The transcriptional targets of Aire and the mechanisms controlling their expression in medullary thymic epithelial cells (mTECs) need to be clarified to understand Aire's tolerogenic function. By using a multi-omics single-cell approach coupled with deep scRNA-seq, we examined how Aire controls the transcription of a wide variety of genes in a small fraction of Aire-expressing cells. We found that chromatin repression by PRC2 is an important step for Aire to achieve stochastic gene expression. Aire unleashed the silenced chromatin configuration caused by PRC2, thereby increasing the expression of its functional targets. Besides this preconditioning for Aire's gene induction, we demonstrated that PRC2 also controls the composition of mTECs that mimic the developmental trait of peripheral tissues, i.e., mimetic cells. Of note, this action of PRC2 was independent of Aire and it was more apparent than Aire. Thus, our study uncovered the essential role of polycomb complex for Aire-mediated promiscuous gene expression and the development of mimetic cells.

Neurological illnesses are debilitating diseases that affect brain function and balance. Due to their complicated aetiologies and progressive nature, neurodegenerative and neuropsychiatric illnesses are difficult to treat. These incurable conditions damage brain functions like mobility, cognition, and emotional regulation, but medication, gene therapy, and physical therapy can manage symptoms. Disruptions in cellular signalling pathways, especially those involving oxidative stress response, memory processing, and neurotransmitter modulation, contribute to these illnesses. This review stresses the interplay between key signalling pathways involved in neurological diseases, such as the Nrf2/Keap1/HO-1/SIRT-1 axis and the p75NTR/PI3K/Akt/MAPK cascade. To protect neurons from oxidative damage and death, the Nrf2 transcription factor promotes antioxidant enzyme production. The Keap1 protein releases Nrf2 during oxidative stress for nuclear translocation and gene activation. The review also discusses how neurotrophin signalling through the p75 neurotrophin receptor (p75NTR) determines cell destiny, whether pro-survival or apoptotic. The article highlights emerging treatment approaches targeting these signalling pathways by mapping these connections. Continued research into these molecular pathways may lead to new neurological disease treatments that restore cellular function and neuronal survival. In addition to enhanced delivery technologies, specific modulators and combination therapies should be developed to fine-tune signalling responses. Understanding these crosstalk dynamics is crucial to strengthening neurological illness treatment options and quality of life.

Traditional Chinese medicine (TCM) utilizes intricate herbal formulations that exemplify the principles of compatibility and synergy. However, the rapid proliferation of herbal data has resulted in redundant information, complicating the understanding of their potential mechanisms. To address this issue, we first established a comprehensive database that encompasses 992 herbs, 18 681 molecules, and 2168 targets. Consequently, we implemented a multi-network strategy based on a core information screening method to elucidate the highly intertwined relationships among the targets of various herbs and to refine herbal target information. Within a non-redundant network framework, separation and overlap analysis demonstrated that the networking of herbs preserves essential clinical information, including their properties, meridians, and therapeutic classifications. Furthermore, two notable trends emerged from the statistical analyses of classical TCM formulas: the separation of herbs and the overlap between herbs and diseases. This phenomenon is termed the herbal combination model (HCM), validated through statistical analyses of two representative case studies: the common cold and rheumatoid arthritis. Additionally, in vivo and in vitro experiments with the new formula YanChuanQin (YanHuSuo-Corydalis Rhizoma, ChuanWu-Aconiti Radix, and QinJiao-Gentianae Macrophyllae Radix) for acute gouty arthritis further support the HCM. Overall, this computational method provides a systematic network strategy for exploring herbal combinations in complex and poorly understood diseases from a non-redundant perspective.

The use of genetically engineered tools, including combinations of Cre-LoxP and Flp-FRT systems, enables the interrogation of complex biology. Steroidogenic factor-1 (SF-1) is expressed in the ventromedial hypothalamic nucleus (VMH). Development of genetic tools, such as mice expressing Flp recombinase (Flp) in SF-1 neurons (Sf-1-Flp), will be useful for future studies that unravel the complex physiology regulated by the VMH. Here, we developed and characterized Sf-1-Flp mice and demonstrated their utility. The Flp sequence was inserted into the Sf-1 locus with P2A. This insertion did not affect Sf-1 mRNA expression levels and Sf-1-Flp mice do not have any visible phenotypes. They are fertile and metabolically comparable to wild-type littermate mice. Optogenetic stimulation using adeno-associated virus (AAV) carrying Flp-dependent channelrhodopsin-2 (ChR2) increased blood glucose and skeletal muscle PGC-1α in Sf-1-Flp mice. This was similar to SF-1 neuronal activation using Sf-1-BAC-Cre and AAV carrying Cre-dependent ChR2. Finally, we generated Sf-1-Flp mice that lack β2-adrenergic receptors (Adrb2) only in skeletal muscle with a combination of Cre/LoxP technology (Sf-1-Flp:SKMΔAdrb2). Optogenetic stimulation of SF-1 neurons failed to increase skeletal muscle PGC-1α in Sf-1-Flp:SKMΔAdrb2 mice, suggesting that Adrb2 in skeletal muscle is required for augmented skeletal muscle PGC-1α by SF-1 neuronal activation. Our data demonstrate that Sf-1-Flp mice are useful for interrogating complex physiology.

