, we discover specific residues within TM5 and TM6 that function as micro-switches to form specific interactions with Gs or Gi. Together, these results present a common mechanism of Gs versus Gi protein coupling selectivity or promiscuity by class A GPCRs and extend the basis of ligand recognition at serotonin receptors.
Subject(s)
Receptors, G-Protein-Coupled , Serotonin , GTP-Binding Proteins/metabolism , Ligands , Receptors, G-Protein-Coupled/metabolism , Receptors, Serotonin/genetics , Receptors, Serotonin/metabolismABSTRACT
The dopamine system, including five dopamine receptors (D1R-D5R), plays essential roles in the central nervous system (CNS), and ligands that activate dopamine receptors have been used to treat many neuropsychiatric disorders. Here, we report two cryo-EM structures of human D3R in complex with an inhibitory G protein and bound to the D3R-selective agonists PD128907 and pramipexole, the latter of which is used to treat patients with Parkinson's disease. The structures reveal agonist binding modes distinct from the antagonist-bound D3R structure and conformational signatures for ligand-induced receptor activation. Mutagenesis and homology modeling illuminate determinants of ligand specificity across dopamine receptors and the mechanisms for Gi protein coupling. Collectively our work reveals the basis of agonist binding and ligand-induced receptor activation and provides structural templates for designing specific ligands to treat CNS diseases targeting the dopaminergic system.
Subject(s)
Cryoelectron Microscopy , GTP-Binding Protein alpha Subunits, Gi-Go/chemistry , Models, Molecular , Multiprotein Complexes/ultrastructure , Receptors, Dopamine D3/chemistry , Benzopyrans/chemistry , HEK293 Cells , Humans , Multiprotein Complexes/chemistry , Oxazines/chemistry , Pramipexole/chemistry , Protein Domains , Structure-Activity RelationshipABSTRACT
Corticotropin-releasing factor (CRF) and the three related peptides urocortins 1-3 (UCN1-UCN3) are endocrine hormones that control the stress responses by activating CRF1R and CRF2R, two members of class B G-protein-coupled receptors (GPCRs). Here, we present two cryoelectron microscopy (cryo-EM) structures of UCN1-bound CRF1R and CRF2R with the stimulatory G protein. In both structures, UCN1 adopts a single straight helix with its N terminus dipped into the receptor transmembrane bundle. Although the peptide-binding residues in CRF1R and CRF2R are different from other members of class B GPCRs, the residues involved in receptor activation and G protein coupling are conserved. In addition, both structures reveal bound cholesterol molecules to the receptor transmembrane helices. Our structures define the basis of ligand-binding specificity in the CRF receptor-hormone system, establish a common mechanism of class B GPCR activation and G protein coupling, and provide a paradigm for studying membrane protein-lipid interactions for class B GPCRs.
Subject(s)
Receptors, Corticotropin-Releasing Hormone/ultrastructure , Amino Acid Sequence , Binding Sites , Corticotropin-Releasing Hormone , Cryoelectron Microscopy/methods , GTP-Binding Protein alpha Subunits, Gs/metabolism , GTP-Binding Proteins/metabolism , Humans , Peptides/metabolism , Receptors, Corticotropin-Releasing Hormone/metabolism , Urocortins/metabolismABSTRACT
G-protein-coupled receptors (GPCRs) have central roles in intercellular communication1,2. Structural studies have revealed how GPCRs can activate G proteins. However, whether this mechanism is conserved among all classes of GPCR remains unknown. Here we report the structure of the class-C heterodimeric GABAB receptor, which is activated by the inhibitory transmitter GABA, in its active form complexed with Gi1 protein. We found that a single G protein interacts with the GB2 subunit of the GABAB receptor at a site that mainly involves intracellular loop 2 on the side of the transmembrane domain. This is in contrast to the G protein binding in a central cavity, as has been observed with other classes of GPCR. This binding mode results from the active form of the transmembrane domain of this GABAB receptor being different from that of other GPCRs, as it shows no outside movement of transmembrane helix 6. Our work also provides details of the inter- and intra-subunit changes that link agonist binding to G-protein activation in this heterodimeric complex.
