A hallmark of neuroinflammatory disorders is mitochondrial dysfunction. Nevertheless, the transcriptional changes underlying this alteration are not well-defined. Microglia activation, a decrease in mitochondrion biogenesis and a subsequent alteration of the redox are common factors in diseases coursing with neuroinflammation. In the last two decades, components of the adenosinergic system have been proposed as potential therapeutic targets to combat neuroinflammation. In this research, we analyzed by RNAseq the gene expression in activated microglia treated with an adenosine A2A receptor antagonist, SCH 582561, and/or an A3 receptor agonist, 2-Cl-IB-MECA, since these receptors are deeply related to neurodegeneration and inflammation. The analysis was focused on genes related to inflammation and REDOX homeostasis. It was detected that in the three conditions (microglia treated with 2-Cl-IB-MECA, SCH 582561, and the combination) more than 40 % of the detected genes codified by the mitochondrial genome were differentially expressed (FDR < 0.05) (14/34, 16/34, and 13/34) respectively, being almost all of them (>85 %) upregulated in the microglia treated with adenosinergic compounds. Also, we analyzed the differential expression of genes related to mitochondrial function and oxidative stress codified by the nuclear genome. Additionally, we evaluated the oxygen consumption rate (OCR) of mitochondria in microglia treated with LPS and IFN-γ, both alone and in combination with adenosinergic compounds. The data showed an improvement in mitochondrial function with the antagonist of the adenosine A2A receptor, compared to the effects of pro-inflammatory stimulus, confirming a functional effect consistent with the RNAseq data.

Guidelines for the management of obesity and type 2 diabetes (T2DM) emphasize the importance of lifestyle changes, including a reduced-calorie diet and increased physical activity. However, for many people, these changes can be difficult to maintain over the long term. Medication options are already available to treat obesity, which can help reduce appetite and/or reduce caloric intake. Incretin-based peptides exert their effect through G-protein-coupled receptors, the receptors for glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), and glucagon peptide hormones are important regulators of insulin secretion and energy metabolism. Understanding the role of intercellular signaling pathways and inflammatory processes is essential for the development of effective pharmacological agents in obesity. GLP-1 receptor agonists have been successfully used, but it is assumed that their effectiveness may be limited by desensitization and downregulation of the target receptor. A growing number of new agents acting on incretin hormones are becoming available for everyday clinical practice, including oral GLP-1 receptor agonists, the dual GLP-1/GIP receptor agonist tirzepatide, and other dual and triple GLP-1/GIP/glucagon receptor agonists, which may show further significant therapeutic potential. This narrative review summarizes the therapeutic effects of different incretin hormones and presents future prospects in the treatment of T2DM and obesity.

Xenorhabdus nematophila is a symbiotic Gammaproteobacterium that produces diverse natural products that facilitate mutualistic and pathogenic interactions in their nematode and insect hosts, respectively. The interplay between X. nematophila secondary metabolism and symbiosis stage is tuned by various global regulators. An example of such a regulator is the LysR-type protein transcription factor LrhA, which regulates amino acid metabolism and is necessary for virulence in insects and normal nematode progeny production. Here, we utilized comparative metabolomics and molecular networking to identify small molecule factors regulated by LrhA and characterized a rare γ-ketoacid (GKA) and two new N-acyl amides, GKA-Arg (1) and GKA-Pro (2) which harbor a γ-keto acyl appendage. A lrhA null mutant produced elevated levels of compound 1 and reduced levels of compound 2 relative to wild type. N-acyl amides 1 and 2 were shown to be selective agonists for the human G-protein-coupled receptors (GPCRs) C3AR1 and CHRM2, respectively. The CHRM2 agonist 2 deleteriously affected the hatch rate and length of Steinernema nematodes. This work further highlights the utility of exploiting regulators of host-bacteria interactions for the identification of the bioactive small molecule signals that they control.
OBJECTIVE: Xenorhabdus bacteria are of interest due to their symbiotic relationship with Steinernema nematodes and their ability to produce a variety of natural bioactive compounds. Despite their importance, the regulatory hierarchy connecting specific natural products and their regulators is poorly understood. In this study, comparative metabolomic profiling was utilized to identify the secondary metabolites modulated by the X. nematophila global regulator LrhA. This analysis led to the discovery of three metabolites, including an N-acyl amide that inhibited the egg hatching rate and length of Steinernema carpocapsae nematodes. These findings support the notion that X. nematophila LrhA influences the symbiosis between X. nematophila and S. carpocapsae through N-acyl amide signaling. A deeper understanding of the regulatory hierarchy of these natural products could contribute to a better comprehension of the symbiotic relationship between X. nematophila and S. carpocapsae.