Gasdermins (GSDMs), functioning as membrane perforating proteins, can be activated by canonical inflammasomes, noncanonical inflammasomes, as well as non-inflammasomes, leading to cell pyroptosis and the subsequent release of inflammatory mediators. Increasing evidence has implicated that GSDMs are associated with chronic kidney disease (CKD), including diabetes nephropathy, lupus nephritis, obstructive nephropathy, and crystalline nephropathy. This review centers on the role of GSDMs-mediated pyroptosis in the pathogenesis of CKD, providing novel ideas for enhancing the prognosis and therapeutic strategies of CKD.

The use of genetically engineered tools, including combinations of Cre-LoxP and Flp-FRT systems, enable the interrogation of complex biology. Steroidogenic factor-1 (SF-1) is expressed in the ventromedial hypothalamic nucleus (VMH). Development of genetic tools, such as mice expressing Flp recombinase (Flp) in SF-1 neurons (Sf-1-Flp), will be useful for future studies that unravel the complex physiology regulated by the VMH. Here, we developed and characterized Sf-1-Flp mice and demonstrated its utility. Flp sequence was inserted into Sf-1 locus with P2A. This insertion did not affect Sf-1 mRNA expression levels and Sf-1-Flp mice do not have any visible phenotypes. They are fertile and metabolically comparable to wild-type littermate mice. Optogenetic stimulation using adeno-associated virus (AAV)-bearing Flp-dependent channelrhodopsin-2 (ChR2) increased blood glucose and skeletal muscle PGC-1α in Sf-1-Flp mice. This was similar to SF-1 neuronal activation using Sf-1-BAC-Cre and AAV-bearing Cre-dependent ChR2. Finally, we generated Sf-1-Flp mice that lack β2-adrenergic receptors (Adrβ2) only in skeletal muscle with a combination of Cre/LoxP technology (Sf-1-Flp::SKMΔAdrβ2). Optogenetic stimulation of SF-1 neurons failed to increase skeletal muscle PGC-1α in Sf-1-Flp::SKMΔAdrβ2 mice, suggesting that Adrβ2 in skeletal muscle is required for augmented skeletal muscle PGC-1α by SF-1 neuronal activation. Our data demonstrate that Sf-1-Flp mice are useful for interrogating complex physiology.

Lysophosphatidylserine (lyso-PS) is a potent hormone-like signaling lysophospholipid, which regulates many facets of mammalian biology and dysregulation in its metabolism is associated with several human neurological and autoimmune diseases. Despite the physiological importance and causal relation with human pathophysiology, little is known about the metabolism of lyso-PS in tissues other than the nervous and immune systems. To address this problem, here, we attempted to identify one (or more) lipase(s) capable of degrading lyso-PS in different mammalian tissues. We found that the membrane fraction of most mammalian tissues possess lyso-PS lipase activity, yet interestingly, the only bona fide lyso-PS lipase ABHD12 displays this enzymatic activity and has control over lyso-PS metabolism only in the mammalian brain. Using an in vitro inhibitor screen against membrane fractions of different tissues, we find that another lipase from the metabolic serine hydrolase family, ABHD6, is a putative lyso-PS lipase in the mouse liver and kidney. Finally, using pharmacological tools, we validate the lyso-PS lipase activity of ABHD6 in vivo, and functionally designate this enzyme as a major lyso-PS lipase in primary hepatocytes, and the mammalian liver and kidneys.

Recent success with the use of glucagon-like peptide-1 (GLP-1) receptor analogs and dipeptidyl peptidase-4 inhibitors for the treatment of patients with diabetes has highlighted the role of the intestine as an endocrine organ. Gut-derived hormones, including glucagon-like peptide-1, glucose-dependent insulinotropic polypeptide, and ghrelin, have important roles in the control of energy metabolism and food intake, and are associated with the metabolic syndrome. In this study, we isolated and identified a new intestine-derived hormone, betagenin, and showed that it stimulates insulin secretion and β-cell proliferation and suppresses β-cell apoptosis. Adenovirus-mediated expression of betagenin restored the blood glucose concentrations and hemoglobin A1c (HbA1c) levels of mice with streptozotocin-induced diabetes to normal and increased their β-cell mass. Transgenic mice overexpressing betagenin exhibited more than three-fold higher β-cell mass than WT mass, whereas that of KO mice was four-fold lower. A synthetic peptide representing the sequence of purified and secreted betagenin enhanced glucose-dependent insulin secretion in human and mouse pancreatic islets and stimulated the proliferation of the pancreatic β-cell line MIN6 through extracellular signal-regulated kinase 1/2-dependent signaling. The intravenous administration of this peptide to streptozotocin mice stimulated the proliferation of pancreatic β-cells in vivo, and the intraperitoneal administration of betagenin ameliorated diabetes and restored β-cell mass. These results indicate that betagenin may reduce blood glucose concentration and induce β-cell regeneration in patients with diabetes.

