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Integrative structural interactomics reveals protein organization and structure in a giant virus
Get PDFGiant viruses are large DNA viruses that infect unicellular and multicellular eukaryotes and form exceptionally large extracellular particles. (Meta)genomics and (meta)transcriptomics have provided insight into their diverse coding repertoire, but many of the proteins remain to be characterized as they lack homology with known proteins. Here, we integrated cross-linking mass spectrometry, quantitative proteomics, computational tools and cryo-EM data to characterize the protein architecture of intact melbournevirus particles. Based on this, we allocated 88 viral proteins to different virion sub-compartments and proposed topologies of 25 inner membrane proteins. We assigned eight components of the capsid in cryo-EM data, including proteins that tether the capsid shell to the membrane, reflecting key points in virion maturation. The data provide a valuable resource and demonstrate the power of an integrative approach to gain system-level structural insights into a poorly characterized biological system
Proneural-mesenchymal hybrid glioblastoma cells are resistant to therapy and dependent on nuclear import
Get PDFBACKGROUND: Despite extensive research efforts, glioblastoma (GBM) remains a deadly disease with poor prognosis. Although previous studies have identified various cell states within GBM tumors, the molecular mechanism underlying adaptive GBM cell plasticity induced by conventional therapy remains unclear. METHODS: We used fluorescent reporters for proneural (PN) and mesenchymal (MES) subtypes to monitor GBM cell plasticity in real-time across multiple patient-derived cell lines. This approach revealed cells that concurrently expressed both PN and MES markers. To investigate this unique hybrid population, we implemented a comprehensive methodological approach encompassing bulk and single-cell RNA sequencing, single-cell ChIP sequencing, nuclear proteomics, high-resolution imaging, orthotopic mouse models, clinical dataset analysis, and pharmacological and genetic techniques. This multifaceted strategy allowed us to gain functional and molecular insights into this distinct cellular population. RESULTS: We showed that these hybrid cells are increased by conventional therapies, and are resistant to these therapies. At the molecular level, hybrid cells display significant alterations in chromatin structure and nuclear protein composition, elevated transcriptional activity, Myc activation, and improved transport between the nucleus and cytoplasm. Genetic and pharmaceutical inhibition of the nuclear import/export shuttling machinery, increased in hybrid cells, effectively suppressed adaptive GBM cell plasticity and hybrid identity, thereby enhancing the sensitivity of GBM cells to therapies. CONCLUSIONS: Our results indicate that GBM hybrid cells play a crucial role in chemoradiation resistance. The nuclear transport machinery presents a potential therapeutic target for hybrid cells, offering a way to counteract the typical resistance to treatment observed in GBM
Hepatitis B virus-infected hepatocytes promote the secretion of collagen VI to the extracellular matrix
Get PDFChronic hepatitis B virus (HBV) infection is a global health problem as it is the major cause of liver fibrosis and its complications cirrhosis and hepatocellular carcinoma. The role of virus-host interactions in liver fibrosis and progression to cancer remains poorly understood. Here we show that HBV infection of permissive cells trigger pathways relevant for extracellular matrix (ECM) remodeling, which is a hallmark of liver fibrosis. We demonstrate that collagen VI (ColVI) is secreted from infected cells and induces a profibrotic phenotype in patient-derived myofibroblasts and identified HBV-induced AKT signaling as a driver of ColVI expression in HBV-infected cells. Consistently, ColVI is upregulated in the liver of HBV patients with fibrosis. Our results suggest a role of ColVI as a driver of HBV-associated liver disease and highlight the potential of ColVI as a biomarker candidate and therapeutic target in HBV-infected patients
Understanding the variability of peanut-oral immunotherapy responses by multi-omics profiling of immune cells