Yeast expression of human G-protein-coupled receptors (GPCRs) can be used as a biosensor platform for the detection of pharmaceuticals. Cannabinoid receptor type 1 (CB1R) is of particular interest, given the cornucopia of natural and synthetic cannabinoids being explored as therapeutics. We show for the first time that engineering the N-terminus of CB1R allows for efficient signal transduction in yeast, and that engineering the sterol composition of the yeast membrane modulates its performance. Using an engineered cannabinoid biosensor, we demonstrate that large libraries of synthetic cannabinoids and terpenes can be quickly screened to elucidate known and novel structure-activity relationships. The biosensor strains offer a ready platform for evaluating the activity of new synthetic cannabinoids, monitoring drugs of abuse, and developing therapeutic molecules.

The primary cilium is a dynamic subcellular compartment templated from the mother centriole or basal body. Cilia are solitary and tiny, but remarkably consequential in cellular pathways regulating proliferation, differentiation, and maintenance. Multiple transmembrane proteins such as G-protein-coupled receptors, channels, enzymes, and membrane-associated lipidated proteins are enriched in the ciliary membrane. The precise regulation of ciliary membrane content is essential for effective signal transduction and maintenance of tissue homeostasis. Surprisingly, a few conserved molecular factors, intraflagellar transport complex A and the tubby family adapter protein TULP3, mediate the transport of most membrane cargoes into cilia. Recent advances in cryogenic electron microscopy provide fundamental insights into these molecular players. Here, we review the molecular players mediating cargo delivery into the ciliary membrane through the lens of structural biology. These mechanistic insights into ciliary transport provide a framework for understanding of disease variants in ciliopathies, enable precise manipulation of cilia-mediated pathways, and provide a platform for the development of targeted therapeutics.

Chemokine receptors are integral to the immune system and prime targets in drug discovery that have undergone extensive structural elucidation in recent years. We outline a timeline of these structural achievements, discuss the intracellular negative allosteric modulation of chemokine receptors, analyze the mechanisms of orthosteric receptor activation, and report on the emerging concept of biased signaling. Additionally, we highlight differences of G-protein binding among chemokine receptors. Intracellular allosteric modulators in chemokine receptors interact with a conserved motif within transmembrane helix 7 and helix 8 and exhibit a two-fold inactivation mechanism that can be harnessed for drug-discovery efforts. Chemokine recognition is a multi-step process traditionally explained by a two-site model within chemokine recognition site 1 (CRS1) and CRS2. Recent structural studies have extended our understanding of this complex mechanism with the identification of CRS1.5 and CRS3. CRS3 is implicated in determining ligand specificity and surrounds the chemokine by almost 180°. Within CRS3 we identified the extracellular loop 2 residue 45.51 as a key interaction mediator for chemokine binding. Y2917.43 on the other hand was shown in CCR1 to be a key determinant of signaling bias which, along with specific chemokine-dependent phosphorylation ensembles at the G-protein coupled receptors (GPCR\'s) C-terminus, seems to play a pivotal role in determining the direction of signal bias in GPCRs.

Neutrophils are not only involved in immune defense against infection but also contribute to the exacerbation of tissue damage after ischemia and reperfusion. We have previously shown that genetic ablation of regulatory Gαi proteins in mice has both protective and deleterious effects on myocardial ischemia reperfusion injury (mIRI), depending on which isoform is deleted. To deepen and analyze these findings in more detail the contribution of Gαi2 proteins in resident cardiac vs circulating blood cells for mIRI was first studied in bone marrow chimeras. In fact, the absence of Gαi2 in all blood cells reduced the extent of mIRI (22,9% infarct size of area at risk (AAR) Gnai2-/- → wt vs 44.0% wt → wt; p < 0.001) whereas the absence of Gαi2 in non-hematopoietic cells increased the infarct damage (66.5% wt → Gnai2-/- vs 44.0% wt → wt; p < 0.001). Previously we have reported the impact of platelet Gαi2 for mIRI. Here, we show that infarct size was substantially reduced when Gαi2 signaling was either genetically ablated in neutrophils/macrophages using LysM-driven Cre recombinase (AAR: 17.9% Gnai2fl/fl LysM-Cre+/tg vs 42.0% Gnai2fl/fl; p < 0.01) or selectively blocked with specific antibodies directed against Gαi2 (AAR: 19.0% (anti-Gαi2) vs 49.0% (IgG); p < 0.001). In addition, the number of platelet-neutrophil complexes (PNCs) in the infarcted area were reduced in both, genetically modified (PNCs: 18 (Gnai2fl/fl; LysM-Cre+/tg) vs 31 (Gnai2fl/fl); p < 0.001) and in anti-Gαi2 antibody-treated (PNCs: 9 (anti-Gαi2) vs 33 (IgG); p < 0.001) mice. Of note, significant infarct-limiting effects were achieved with a single anti-Gαi2 antibody challenge immediately prior to vessel reperfusion without affecting bleeding time, heart rate or cellular distribution of neutrophils. Finally, anti-Gαi2 antibody treatment also inhibited transendothelial migration of human neutrophils (25,885 (IgG) vs 13,225 (anti-Gαi2) neutrophils; p < 0.001), collectively suggesting that a therapeutic concept of functional Gαi2 inhibition during thrombolysis and reperfusion in patients with myocardial infarction should be further considered.