Zinc is a critical trace element that is important for various biological functions including male and female reproductive systems, but the molecular mechanisms that underlie fertility have been unclear. We show here that zinc signaling in the endometrial tissue is indispensable for successful pregnancy in mice. We observed that a uterine-specific genetic deletion of Slc39a10/Zip10, which encodes one of the zinc transporters to elevate the cytoplasmic level of zinc, results in female infertility due to failure of embryo invasion into the endometrium and subsequent embryonic loss. Zip10 mRNA is expressed in uterine tissues, especially in the decidualizing stromal cells during embryo implantation. The absence of ZIP10 leads to attenuation of progesterone-progesterone receptor (PGR) signals between the epithelium and the stroma, including abnormal expression of the PGR and its target molecules in both the epithelium and stroma in vivo. We found that depletion of intracytoplasmic zinc ions due to loss of ZIP10 disrupts the change in nuclear-to-cytoplasmic localization of GLI1, which is critical for PGR signaling in the decidualizing stromal cells in vitro not only in mice but also in humans. Our findings (i) highlight a biological relevance of ZIP10-mediated zinc homeostatic regulation in the establishment of a successful pregnancy and (ii) will help to prevent infertility in humans.

α-Synucleinopathies constitute a spectrum of neurodegenerative disorders, including Parkinson's disease (PD), Lewy body dementia (LBD), Multiple System Atrophy (MSA), and Alzheimer's disease concurrent with LBD (AD-LBD). These disorders are unified by a pathological hallmark: aberrant misfolding and accumulation of α-synuclein (α-syn). This review delves into the pivotal role of α-syn, the key agent in α-synucleinopathy pathophysiology, and provides a survey of potential therapeutics that target cell-to-cell spread of pathologic α-syn. Recognizing the intricate complexity and multifactorial etiology of α-synucleinopathy, the review illuminates the potential of various membrane receptors, proteins, intercellular spreading pathways, and pathological agents for therapeutic interventions. While significant progress has been made in understanding α-synucleinopathy, the pursuit of efficacious treatments remains challenging. Several strategies involving decreasing α-syn production and aggregation, increasing α-syn degradation, lowering extracellular α-syn, and inhibiting cellular uptake of α-syn are presented. The paper underscores the necessity of meticulous and comprehensive investigations to advance our knowledge of α-synucleinopathy pathology and ultimately develop innovative therapeutic strategies for α-synucleinopathies.

Lymphocyte activation gene 3 (LAG3) is a key receptor involved in the propagation of pathological proteins in Parkinson's disease (PD). This study investigates the role of neuronal LAG3 in mediating the binding, uptake, and propagation of α-synuclein (αSyn) preformed fibrils (PFFs). Using neuronal LAG3 conditional knockout mice and human induced pluripotent stem cells-derived dopaminergic (DA) neurons, we demonstrate that LAG3 expression is critical for pathogenic αSyn propagation. Our results show that the absence of neuronal LAG3 significantly reduces αSyn pathology, alleviates motor dysfunction, and inhibits neurodegeneration in vivo. Electrophysiological recordings revealed that αSyn PFFs induce pronounced neuronal hyperactivity in wild-type (WT) neurons, increasing firing rates in cell-attached and whole-cell configurations, and reducing miniature excitatory postsynaptic currents. In contrast, neurons lacking LAG3 resisted these electrophysiological effects. Moreover, treatment with an anti-human LAG3 antibody in human DA neurons inhibited αSyn PFFs binding and uptake, preventing pathology propagation. These findings confirm the essential function of neuronal LAG3 in mediating αSyn propagation and associated disruptions, identifying LAG3 as a potential therapeutic target for PD and related α-synucleinopathies.

House dust mite (HDM) is the leading allergen for allergic rhinitis (AR). Although allergic sensitisation by inhaled allergens renders susceptible individuals prone to developing AR, the molecular mechanisms driving this process remain incompletely elucidated.

This study aimed to elucidate the molecular mechanisms underlying HDM-induced AR.

We examined the expression of cytidine/uridine monophosphate kinase 2 (CMPK2), STING and the NLRP3 inflammasome in both AR patients and mice. Additionally, we investigated the role of CMPK2 and STING in the activation of the NLRP3 inflammasome in AR.

The expression of CMPK2, STING and the NLRP3 inflammasome was significantly increased in the nasal mucosa of AR patients compared to non-AR controls. A positive correlation was found between CMPK2 expression and the levels of STING, NLRP3, ASC, CASP1 and IL-1β. HDM treatment up-regulated the expression of CMPK2, and CMPK2 overexpression enhanced NLRP3 inflammasome activation in human nasal epithelial cells (HNEPCs). Additionally, mitochondrial reactive oxygen species (mtROS) production following HDM exposure contributed to mitochondrial dysfunction and the release of mitochondrial DNA (mtDNA), which activated the cyclic GMP-AMP synthase (cGAS)-STING pathway. Remarkably, depletion of mtDNA or inhibition of STING signalling reduced HDM-induced NLRP3 inflammasome activation in HNEPCs. In vivo, genetic knockout of CMPK2 or STING alleviated NLRP3 inflammasome activation and ameliorated clinical symptoms of AR in mice.

Our results suggest that HDM promotes the activation of NLRP3 inflammasome through the up-regulation of CMPK2 and ensuing mtDNA-STING signalling pathway, hence revealing additional therapeutic target for AR.

Cytidine/uridine monophosphate kinase 2 (CMPK2) expression is up-regulated in the nasal mucosa of patients and mice with allergic rhinitis (AR). CMPK2 caused NLRP3 inflammasome activation via mitochondrial DNA (mtDNA)-STING pathway. Blocking CMPK2 or STING signalling significantly reduced the activation of NLRP3 in house dust mite (HDM)-challenged mice and human nasal epithelial cells (HNEPCs).

