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Nature Communications Feb 2024Circulating monocytes participate in pain chronification but the molecular events that cause their deployment are unclear. Using a mouse model of hyperalgesic priming...
Circulating monocytes participate in pain chronification but the molecular events that cause their deployment are unclear. Using a mouse model of hyperalgesic priming (HP), we show that monocytes enable progression to pain chronicity through a mechanism that requires transient activation of the hydrolase, N-acylethanolamine acid amidase (NAAA), and the consequent suppression of NAAA-regulated lipid signaling at peroxisome proliferator-activated receptor-α (PPAR-α). Inhibiting NAAA in the 72 hours following administration of a priming stimulus prevented HP. This effect was phenocopied by NAAA deletion and depended on PPAR-α recruitment. Mice lacking NAAA in CD11b cells - monocytes, macrophages, and neutrophils - were resistant to HP induction. Conversely, mice overexpressing NAAA or lacking PPAR-α in the same cells were constitutively primed. Depletion of monocytes, but not resident macrophages, generated mice that were refractory to HP. The results identify NAAA-regulated signaling in monocytes as a control node in the induction of HP and, potentially, the transition to pain chronicity.
Topics: Humans; Amidohydrolases; Enzyme Inhibitors; Hyperalgesia; Lipids; Monocytes; Pain; PPAR alpha; Animals; Mice
PubMed: 38402219
DOI: 10.1038/s41467-024-46139-5 -
Nature Communications Feb 2024Fusion-positive rhabdomyosarcoma (FP-RMS) is an aggressive pediatric sarcoma driven primarily by the PAX3-FOXO1 fusion oncogene, for which therapies targeting PAX3-FOXO1...
Fusion-positive rhabdomyosarcoma (FP-RMS) is an aggressive pediatric sarcoma driven primarily by the PAX3-FOXO1 fusion oncogene, for which therapies targeting PAX3-FOXO1 are lacking. Here, we screen 62,643 compounds using an engineered cell line that monitors PAX3-FOXO1 transcriptional activity identifying a hitherto uncharacterized compound, P3FI-63. RNA-seq, ATAC-seq, and docking analyses implicate histone lysine demethylases (KDMs) as its targets. Enzymatic assays confirm the inhibition of multiple KDMs with the highest selectivity for KDM3B. Structural similarity search of P3FI-63 identifies P3FI-90 with improved solubility and potency. Biophysical binding of P3FI-90 to KDM3B is demonstrated using NMR and SPR. P3FI-90 suppresses the growth of FP-RMS in vitro and in vivo through downregulating PAX3-FOXO1 activity, and combined knockdown of KDM3B and KDM1A phenocopies P3FI-90 effects. Thus, we report KDM inhibitors P3FI-63 and P3FI-90 with the highest specificity for KDM3B. Their potent suppression of PAX3-FOXO1 activity indicates a possible therapeutic approach for FP-RMS and other transcriptionally addicted cancers.
Topics: Child; Humans; Paired Box Transcription Factors; Rhabdomyosarcoma, Alveolar; Cell Line, Tumor; Rhabdomyosarcoma; Forkhead Box Protein O1; Oncogene Proteins, Fusion; Gene Expression Regulation, Neoplastic; PAX3 Transcription Factor; Jumonji Domain-Containing Histone Demethylases; Histone Demethylases
PubMed: 38402212
DOI: 10.1038/s41467-024-45902-y -
International Journal of Molecular... Feb 2024The circadian rhythm is necessary for the homeostasis and health of living organisms. Molecular clocks interconnected by transcription/translation feedback loops exist... (Review)
Review
The circadian rhythm is necessary for the homeostasis and health of living organisms. Molecular clocks interconnected by transcription/translation feedback loops exist in most cells of the body. A puzzling exemption to this, otherwise, general biological hallmark is given by the cell physiology of pluripotent stem cells (PSCs) that lack circadian oscillations gradually acquired following their in vivo programmed differentiation. This process can be nicely phenocopied following in vitro commitment and reversed during the reprogramming of somatic cells to induce PSCs. The current understanding of how and why pluripotency is "time-uncoupled" is largely incomplete. A complex picture is emerging where the circadian core clockwork is negatively regulated in PSCs at the post-transcriptional/translational, epigenetic, and other-clock-interaction levels. Moreover, non-canonical functions of circadian core-work components in the balance between pluripotency identity and metabolic-driven cell reprogramming are emerging. This review selects and discusses results of relevant recent investigations providing major insights into this context.