Subject(s)
GTP-Binding Proteins/chemistry , Receptors, GABA-B/chemistry , Cryoelectron Microscopy , Humans , Protein Binding , Protein Domains , Protein Multimerization , Protein Structure, TertiaryABSTRACT
Serotonin, or 5-hydroxytryptamine (5-HT), is an important neurotransmitter1,2 that activates the largest subtype family of G-protein-coupled receptors3. Drugs that target 5-HT1A, 5-HT1D, 5-HT1E and other 5-HT receptors are used to treat numerous disorders4. 5-HT receptors have high levels of basal activity and are subject to regulation by lipids, but the structural basis for the lipid regulation and basal activation of these receptors and the pan-agonism of 5-HT remains unclear. Here we report five structures of 5-HT receptor-G-protein complexes: 5-HT1A in the apo state, bound to 5-HT or bound to the antipsychotic drug aripiprazole; 5-HT1D bound to 5-HT; and 5-HT1E in complex with a 5-HT1E- and 5-HT1F-selective agonist, BRL-54443. Notably, the phospholipid phosphatidylinositol 4-phosphate is present at the G-protein-5-HT1A interface, and is able to increase 5-HT1A-mediated G-protein activity. The receptor transmembrane domain is surrounded by cholesterol molecules-particularly in the case of 5-HT1A, in which cholesterol molecules are directly involved in shaping the ligand-binding pocket that determines the specificity for aripiprazol. Within the ligand-binding pocket of apo-5-HT1A are structured water molecules that mimic 5-HT to activate the receptor. Together, our results address a long-standing question of how lipids and water molecules regulate G-protein-coupled receptors, reveal how 5-HT acts as a pan-agonist, and identify the determinants of drug recognition in 5-HT receptors.
Subject(s)
Cryoelectron Microscopy , Ligands , Lipids , Receptors, Serotonin, 5-HT1/metabolism , Receptors, Serotonin, 5-HT1/ultrastructure , Apoproteins/chemistry , Apoproteins/metabolism , Apoproteins/ultrastructure , Aripiprazole/metabolism , Aripiprazole/pharmacology , Binding Sites , Cholesterol/pharmacology , Heterotrimeric GTP-Binding Proteins/chemistry , Heterotrimeric GTP-Binding Proteins/metabolism , Heterotrimeric GTP-Binding Proteins/ultrastructure , Humans , Models, Molecular , Phosphatidylinositol Phosphates/chemistry , Phosphatidylinositol Phosphates/metabolism , Phosphatidylinositol Phosphates/pharmacology , Receptor, Serotonin, 5-HT1A/chemistry , Receptor, Serotonin, 5-HT1A/metabolism , Receptor, Serotonin, 5-HT1A/ultrastructure , Receptors, Serotonin, 5-HT1/chemistry , Serotonin 5-HT1 Receptor Agonists/chemistry , Serotonin 5-HT1 Receptor Agonists/metabolism , Serotonin 5-HT1 Receptor Agonists/pharmacology , Water/chemistryABSTRACT
Adhesion G-protein-coupled receptors (GPCRs) are a major family of GPCRs, but limited knowledge of their ligand regulation or structure is available1-3. Here we report that glucocorticoid stress hormones activate adhesion G-protein-coupled receptor G3 (ADGRG3; also known as GPR97)4-6, a prototypical adhesion GPCR. The cryo-electron microscopy structures of GPR97-Go complexes bound to the anti-inflammatory drug beclomethasone or the steroid hormone cortisol revealed that glucocorticoids bind to a pocket within the transmembrane domain. The steroidal core of glucocorticoids is packed against the 'toggle switch' residue W6.53, which senses the binding of a ligand and induces activation of the receptor. Active GPR97 uses a quaternary core and HLY motif to fasten the seven-transmembrane bundle and to mediate G protein coupling. The cytoplasmic side of GPR97 has an open cavity, where all three intracellular loops interact with the Go protein, contributing to the high basal activity of GRP97. Palmitoylation at the cytosolic tail of the Go protein was found to be essential for efficient engagement with GPR97 but is not observed in other solved GPCR complex structures. Our work provides a structural basis for ligand binding to the seven-transmembrane domain of an adhesion GPCR and subsequent G protein coupling.