Mitochondria are essential to cellular function as they generate the majority of cellular ATP, mediated through oxidative phosphorylation, which couples proton pumping of the electron transport chain (ETC) to ATP production. The ETC generates an electrochemical gradient, known as the proton motive force, consisting of the mitochondrial membrane potential (ΔΨm, the major component in mammals) and ΔpH across the inner mitochondrial membrane. Both ATP production and reactive oxygen species (ROS) are linked to ΔΨm, and it has been shown that an imbalance in ΔΨm beyond the physiological optimal intermediate range results in excessive ROS production. The reaction of cytochrome c oxidase (COX) of the ETC with its small electron donor cytochrome c (Cytc) is the proposed rate-limiting step in mammals under physiological conditions. The rate at which this redox reaction occurs controls ΔΨm and thus ATP and ROS production. Multiple mechanisms are in place that regulate this reaction to meet the cell's energy demand and respond to acute stress. COX and Cytc have been shown to be regulated by all three main mechanisms, which we discuss in detail: allosteric regulation, tissue-specific isoforms, and post-translational modifications for which we provide a comprehensive catalog and discussion of their functional role with 55 and 50 identified phosphorylation and acetylation sites on COX, respectively. Disruption of these regulatory mechanisms has been found in several common human diseases, including stroke and myocardial infarction, inflammation including sepsis, and diabetes, where changes in COX or Cytc phosphorylation lead to mitochondrial dysfunction contributing to disease pathophysiology. Identification and subsequent targeting of the underlying signaling pathways holds clear promise for future interventions to improve human health. An example intervention is the recently discovered noninvasive COX-inhibitory infrared light therapy that holds promise to transform the current standard of clinical care in disease conditions where COX regulation has gone awry.

The P4HTM gene encodes a transmembrane prolyl 4-hydroxylase, which is responsible for the degradation of hypoxia-inducible transcription factors (HIF) under normoxia. Clinically, biallelic P4HTM variants have been identified in patients with hypotonia, hypoventilation, intellectual disabilities, dysautonomia, epilepsy, and eye abnormalities (HIDEA syndrome). Seizure was one of the most prominent symptoms. However, the clinical features of patients with epilepsy associated with P4HTM variants remain unclear. In this report, we describe a one-month-old infant with HIDEA syndrome caused by compound heterozygous P4HTM variants (c.300dupG/p.Gly103Argfs*22 and c.488C > T/p.Ala163Val). The infant presented with clonic seizures of focal onset that responded well to valproate, but with profound intellectual disability and global developmental delay at the last follow-up at 3 years old. A review of the existing literature indicates that seizures in this population typically begin early in infancy, manifest in multiple types, and are relatively well controlled. Epilepsy seemed unrelated to developmental outcomes or disease progression. Valproate, which has HIF-1α inhibiting properties, may be a promising treatment avenue for this population.

Bioassay-guided fractionation approach led to identification of two novel compounds; (4-(hydroxymethyl)-3-methoxy-1H-pyrazol (1) and mycalene (2), alongside with four known metabolites; octadecane (3), hexatriacontane (4), 1-heneicosanol (5) and heptatriacontanoic acid (6) from the Red Sea marine sponge Hemimycale sp. The ethyl acetate fraction showed a noticeable cytotoxic activity against the lung cancer cell line (A549) with IC50 value of 75.54 µg/ mL. Structural elucidation was achieved using a combination of 1D and 2D nuclear magnetic resonance (NMR) spectroscopy and high-resolution electrospray ionization-mass spectrometry (HR-ESI-MS). To elucidate the potential mechanism of action behind the cytotoxic effects against lung cancer, a multi-faceted approach combining in silico network pharmacology, experimental validation, and molecular docking studies were employed. Both compounds, designated as 1 and 2, demonstrated significant binding affinities to predicted target proteins, with docking scores of -4.789 and - 4.421 kcal/mol, respectively. These results lay the groundwork for further investigation into the therapeutic potential of these novel compounds from Hemimycale sp. as promising candidates for lung cancer treatment.

Single-cell RNA-sequencing (scRNA-seq) provides gene expression profiles of individual cells from complex samples, facilitating the detection of cell type-specific marker genes. In scRNA-seq experiments with multiple donors, the population level variation brings an extra layer of complexity in cell type-specific gene detection, for example, they may not appear in all donors. Motivated by this observation, we develop a statistical model named scCTS to identify cell type-specific genes from population-level scRNA-seq data. Extensive data analyses demonstrate that the proposed method identifies more biologically meaningful cell type-specific genes compared to traditional methods.