Topics: Circadian Rhythm; Cell Differentiation; Pluripotent Stem Cells; Cellular Reprogramming; Circadian Clocks
PubMed: 38396740
DOI: 10.3390/ijms25042063 -
Cells Feb 2024Platelet function at vascular injury sites is tightly regulated through the actin cytoskeleton. The Wiskott-Aldrich syndrome protein-family verprolin-homologous protein...
Platelet function at vascular injury sites is tightly regulated through the actin cytoskeleton. The Wiskott-Aldrich syndrome protein-family verprolin-homologous protein (WAVE)-regulatory complex (WRC) activates lamellipodia formation via ARP2/3, initiated by GTP-bound RAC1 interacting with the WRC subunit CYFIP1. The protein FAM49b (Family of Unknown Function 49b), also known as CYRI-B (CYFIP-Related RAC Interactor B), has been found to interact with activated RAC1, leading to the negative regulation of the WRC in mammalian cells. To investigate the role of FAM49b in platelet function, we studied platelet-specific -, -, and /-mice. Platelet counts and activation of mice were comparable to those of control mice. On fully fibrinogen-coated surfaces, -platelets spread faster with an increased mean projected cell area than control platelets, whereas /-platelets did not form lamellipodia, phenocopying the -platelets. However, -platelets often assumed a polarized shape and were more prone to migrate on fibrinogen-coated surfaces. On 2D structured micropatterns, however, -platelets displayed reduced spreading, whereas spreading of - and /-platelets was enhanced. In summary, FAM49b contributes to the regulation of morphology and migration of spread platelets, but to exert its inhibitory effect on actin polymerization, the functional WAVE complex must be present.
Topics: Animals; Mice; Actin Cytoskeleton; Adaptor Proteins, Signal Transducing; Blood Platelets; Carrier Proteins; Fibrinogen; Mammals; rac1 GTP-Binding Protein
PubMed: 38391912
DOI: 10.3390/cells13040299 -
Arteriosclerosis, Thrombosis, and... Apr 2024Venous thromboembolism is a major health problem. After thrombus formation, its resolution is essential to re-establish blood flow, which is crucially mediated by...
BACKGROUND
Venous thromboembolism is a major health problem. After thrombus formation, its resolution is essential to re-establish blood flow, which is crucially mediated by infiltrating neutrophils and monocytes in concert with activated platelets and endothelial cells. Thus, we aimed to modulate leukocyte function during thrombus resolution post-thrombus formation by blocking P-selectin/CD62P-mediated cell interactions.
METHODS
Thrombosis was induced by inferior vena cava stenosis through ligation in mice. After 1 day, a P-selectin-blocking antibody or isotype control was administered and thrombus composition and resolution were analyzed.
RESULTS
Localizing neutrophils and macrophages in thrombotic lesions of wild-type mice revealed that these cells enter the thrombus and vessel wall from the caudal end. Neutrophils were predominantly present 1 day and monocytes/macrophages 3 days after vessel ligation. Blocking P-selectin reduced circulating platelet-neutrophil and platelet-Ly6C monocyte aggregates near the thrombus, and diminished neutrophils and Ly6C macrophages in the cranial thrombus part compared with isotype-treated controls. Depletion of neutrophils 1 day after thrombus initiation did not phenocopy P-selectin inhibition but led to larger thrombi compared with untreated controls. In vitro, P-selectin enhanced human leukocyte function as P-selectin-coated beads increased reactive oxygen species production by neutrophils and tissue factor expression of classical monocytes. Accordingly, P-selectin inhibition reduced oxidative burst in the thrombus and tissue factor expression in the adjacent vessel wall. Moreover, blocking P-selectin reduced thrombus density determined by scanning electron microscopy and increased urokinase-type plasminogen activator levels in the thrombus, which accelerated caudal fibrin degradation from day 3 to day 14. This accelerated thrombus resolution as thrombus volume declined more rapidly after blocking P-selectin.