Bioluminescence and fluorescence resonance energy transfer (BRET and FRET) together with the proximity ligation method revealed the existence of G-protein-coupled receptors, Ionotropic and Receptor tyrosine kinase heterocomplexes, e.g., A2AR-D2R, GABAA-D5R, and FGFR1-5-HT1AR heterocomplexes. Molecular integration takes place through allosteric receptor-receptor interactions in heteroreceptor complexes of synaptic and extra-synaptic regions. It involves the modulation of receptor protomer recognition, signaling and trafficking, as well as the modulation of behavioral responses. Allosteric receptor-receptor interactions in hetero-complexes give rise to concepts like meta-modulation and protein modulation. The introduction of receptor-receptor interactions was the origin of the concept of meta-modulation provided by Katz and Edwards in 1999, which stood for the fine-tuning or modulation of nerve cell transmission. In 2000-2010, Ribeiro and Sebastiao, based on a series of papers, provided strong support for their view that adenosine can meta-modulate (fine-tune) synaptic transmission through adenosine receptors. However, another term should also be considered: protein modulation, which is the key feature of allosteric receptor-receptor interactions leading to learning and consolidation by novel adapter proteins to memory. Finally, it must be underlined that allosteric receptor-receptor interactions and their involvement both in brain disease and its treatment are of high interest. Their pathophysiological relevance has been obtained, especially for major depressive disorder, cocaine use disorder, and Parkinson\'s disease.

UNASSIGNED: Chronic liver disease (CLD) remains a global health issue associated with a significant disease burden. Liver fibrosis, a hallmark of CLD, is characterised by the activation of hepatic stellate cells (HSCs) that gain profibrotic characteristics including increased production of extracellular matrix protein. Currently, no antifibrotic therapies are available clinically, in part because of the lack of HSC-specific drug targets. Here, we aimed to identify HSC-specific membrane proteins that can serve as targets for antifibrotic drug development.
UNASSIGNED: Small interfering RNA-mediated knockdown of GPR176 was used to assess the in vitro function of GPR176 in HSCs and in precision cut liver slices (PCLS). The in vivo role of GPR176 was assessed using the carbon tetrachloride (CCl4) and common bile duct ligation (BDL) models in wild-type and GPR176 knockout mice. GPR176 in human CLD was assessed by immunohistochemistry of diseased human livers and RNA expression analysis in human primary HSCs and transcriptomic data sets.
UNASSIGNED: We identified Gpr176, an orphan G-protein coupled receptor, as an HSC-enriched activation associated gene. In vitro, Gpr176 is strongly induced upon culture-induced and hepatocyte-damage-induced activation of primary HSCs. Knockdown of GPR176 in primary mouse HSCs or PCLS cultures resulted in reduced fibrogenic characteristics. Absence of GPR176 did not influence liver homeostasis, but Gpr176-/- mice developed less severe fibrosis in CCl4 and BDL fibrosis models. In humans, GPR176 expression was correlated with in vitro HSC activation and with fibrosis stage in patients with CLD.
UNASSIGNED: GPR176 is a functional protein during liver fibrosis and reducing its activity attenuates fibrogenesis. These results highlight the potential of GPR176 as an HSC-specific antifibrotic candidate to treat CLD.
UNASSIGNED: The lack of effective antifibrotic drugs is partly attributed to the insufficient knowledge about the mechanisms involved in the development of liver fibrosis. We demonstrate that the G-protein coupled receptor GPR176 contributes to fibrosis development. Since GPR176 is specifically expressed on the membrane of activated hepatic stellate cells and is linked with fibrosis progression in humans, it opens new avenues for the development of targeted interventions.

Shifts in the magnitude and nature of gut microbial metabolites have been implicated in Alzheimer\'s disease (AD), but the host receptors that sense and respond to these metabolites are largely unknown. Here, we develop a systems biology framework that integrates machine learning and multi-omics to identify molecular relationships of gut microbial metabolites with non-olfactory G-protein-coupled receptors (termed the \"GPCRome\"). We evaluate 1.09 million metabolite-protein pairs connecting 408 human GPCRs and 335 gut microbial metabolites. Using genetics-derived Mendelian randomization and integrative analyses of human brain transcriptomic and proteomic profiles, we identify orphan GPCRs (i.e., GPR84) as potential drug targets in AD and that triacanthine experimentally activates GPR84. We demonstrate that phenethylamine and agmatine significantly reduce tau hyperphosphorylation (p-tau181 and p-tau205) in AD patient induced pluripotent stem cell-derived neurons. This study demonstrates a systems biology framework to uncover the GPCR targets of human gut microbiota in AD and other complex diseases if broadly applied.