The pyroptosis of cardiomyocytes has become an essential topic in heart failure research. The abnormal accumulation of these biological factors, including angiotensin II, advanced glycation end products, and various growth factors (such as connective tissue growth factor, vascular endothelial growth factor, transforming growth factor beta, among others), activates the nuclear factor-κB (NF-κB) signaling pathway in cardiovascular diseases, ultimately leading to pyroptosis of cardiomyocytes. Therefore, exploring the underlying molecular biological mechanisms is essential for developing novel drugs and therapeutic strategies. However, our current understanding of the precise regulatory mechanism of this complex signaling pathway in cardiomyocyte pyroptosis is still limited. Given this, this study reviews the milestone discoveries in the field of pyroptosis research since 1986, analyzes in detail the similarities, differences, and interactions between pyroptosis and other cell death modes (such as apoptosis, necroptosis, autophagy, and ferroptosis), and explores the deep connection between pyroptosis and heart failure. At the same time, it depicts in detail the complete pathway of the activation, transmission, and eventual cardiomyocyte pyroptosis of the NF-κB signaling pathway in the process of heart failure. In addition, the study also systematically summarizes various therapeutic approaches that can inhibit NF-κB to reduce cardiomyocyte pyroptosis, including drugs, natural compounds, small molecule inhibitors, gene editing, and other cutting-edge technologies, aiming to provide solid scientific support and new research perspectives for the prevention and treatment of heart failure.

A major fraction of loci identified by genome-wide association studies (GWASs) mediate alternative splicing, but mechanistic interpretation is hindered by the technical limitations of short-read RNA sequencing (RNA-seq), which cannot directly link splicing events to full-length protein isoforms. Long-read RNA-seq represents a powerful tool to characterize transcript isoforms, and recently, infer protein isoform existence. Here, we present an approach that integrates information from GWASs, splicing quantitative trait loci (sQTLs), and PacBio long-read RNA-seq in a disease-relevant model to infer the effects of sQTLs on the ultimate protein isoform products they encode. We demonstrate the utility of our approach using bone mineral density (BMD) GWAS data. We identified 1,863 sQTLs from the Genotype-Tissue Expression (GTEx) project in 732 protein-coding genes that colocalized with BMD associations (H4PP ≥ 0.75). We generated PacBio Iso-Seq data (N = ∼22 million full-length reads) on human osteoblasts, identifying 68,326 protein-coding isoforms, of which 17,375 (25%) were unannotated. By casting the sQTLs onto protein isoforms, we connected 809 sQTLs to 2,029 protein isoforms from 441 genes expressed in osteoblasts. Overall, we found that 74 sQTLs influenced isoforms likely impacted by nonsense-mediated decay and 190 that potentially resulted in the expression of unannotated protein isoforms. Finally, we functionally validated colocalizing sQTLs in TPM2, in which siRNA-mediated knockdown in osteoblasts showed two TPM2 isoforms with opposing effects on mineralization but exhibited no effect upon knockdown of the entire gene. Our approach should be to generalize across diverse clinical traits and to provide insights into protein isoform activities modulated by GWAS loci.

Retinol binding protein 4 (RBP4) is an adipokine that has been explored as a key biomarker of type 2 diabetes mellitus (T2DM) in recent years. Researchers have conducted a series of experiments to understand the interplay between RBP4 and T2DM, including its role in insulin resistance and pancreatic β-cell function. The results of these studies indicate that RBP4 has a significant influence on T2DM and is considered a potential biomarker of T2DM. However, there have also been some controversies about the relationship between RBP4 levels and T2DM. In this review, we update and summarize recent studies focused on the relationship between RBP4 and T2DM and its role in insulin resistance and pancreatic β-cell function to clarify the existing controversy and provide evidence for future studies. We also assessed the potential therapeutic applications of RBP4 in treating T2DM.

A narrative review.

Overall, there were significant associations between RBP4 levels, insulin resistance, pancreatic β-cell function, and T2DM.

More mechanistic studies are needed to determine the role of RBP4 in the onset of T2DM, especially in terms of pancreatic β-cell function. In addition, further studies are required to evaluate the effects of drug intervention, lifestyle intervention, and bariatric surgery on RBP4 levels to control T2DM and the role of reducing RBP4 levels in improving insulin sensitivity and pancreatic β-cell function.

Nectin-4 (Nectin cell adhesion molecule 4), a type I transmembrane cell adhesion protein, was demonstrated to be overexpressed in a variety of tumors, making it an attractive antigen for targeted therapies such as antibody-drug conjugates (ADCs). Of great note, the US Food and Drug Administration (FDA)-approval of the first Nectin-4-directed ADC, enfortumab vedotin (EV), in urothelial cancer (UC) not only introduced Nectin-4 as a clinically validated and reliable target antigen but also confirmed the evolving role of Nectin-4-directed ADCs as novel and promising cancer therapeutics. In addition to EV, there have been or are currently being seven and eleven Nectin-4-directed ADCs, respectively, in various stages of clinical trials and preclinical development, offering a promising future for the treatment of Nectin-4-positive cancer patients. This study reviewed clinical- and preclinical-stage Nectin-4-directed ADCs.