CONCLUSIONS
Inhibition of P-selectin-dependent activation of monocytes and neutrophils accelerates venous thrombosis resolution due to reduced infiltration and activation of innate immune cells at the site of thrombus formation, which prevents early thrombus stabilization and facilitates fibrinolysis.
Topics: Mice; Humans; Animals; Monocytes; P-Selectin; Endothelial Cells; Thromboplastin; Neutrophil Infiltration; Thrombosis; Neutrophils
PubMed: 38385292
DOI: 10.1161/ATVBAHA.123.320016 -
JACC. Case Reports Feb 2024Noninvasive imaging is crucial for diagnosing and managing arrhythmogenic cardiomyopathy. Despite advanced multimodality imaging tools, challenges persist in...
Noninvasive imaging is crucial for diagnosing and managing arrhythmogenic cardiomyopathy. Despite advanced multimodality imaging tools, challenges persist in differentiating it from other arrhythmogenic diseases (eg, cardiac sarcoidosis). We present a case of arrhythmogenic cardiomyopathy with an variant of uncertain significance exhibiting a phenocopy of cardiac sarcoidosis.
PubMed: 38379651
DOI: 10.1016/j.jaccas.2023.102198 -
MSphere Mar 2024Fungal infections cause a large health burden but are treated by only a handful of antifungal drug classes. Chromatin factors have emerged as possible targets for new...
Fungal infections cause a large health burden but are treated by only a handful of antifungal drug classes. Chromatin factors have emerged as possible targets for new antifungals. These targets include the reader proteins, which interact with posttranslationally modified histones to influence DNA transcription and repair. The YEATS domain is one such reader recognizing both crotonylated and acetylated histones. Here, we performed a detailed structure/function analysis of the YEATS domain reader Yaf9, a subunit of the NuA4 histone acetyltransferase and the SWR1 chromatin remodeling complex. We have previously demonstrated that the homozygous deletion mutant displays growth defects and is avirulent in mice. Here we show that a YEATS domain mutant expected to inactivate Yaf9's chromatin binding does not display strong phenotypes , nor during infection of immune cells or in a mouse systemic infection model, with only a minor virulence reduction . In contrast to the YEATS domain mutation, deletion of the C-terminal domain of Yaf9, a protein-protein interaction module necessary for its interactions with SWR1 and NuA4, phenocopies the null mutant. This shows that the C-terminal domain is essential for Yaf9 roles and , including virulence. Our study informs on the strategies for therapeutic targeting of Yaf9, showing that approaches taken for the mammalian YEATS domains by disrupting their chromatin binding might not be effective in , and provides a foundation for studying YEATS proteins in human fungal pathogens.IMPORTANCEThe scarcity of available antifungal drugs and rising resistance demand the development of therapies with new modes of action. In this context, chromatin regulation may be a target for novel antifungal therapeutics. To realize this potential, we must better understand the roles of chromatin regulators in fungal pathogens. Toward this goal, here, we studied the YEATS domain chromatin reader Yaf9 in . Yaf9 uses the YEATS domain for chromatin binding and a C-terminal domain to interact with chromatin remodeling complexes. By constructing mutants in these domains and characterizing their phenotypes, our data indicate that the Yaf9 YEATS domain might not be a suitable therapeutic drug target. Instead, the Yaf9 C-terminal domain is critical for virulence. Collectively, our study informs how a class of chromatin regulators performs their cellular and pathogenesis roles in and reveals strategies to inhibit them.
Topics: Humans; Animals; Mice; Chromatin; Histones; Candida albicans; Saccharomyces cerevisiae Proteins; Antifungal Agents; Homozygote; Sequence Deletion; Transcription Factors; Protein Interaction Domains and Motifs; Mammals
PubMed: 38376217
DOI: 10.1128/msphere.00696-23 -
BioRxiv : the Preprint Server For... Feb 2024The role of splicing dysregulation in cancer is underscored by splicing factor mutations; however, its impact in the absence of such rare mutations is poorly understood....