Get PDFBACKGROUND: Oral immunotherapy (OIT) induces desensitization in peanut allergy, yet 15%–30% of patients do not respond, and a significant risk of anaphylaxis due to treatment remains. In a placebo-controlled peanut OIT trial, this study identifies molecular drivers of OIT responsiveness through multi-omics profiling in immune cells. METHODS: Immunoglobulins, cytokines, transcriptome, and DNA methylome profiles were analyzed in peanut-stimulated and unstimulated peripheral blood mononuclear cells isolated from peanut-allergic children before and after treatment. Multi-omics profiling focused on OIT responsiveness within the active treatment arm. Additional subgroup analyses were performed to further elucidate molecular mechanisms and potential biomarkers. RESULTS: Complete responders, tolerating 4500 mg of peanut protein, exhibited lower pre-treatment peanut-specific IgE and Th2 cytokine production (IL-4, IL-5) compared to incomplete responders who tolerated ≤ 1000 mg of peanut protein after treatment. Our primary analysis identified 184 differentially expressed genes and 1001 differentially methylated genes, enriched for innate (ILC3) and adaptive (CD8αα subset of CD8+ T cells) immune cells, alongside γδ T cells and exosomes, highlighting gastrointestinal regulatory processes as central to OIT success. We found a marked downregulation of immunoglobulin genes in patients receiving peanut compared to placebo, suggesting OIT-induced modulation of B-cell activity. Functional networks revealed a marked imbalance contrasting regulatory T-cell responses and B-cell suppression in the complete responders with innate immune signaling and metabolic stress in the incomplete responders. CONCLUSION: This multi-omics approach underscores the importance of gastrointestinal immune mechanisms underlying the variation in peanut oral immunotherapy responses and offers potential biomarkers for improving treatment strategies
Attenuation of Ca(2+) signaling by overexpression of PMCA2 affects the microglial response to pathological events
No full textMicroglia strongly impact the pathologic course of brain diseases and injuries. Intracellular Ca(2+)dynamics serve as central integrators, connecting microglial sensing capacity to their responses. We generated a mouse line with microglial overexpression of plasma membrane Ca(2+)-ATPase (PMCA)2, a central regulator of cytoplasmic Ca(2+) homeostasis. This manipulation significantly attenuated ATP-evoked Ca(2+)signals in vitro and spontaneous Ca(2+) transients in vivo. Notably, in contrast to astrocytes, PMCA2 overexpression in microglia/macrophages did not affect animal behavior and survival. It had, however, a profound impact on microglial reactivity in pathological contexts, including reduced inflammatory responses following lipopolysaccharide challenge and diminished microglial proliferation at sites of acute injury. In an Alzheimer’s disease model, PMCA2 overexpression attenuated the disease-associated microglial signature, reducing amyloid plaque burden and plaque-associated neuritic dystrophy. These findings highlight the importance of Ca(2+)-mediated signaling for modulating the microglial response to pathologic events. Attenuating microglial Ca(2+)signaling by PMCA2 overexpression is a potential strategy to promote beneficial microglial phenotypes in brain inflammation or degeneration
Pharmacological inhibitors of the gamma-secretase enzyme complex disrupt epithelial cell function triggering colitis in mice
No full textBACKGROUND AND AIMS: Inhibiting γ-secretase-mediated Notch signaling has been explored as a potential treatment for Alzheimer's disease and cancer. However, clinical trials have revealed that this approach can lead to side effects, such as gut inflammation. Notch signaling has been shown to be a key mediator of intestinal epithelial homeostasis. We aimed to investigate the molecular mechanisms of γ-secretase inhibition-associated colitis. METHODS: Mice and small intestinal organoids were treated with γ-secretase inhibitors and analyzed for intestinal epithelial cell (IEC) differentiation and inflammation-associated markers using different molecular and histological approaches, along with transcriptomic and proteomic analyses. To evaluate the role of the microbiome in colitis development, mice undergoing pharmacological γ-secretase inhibition were treated with antibiotics. Additionally, inflammatory bowel disease (IBD) patient samples and control samples were analyzed to assess the expression of Notch signaling pathway components in IECs. RESULTS: This study shows that pharmacological γ-secretase inhibition induces inflammation in both the small and large intestine of mice, a phenotype that could be rescued upon microbiota depletion. Inhibiting the γ-secretase induced structural disruption of the epithelium and inflammatory cytokine release. On a molecular level, epithelial organoids exhibited disrupted IEC differentiation and impaired proliferation, associated with defective Notch signaling. Finally, analysis of IBD patients revealed deregulation of Notch pathway components within IECs. CONCLUSIONS: In conclusion, systemic use of γ-secretase inhibitors disrupts epithelial cell function by impairing IEC differentiation and triggering gut inflammation in mice. These findings should be considered when designing future therapeutic interventions involving γ-secretase inhibitors