KIF2C/MCAK (KIF2C) is the most well-characterized member of the kinesin-13 family, which is critical in the regulation of microtubule (MT) dynamics during mitosis, as well as interphase. This systematic review briefly describes the important structural elements of KIF2C, its regulation by multiple molecular mechanisms, and its broad cellular functions. Furthermore, it systematically summarizes its oncogenic potential in malignant progression and performs a meta-analysis of its prognostic value in cancer patients. KIF2C was shown to be involved in multiple crucial cellular processes including cell migration and invasion, DNA repair, senescence induction and immune modulation, which are all known to be critical during the development of malignant tumors. Indeed, an increasing number of publications indicate that KIF2C is aberrantly expressed in multiple cancer entities. Consequently, we have highlighted its involvement in at least five hallmarks of cancer, namely: genome instability, resisting cell death, activating invasion and metastasis, avoiding immune destruction and cellular senescence. This was followed by a systematic search of KIF2C/MCAK's expression in various malignant tumor entities and its correlation with clinicopathologic features. Available data were pooled into multiple weighted meta-analyses for the correlation between KIF2Chigh protein or gene expression and the overall survival in breast cancer, non-small cell lung cancer and hepatocellular carcinoma patients. Furthermore, high expression of KIF2C was correlated to disease-free survival of hepatocellular carcinoma. All meta-analyses showed poor prognosis for cancer patients with KIF2Chigh expression, associated with a decreased overall survival and reduced disease-free survival, indicating KIF2C's oncogenic potential in malignant progression and as a prognostic marker. This work delineated the promising research perspective of KIF2C with modern in vivo and in vitro technologies to further decipher the function of KIF2C in malignant tumor development and progression. This might help to establish KIF2C as a biomarker for the diagnosis or evaluation of at least three cancer entities.

Development of the anterior segment of the eye requires reciprocal sequential interactions between the arising tissues, facilitated by numerous genetic factors. Disruption of any of these processes results in congenital anomalies in the affected tissue(s) leading to anterior segment disorders (ASD) including aniridia, Axenfeld-Rieger anomaly, congenital corneal opacities (Peters anomaly, cornea plana, congenital primary aphakia), and primary congenital glaucoma. Current understanding of the genetic factors involved in ASD remains incomplete, with approximately 50% overall receiving a genetic diagnosis. While some genes are strongly associated with a specific clinical diagnosis, the majority of known factors are linked with highly variable phenotypic presentations, with pathogenic variants in FOXC1, CYP1B1, and PITX2 associated with the broadest spectrum of ASD conditions. This review discusses typical clinical presentations including associated systemic features of various forms of ASD; the latest functional data and genotype-phenotype correlations related to 25 ASD factors including newly identified genes; promising novel candidates; and current and emerging treatments for these complex conditions. Recent developments of interest in the genetics of ASD include identification of phenotypic expansions for several factors, discovery of multiple modes of inheritance for some genes, and novel mechanisms including a growing number of non-coding variants and alleles affecting specific domains/residues and requiring further studies.

The otopetrin (OTOP) proteins were recently characterized as extracellular proton-activated proton channels. Several recent OTOP channel structures demonstrated that the channels form a dimer with each subunit adopting a double-barrel architecture. However, the structural mechanisms underlying some basic functional properties of the OTOP channels remain unresolved, including extracellular pH activation, proton conducting pathway, and rapid desensitization. In this study, we performed structural and functional characterization of the Caenorhabditis elegans OTOP8 (CeOTOP8) and mouse OTOP2 (mOTOP2) and illuminated a set of conformational changes related to the proton-conducting process in OTOP. The structures of CeOTOP8 reveal the conformational change at the N-terminal part of TM12 that renders the channel in a transiently proton-transferring state, elucidating an inter-barrel, Glu/His-bridged proton passage within each subunit. The structures of mOTOP2 reveal the conformational change at the N-terminal part of TM6 that exposes the central glutamate to the extracellular solution for protonation. In addition, the structural comparison between CeOTOP8 and mOTOP2, along with the structure-based mutagenesis, demonstrates that an inter-subunit movement at the OTOP channel dimer interface plays a central role in regulating channel activity. Combining the structural information from both channels, we propose a working model describing the multi-step conformational changes during the proton conducting process.

Postoperative pain affects most patients after major surgery and can transition to chronic pain. The considerable side effects and limited efficacy of current treatments underline the need for new therapeutic options. We observed increased amounts of the metabolites BH4 and serotonin after skin injury. Mast cells were primary postoperative sources of Gch1, the rate-limiting enzyme in BH4 synthesis, itself an obligate cofactor in serotonin production by tryptophan hydroxylase (Tph1). Mice deficient in mast cells or in mast cell-specific Gch1 or Tph1 showed drastically decreased postoperative pain. We found that injury induced the nociceptive neuropeptide substance P, mast cell degranulation, and granule nerve colocalization. Substance P triggered serotonin release in mouse and human mast cells, and substance P receptor blockade substantially ameliorated pain hypersensitivity. Our findings highlight the importance of mast cells at the neuroimmune interface and substance P-driven mast cell BH4 and serotonin production as a therapeutic target for postoperative pain treatment.

N-lactoyl-phenylalanine (Lac-Phe) is a lactate-derived metabolite that suppresses food intake and body weight. Little is known about the mechanisms that mediate Lac-Phe transport across cell membranes. Here we identify SLC17A1 and SLC17A3, two kidney-restricted plasma membrane-localized solute carriers, as physiologic urine Lac-Phe transporters. In cell culture, SLC17A1/3 exhibit high Lac-Phe efflux activity. In humans, levels of Lac-Phe in urine exhibit a strong genetic association with the SLC17A1-4 locus. Urine Lac-Phe levels are increased following a Wingate sprint test. In mice, genetic ablation of either SLC17A1 or SLC17A3 reduces urine Lac-Phe levels. Despite these differences, both knockout strains have normal blood Lac-Phe and body weights, demonstrating SLC17A1/3-dependent de-coupling of urine and plasma Lac-Phe pools. Together, these data establish SLC17A1/3 family members as the physiologic urine Lac-Phe transporters and uncover a biochemical pathway for the renal excretion of this signaling metabolite.