The role of splicing dysregulation in cancer is underscored by splicing factor mutations; however, its impact in the absence of such rare mutations is poorly understood. To reveal complex patient subtypes and putative regulators of pathogenic splicing in Acute Myeloid Leukemia (AML), we developed a new approach called OncoSplice. Among diverse new subtypes, OncoSplice identified a biphasic poor prognosis signature that partially phenocopies -mutant splicing, impacting thousands of genes in over 40% of adult and pediatric AML cases. -like splicing co-opted a healthy circadian splicing program, was stable over time and induced a leukemia stem cell (LSC) program. Pharmacological inhibition of the implicated -like splicing regulator, PRMT5, rescued leukemia mis-splicing and inhibited leukemic cell growth. Genetic deletion of IRAK4, a common target of -like and PRMT5 treated cells, blocked leukemia development in xenograft models and induced differentiation. These analyses reveal a new prognostic alternative-splicing mechanism in malignancy, independent of splicing-factor mutations.
PubMed: 38370617
DOI: 10.1101/2024.02.04.578798 -
Heliyon Feb 2024Sushi, von Willebrand factor type A, EGF and pentraxin domain containing 1 (SVEP1) is an extracellular matrix protein that causally promotes cardiovascular disease in...
Sushi, von Willebrand factor type A, EGF and pentraxin domain containing 1 (SVEP1) is an extracellular matrix protein that causally promotes cardiovascular disease in humans and mice. However, the receptor mediating the effect of SVEP1 on the development of disease remains unclear. We previously demonstrated that depleting either vascular smooth muscle cell (VSMC)- or myeloid cell-derived integrin α9β1, the first receptor that was identified to interact with SVEP1, did not phenocopy the disease-abrogating effect of depleting SVEP1. Due to its wide expression in tissues and cell types, here we extend this line of investigation to definitively determine if integrin α9β1 impacts the development of atherosclerosis. In a mouse model of atherosclerosis, we found that depleting integrin α9β1 in all cells did not alter plaque size or characteristics of plaque complexity when compared to wild type mice. Further, the significant SVEP1-mediated effects on increase in macrophage content and VSMC proliferation within the atherosclerotic plaque were not altered in animals lacking integrin α9β1. Together, these findings strongly suggest that integrin α9β1 is not responsible for mediating the SVEP1-induced promotion of atherosclerosis and support further studies aimed at characterizing other receptors whose interaction with SVEP1 may represent a therapeutically targetable interaction.
PubMed: 38370227
DOI: 10.1016/j.heliyon.2024.e25760 -
Development (Cambridge, England) Feb 2024Malrotation of the intestine is a prevalent birth anomaly, the etiology of which remains poorly understood. Here, we show that late-stage exposure of Xenopus embryos to...
Malrotation of the intestine is a prevalent birth anomaly, the etiology of which remains poorly understood. Here, we show that late-stage exposure of Xenopus embryos to atrazine, a widely used herbicide that targets electron transport chain (ETC) reactions, elicits intestinal malrotation at high frequency. Interestingly, atrazine specifically inhibits the cellular morphogenetic events required for gut tube elongation, including cell rearrangement, differentiation and proliferation; insufficient gut lengthening consequently reorients the direction of intestine rotation. Transcriptome analyses of atrazine-exposed intestines reveal misexpression of genes associated with glycolysis and oxidative stress, and metabolomics shows that atrazine depletes key glycolytic and tricarboxylic acid cycle metabolites. Moreover, cellular bioenergetics assays indicate that atrazine blocks a crucial developmental transition from glycolytic ATP production toward oxidative phosphorylation. Atrazine-induced defects are phenocopied by rotenone, a known ETC Complex I inhibitor, accompanied by elevated reactive oxygen species, and rescued by antioxidant supplementation, suggesting that malrotation may be at least partly attributable to redox imbalance. These studies reveal roles for metabolism in gut morphogenesis and implicate defective gut tube elongation and/or metabolic perturbations in the etiology of intestinal malrotation.
Topics: Atrazine; Rotation; Herbicides; Oxidation-Reduction; Gene Expression Profiling
PubMed: 38369735
DOI: 10.1242/dev.202020