Microbiota and kidney disease: the road ahead
No full textMore than 850 million individuals worldwide, accounting for 10–15% of the adult population, are estimated to have chronic kidney disease. Each of these individuals is host to tens of trillions of microorganisms that are collectively referred to as microbiota — a dynamic ecosystem that both influences host health and is itself influenced by changes in the host. Available evidence supports the existence of functional connections between resident microorganisms and kidney health that are altered in the context of specific kidney diseases, including acute kidney injury, chronic kidney disease and renal stone disease. Moreover, promising data from preclinical studies suggest that targeting of gut microbial pathways may provide new therapeutic opportunities for the treatment of kidney disease. This Roadmap describes current understanding of the mechanisms by which microorganisms regulate host organ function, the effects of kidney disease on the gut microbiome, and how these insights may contribute to the development of microbe-targeted therapeutics. We highlight key knowledge gaps that remain to be addressed and strategies for addressing these, outlining both the promise and the potential pitfalls of leveraging our understanding of the gut microbiota to better understand and treat kidney disease
Pressure-adjusted static compression: aerobic metabolism and microvascular perfusion in the context of chemotherapy-induced neuropathy
Get PDFBACKGROUND: Chemotherapy-induced peripheral neuropathy (CIPN) involves impaired microvascular neuronal perfusion and reduced bioenergetics. Compression and cryotherapy are potential preventive measures, yet their mechanism of acral temperature reduction remains unclear. This study aims to unravel the effects of pressure-adjusted static compression (PSC) on aerobic metabolic and endothelial responses in patients undergoing chemotherapy (CTX). METHODS: Cancer patients with CTX above the CIPN-threshold dose (n = 24, 50% male; age 64 [61–71] years) and healthy controls (n = 53, 45% male; age 23 (18 to 87) years) had PSC applied on upper extremities. Tissue oxygenation and metabolism were derived by measuring oxygen supply (O(2)Hb), oxygen demand (HHb), tissue oxygenation (TOI) and microvascular perfusion (THb) with quantitative time-resolved near-infrared spectroscopy (NIRS) and temperature with thermography. Effects were compared to cryoapplication and intermittent pneumatic compression (IPC). Endothelial function was quantified during vascular occlusion test (VOT). RESULTS: PSC, in contrast to undersized surgical gloves (SG), uniformly creates pressure on hands and leads to a more pronounced reduction of hand temperature. Furthermore, PSC significantly increased microvascular perfusion, O(2)-supply and reduced O(2)-demand and aerobic metabolism, thus raised local tissue oxygenation (p < 0.05 each). CTX lead to impaired metabolic and vascular reaction to PSC with only significant reduction of O(2)-demand (p < 0.05) during PSC. PSC is preferable regarding comfort (p < 0.05) compared to SG. Cooling of hands (cooling gloves) had different action (p < 0.001) to PSC with significantly reduced microvascular perfusion, O(2)-supply and O(2)-demand (p < 0.05 each). Comparable local significant effects (p < 0.05) were seen during IPC. CTX exhibited endothelial dysfunction with impaired microvascular reactivity, which limited their capacity to enhance tissue oxygenation. CONCLUSIONS: Reduction of oxygen demand represents an important mechanism in interventions targeting prevention of CIPN. PSC, comparable to cryoapplication and IPC attenuates energy metabolism and enhances tissue oxygenation. PSC’s impact on the combination of vascular and energy metabolism suggests its potential to alleviate CIPN burden. These findings support PSC’s role in reducing acral CIPN through distinct mechanisms
Additional file 10 of ERV3-MLT1 provides cis-regulatory elements for human placental functioning and are commonly dysregulated in human-specific preeclampsia
No full textAdditional file 10: Table S4. Protein encoding isoforms of EPS8L1