Rabies, caused by lyssavirus rabies (Rabies lyssavirus, RABV), is a fatal disease among humans and almost all warm-blooded animals. In this study, we found that RABV infection induces the up-regulation of receptor transporter protein 4 (RTP4) in mouse brains and different cells of nervous tissue. Over-expression of RTP4 reduces the viral titer of RABV in different neuronal cells. Furthermore, a recombinant RABV expressing RTP4, named rRABV-RTP4, was constructed and displayed a lower viral titer in different neuronal cells due to the expression of RTP4. Moreover, the survival rates of mice infected with rRABV-RTP4 were significantly higher than those of mice infected with parent virus rRABV or control virus rRABV-RTP4(-). In terms of mechanism, RTP4 could bind viral genomic RNA (vRNA) of RABV, and suppress the whole viral genome amplification. In addition, we found that the zinc finger domain (ZFD) of RTP4 exerts the antiviral function by truncation analysis, and an important amino acids site (C95) in the RTP4 3CxxC motif which is essential for its antiviral function was identified by mutation analysis. This study contributes to our understanding of how RTP4 or other RTP proteins play a role in defense against the invasion of RABV or other viruses.

The inhibitors, CK-666 and CK-869, are widely used to probe the function of Arp2/3 complex mediated actin nucleation in vitro and in cells. However, in mammals, the Arp2/3 complex consists of 8 iso-complexes, as three of its subunits (Arp3, ArpC1, ArpC5) are encoded by two different genes. Here, we used recombinant Arp2/3 with defined composition to assess the activity of CK-666 and CK-869 against iso-complexes. We demonstrate that both inhibitors prevent linear actin filament formation when ArpC1A- or ArpC1B-containing complexes are activated by SPIN90. In contrast, inhibition of actin branching depends on iso-complex composition. Both drugs prevent actin branch formation by complexes containing ArpC1A, but only CK-869 can inhibit ArpC1B-containing complexes. Consistent with this, in bone marrow-derived macrophages which express low levels of ArpC1A, CK-869 but not CK-666, impacted phagocytosis and cell migration. CK-869 also only inhibits Arp3- but not Arp3B-containing iso-complexes. Our findings have important implications for the interpretation of results using CK-666 and CK-869, given that the relative expression levels of ArpC1 and Arp3 isoforms in cells and tissues remains largely unknown.

Inflammatory bowel disease (IBD) is a chronic disease of the gastrointestinal tract affecting millions of people. Here, we investigated the expression and functions of poly(ADP-ribose) polymerase 14 (Parp14), an important regulatory protein in immune cells, with an IBD patient cohort as well as two mouse colitis models, that is, IBD-mimicking oral dextran sulfate sodium (DSS) exposure and oral Salmonella infection. Parp14 was expressed in the human colon by cells in the lamina propria, but, in particular, by the epithelial cells with a granular staining pattern in the cytosol. The same expression pattern was evidenced in both mouse models. Parp14-deficiency caused increased rectal bleeding as well as stronger epithelial erosion, Goblet cell loss, and immune cell infiltration in DSS-exposed mice. The absence of Parp14 did not affect the mouse colon bacterial microbiota. Also, the colon leukocyte populations of Parp14-deficient mice were normal. In contrast, bulk tissue RNA-Seq demonstrated that the colon transcriptomes of Parp14-deficient mice were dominated by abnormalities in inflammation and infection responses both prior and after the DSS exposure. Overall, the data indicate that Parp14 has an important role in the maintenance of colon epithelial barrier integrity. The prognostic and predictive biomarker potential of Parp14 in IBD merits further investigation.

Diabetic kidney disease (DKD) is the primary cause of end-stage renal disease. The Astragalus-Coptis drug pair is frequently employed in the management of DKD. However, the precise molecular mechanism underlying its therapeutic effect remains elusive.

To investigate the synergistic effects of multiple active ingredients in the Astragalus-Coptis drug pair on DKD through multiple targets and pathways.

The ingredients of the Astragalus-Coptis drug pair were collected and screened using the TCMSP database and the SwissADME platform. The targets were predicted using the SwissTargetPrediction database, while the DKD differential gene expression analysis was obtained from the Gene Expression Omnibus database. DKD targets were acquired from the GeneCards, Online Mendelian Inheritance in Man database, and DisGeNET databases, with common targets identified through the Venny platform. The protein-protein interaction network and the "disease-active ingredient-target" network of the common targets were constructed utilizing the STRING database and Cytoscape software, followed by the analysis of the interaction relationships and further screening of key targets and core active ingredients. Gene Ontology (GO) function and Kyoto Ency-clopedia of Genes and Genomes (KEGG) pathway enrichments were performed using the DAVID database. The tissue and organ distributions of key targets were evaluated. PyMOL and AutoDock software validate the molecular docking between the core ingredients and key targets. Finally, molecular dynamics (MD) simulations were conducted to simulate the optimal complex formed by interactions between core ingredients and key target proteins.