Subject(s)
Cryoelectron Microscopy , GTP-Binding Protein alpha Subunits, Gi-Go/chemistry , GTP-Binding Protein alpha Subunits, Gi-Go/metabolism , Glucocorticoids/chemistry , Glucocorticoids/metabolism , Receptors, G-Protein-Coupled/chemistry , Receptors, G-Protein-Coupled/ultrastructure , Binding Sites , GTP-Binding Protein alpha Subunits, Gi-Go/ultrastructure , Humans , Ligands , Lipoylation , Models, Molecular , Protein Binding , Receptors, G-Protein-Coupled/metabolismABSTRACT
BACKGROUND: Varicose veins (VV) are one of the common human diseases, but the role of genetics in its development is not fully understood. METHODS: We conducted an exome-wide association study of VV using whole-exome sequencing data from the UK Biobank, and focused on common and rare variants using single-variant association analysis and gene-level collapsing analysis. FINDINGS: A total of 13,823,269 autosomal genetic variants were obtained after quality control. We identified 36 VV-related independent common variants mapping to 34 genes by single-variant analysis and three rare variant genes (PIEZO1, ECE1, FBLN7) by collapsing analysis, and most associations between genes and VV were replicated in FinnGen. PIEZO1 was the closest gene associated with VV (P = 5.05 × 10-31), and it was found to reach exome-wide significance in both single-variant and collapsing analyses. Two novel rare variant genes (ECE1 and METTL21A) associated with VV were identified, of which METTL21A was associated only with females. The pleiotropic effects of VV-related genes suggested that body size, inflammation, and pulmonary function are strongly associated with the development of VV. CONCLUSIONS: Our findings highlight the importance of causal genes for VV and provide new directions for treatment.
Subject(s)
Exome Sequencing , Exome , Genetic Predisposition to Disease , Genome-Wide Association Study , Varicose Veins , Humans , Varicose Veins/genetics , Female , Male , Exome/genetics , Polymorphism, Single Nucleotide , Endothelin-Converting Enzymes/genetics , Middle Aged , Genetic Variation , Adult , Ion ChannelsABSTRACT
Selective macroautophagy of the endoplasmic reticulum (ER) and the nucleus, known as ER-phagy and nucleophagy, respectively, are processes whose mechanisms remain inadequately understood. Through an imaging-based screen, we find that in the fission yeast Schizosaccharomyces pombe, Yep1 (also known as Hva22 or Rop1), the ortholog of human REEP1-4, is essential for ER-phagy and nucleophagy but not for bulk autophagy. In the absence of Yep1, the initial phase of ER-phagy and nucleophagy proceeds normally, with the ER-phagy/nucleophagy receptor Epr1 coassembling with Atg8. However, ER-phagy/nucleophagy cargos fail to reach the vacuole. Instead, nucleus- and cortical-ER-derived membrane structures not enclosed within autophagosomes accumulate in the cytoplasm. Intriguingly, the outer membranes of nucleus-derived structures remain continuous with the nuclear envelope-ER network, suggesting a possible outer membrane fission defect during cargo separation from source compartments. We find that the ER-phagy role of Yep1 relies on its abilities to self-interact and shape membranes and requires its C-terminal amphipathic helices. Moreover, we show that human REEP1-4 and budding yeast Atg40 can functionally substitute for Yep1 in ER-phagy, and Atg40 is a divergent ortholog of Yep1 and REEP1-4. Our findings uncover an unexpected mechanism governing the autophagosomal enclosure of ER-phagy/nucleophagy cargos and shed new light on the functions and evolution of REEP family proteins.
Subject(s)
Schizosaccharomyces , Humans , Schizosaccharomyces/genetics , Schizosaccharomyces/metabolism , Autophagy/genetics , Endoplasmic Reticulum/metabolism , Autophagosomes/metabolism , Autophagy-Related Protein 8 Family/genetics , Autophagy-Related Protein 8 Family/metabolism , Endoplasmic Reticulum Stress , Membrane Transport Proteins/metabolismABSTRACT
The G-protein-coupled bile acid receptor (GPBAR) conveys the cross-membrane signalling of a vast variety of bile acids and is a signalling hub in the liver-bile acid-microbiota-metabolism axis1-3. Here we report the cryo-electron microscopy structures of GPBAR-Gs complexes stabilized by either the high-affinity P3954 or the semisynthesized bile acid derivative INT-7771,3 at 3 Å resolution. These structures revealed a large oval pocket that contains several polar groups positioned to accommodate the amphipathic cholic core of bile acids, a fingerprint of key residues to recognize diverse bile acids in the orthosteric site, a putative second bile acid-binding site with allosteric properties and structural features that contribute to bias properties. Moreover, GPBAR undertakes an atypical mode of activation and G protein coupling that features a different set of key residues connecting the ligand-binding pocket to the Gs-coupling site, and a specific interaction motif that is localized in intracellular loop 3. Overall, our study not only reveals unique structural features of GPBAR that are involved in bile acid recognition and allosteric effects, but also suggests the presence of distinct connecting mechanisms between the ligand-binding pocket and the G-protein-binding site in the G-protein-coupled receptor superfamily.