A total of 27 active ingredients and 512 potential targets of the Astragalus-Coptis drug pair were identified. There were 273 common targets between DKD and the Astragalus-Coptis drug pair. Through protein-protein interaction network topology analysis, we identified 9 core active ingredients and 10 key targets. GO and KEGG pathway enrichment analyses revealed that Astragalus-Coptis drug pair treatment for DKD involves various biological processes, including protein phosphorylation, negative regulation of apoptosis, inflammatory response, and endoplasmic reticulum unfolded protein response. These pathways are mainly associated with the advanced glycation end products (AGE)-receptor for AGE products signaling pathway in diabetic complications, as well as the Lipid and atherosclerosis. Molecular docking and MD simulations demonstrated high affinity and stability between the core active ingredients and key targets. Notably, the quercetin-AKT serine/threonine kinase 1 (AKT1) and quercetin-tumor necrosis factor (TNF) protein complexes exhibited exceptional stability.

This study demonstrated that DKD treatment with the Astragalus-Coptis drug pair involves multiple ingredients, targets, and signaling pathways. We propose a novel approach for investigating the molecular mechanism underlying the therapeutic effects of the Astragalus-Coptis drug pair on DKD. Furthermore, we suggest that quercetin is the most potent active ingredient and specifically targets AKT1 and TNF, providing a theoretical foundation for further exploration of pharmacologically active ingredients and elucidating their molecular mechanisms in DKD treatment.

Metabolic syndrome (MetS) represents a constellation of metabolic abnormalities, typified by obesity, hypertension, hyperglycemia, and hyperlipidemia. It stems from intricate dysregulations in metabolic pathways governing energy and substrate metabolism. While comprehending the precise etiological mechanisms of MetS remains challenging, evidence underscores the pivotal roles of aberrations in lipid metabolism and insulin resistance (IR) in its pathogenesis. Notably, nicotinamide N-methyltransferase (NNMT) has recently surfaced as a promising therapeutic target for addressing MetS. Single nucleotide variants in the NNMT gene are significantly correlated with disturbances in energy metabolism, obesity, type 2 diabetes (T2D), hyperlipidemia, and hypertension. Elevated NNMT gene expression is notably observed in the liver and white adipose tissue (WAT) of individuals with diabetic mice, obesity, and rats afflicted with MetS. Knockdown of NNMT elicits heightened energy expenditure in adipose and hepatic tissues, mitigates lipid accumulation, and enhances insulin sensitivity. NNMT catalyzes the methylation of nicotinamide (NAM) using S-adenosyl-methionine (SAM) as the donor methyl group, resulting in the formation of S-adenosyl-l-homocysteine (SAH) and methylnicotinamide (MNAM). This enzymatic process results in the depletion of NAM, a precursor of nicotinamide adenine dinucleotide (NAD+), and the generation of SAH, a precursor of homocysteine (Hcy). Consequently, this cascade leads to reduced NAD+ levels and elevated Hcy levels, implicating NNMT in the pathogenesis of MetS. Moreover, experimental studies employing RNA interference (RNAi) strategies and small molecule inhibitors targeting NNMT have underscored its potential as a therapeutic target for preventing or treating MetS-related diseases. Nonetheless, the precise mechanistic underpinnings remain elusive, and as of yet, clinical trials focusing on NNMT have not been documented. Therefore, further investigations are warranted to elucidate the intricate roles of NNMT in MetS and to develop targeted therapeutic interventions.

Pathologic α-synuclein (α-syn) spreads from cell-to-cell, in part, through binding to the lymphocyte-activation gene 3 (Lag3). Here we report that amyloid β precursor-like protein 1 (Aplp1) interacts with Lag3 that facilitates the binding, internalization, transmission, and toxicity of pathologic α-syn. Deletion of both Aplp1 and Lag3 eliminates the loss of dopaminergic neurons and the accompanying behavioral deficits induced by α-syn preformed fibrils (PFF). Anti-Lag3 prevents the internalization of α-syn PFF by disrupting the interaction of Aplp1 and Lag3, and blocks the neurodegeneration induced by α-syn PFF in vivo. The identification of Aplp1 and the interplay with Lag3 for α-syn PFF induced pathology deepens our insight about molecular mechanisms of cell-to-cell transmission of pathologic α-syn and provides additional targets for therapeutic strategies aimed at preventing neurodegeneration in Parkinson's disease and related α-synucleinopathies.

The suprachiasmatic nucleus (SCN) encodes time of day through changes in daily firing; however, the molecular mechanisms by which the SCN times behavior are not fully understood. To identify factors that could encode day/night differences in activity, we combine patch-clamp recordings and single-cell sequencing of individual SCN neurons in mice. We identify PiT2, a phosphate transporter, as being upregulated in a population of Vip+Nms+ SCN neurons at night. Although nocturnal and typically showing a peak of activity at lights off, mice lacking PiT2 (PiT2-/-) do not reach the activity level seen in wild-type mice during the light/dark transition. PiT2 loss leads to increased SCN neuronal firing and broad changes in SCN protein phosphorylation. PiT2-/- mice display a deficit in seasonal entrainment when moving from a simulated short summer to longer winter nights. This suggests that PiT2 is responsible for timing activity and is a driver of SCN plasticity allowing seasonal entrainment.