Subject(s)
Bile Acids and Salts/metabolism , Cryoelectron Microscopy , Receptors, G-Protein-Coupled/metabolism , Receptors, G-Protein-Coupled/ultrastructure , Allosteric Regulation/drug effects , Bile Acids and Salts/chemistry , Binding Sites/drug effects , Cholic Acids/chemistry , Cholic Acids/pharmacology , GTP-Binding Protein alpha Subunits, Gs/chemistry , GTP-Binding Protein alpha Subunits, Gs/metabolism , GTP-Binding Protein alpha Subunits, Gs/ultrastructure , Humans , Ligands , Models, Molecular , Protein Binding , Receptors, G-Protein-Coupled/agonists , Receptors, G-Protein-Coupled/chemistry , Substrate SpecificityABSTRACT
Microorganisms encode most of the functions of life on Earth. However, conventional research has primarily focused on specific environments such as humans, soil and oceans, leaving the distribution of functional families throughout the global biosphere poorly comprehended. Here, we present the database of the global distribution of prokaryotic protein families (GDPF, http://bioinfo.qd.sdu.edu.cn/GDPF/), a data resource on the distribution of functional families across the global biosphere. GDPF provides global distribution information for 36 334 protein families, 19 734 superfamilies and 12 089 KEGG (Kyoto Encyclopedia of Genes and Genomes) orthologs from multiple source databases, covering typical environments such as soil, oceans, animals, plants and sediments. Users can browse, search and download the distribution data of each entry in 10 000 global microbial communities, as well as conduct comparative analysis of distribution disparities among multiple entries across various environments. The GDPF data resource contributes to uncovering the geographical distribution patterns, key influencing factors and macroecological principles of microbial functions at a global level, thereby promoting research in Earth ecology and human health.
Subject(s)
Ecology , Prokaryotic Cells , Proteins , Animals , Humans , Soil , Multigene Family , Proteins/geneticsABSTRACT
Liver can sense the nutrient status and send signals to other organs to regulate overall metabolic homoeostasis. Herein, we demonstrate that ketone bodies act as signals released from the liver that specifically determine the distribution of excess lipid in epididymal white adipose tissue (eWAT) when exposed to a ketogenic diet (KD). An acute KD can immediately result in excess lipid deposition in the liver. Subsequently, the liver sends the ketone body ß-hydroxybutyrate (BHB) to regulate white adipose expansion, including adipogenesis and lipogenesis, to alleviate hepatic lipid accumulation. When ketone bodies are depleted by deleting 3-hydroxy-3-methylglutaryl-CoA synthase 2 gene in the liver, the enhanced lipid deposition in eWAT but not in inguinal white adipose tissue is preferentially blocked, while lipid accumulation in liver is not alleviated. Mechanistically, ketone body BHB can significantly decrease lysine acetylation of peroxisome proliferator-activated receptor gamma in eWAT, causing enhanced activity of peroxisome proliferator-activated receptor gamma, the key adipogenic transcription factor. These observations suggest that the liver senses metabolic stress first and sends a corresponding signal, that is, ketone body BHB, to specifically promote eWAT expansion to adapt to metabolic challenges.
Subject(s)
Adipose Tissue, White , Diet, Ketogenic , Fatty Liver , Ketone Bodies , Humans , Adipose Tissue, White/metabolism , Fatty Liver/metabolism , Ketone Bodies/metabolism , Lipids , Liver/metabolism , PPAR gamma/metabolismABSTRACT
Fish possess a powerful IFN system to defend against aquatic virus infections. Nevertheless, spring viremia of carp virus (SVCV) causes large-scale mortality in common carp and significant economic losses to aquaculture. Therefore, it is necessary to investigate the strategies used by SVCV to escape the IFN response. In this study, we show that the SVCV nucleoprotein (N protein) negatively regulates cellular IFN production by degrading stimulator of IFN genes (STING) via the autophagy-lysosome-dependent pathway. First, overexpression of N protein inhibited the IFN promoter activation induced by polyinosinic-polycytidylic acid and STING. Second, the N protein associated with STING and experiments using a dominant-negative STING mutant demonstrated that the N-terminal transmembrane domains of STING were indispensable for this interaction. Then, the N protein degraded STING in a dose-dependent and autophagy-lysosome-dependent manner. Intriguingly, in the absence of STING, individual N proteins could not elicit host autophagic flow. Furthermore, the autophagy factor Beclin1 was found to interact with the N protein to attenuate N protein-mediated STING degradation after beclin1 knockdown. Finally, the N protein remarkably weakened STING-enhanced cellular antiviral responses. These findings reveal that SVCV uses the host autophagic process to achieve immune escape, thus broadening our understanding of aquatic virus pathogenesis.