Molecular omics最新文献

To illustrate epigenetic heterogeneity, versatile tools of single-cell ChIP-seq (scChIP-seq) are essential for both convenience and accuracy. We developed MobiChIP, a compatible ChIP-seq library construction method based on current sequencing platforms for single-cell applications. MobiChIP efficiently captures fragments from tagmented nuclei across various species and allows sample mixing from different tissues or species. This strategy offers robust nucleosome amplification and flexible sequencing without customized primers. MobiChIP reveals regulatory landscapes of chromatin with active (H3K27ac) and repressive (H3K27me3) histone modification in peripheral blood mononuclear cells (PBMCs) and accurately identifies epigenetic repression of the Hox gene cluster, outperforming ATAC-seq. Meanwhile, we also integrated scChIP-seq with scRNA-seq to further illustrate cellular genetic and epigenetic heterogeneity.

Obstructive sleep apnea (OSA) is characterized by intermittent hypoxic environments at the cellular level and is an independent risk factor for the development of cardiovascular disease. Endothelial cell (EC) dysfunction precedes the development of cardiovascular disease; however, the mechanisms by which ECs respond to these intermittent hypoxic events are poorly understood. To better understand EC responses to hypoxia, we examined the effects of sustained hypoxia (SH) and intermittent hypoxia (IH) on the activation of HIF-1α in ECs. While SH stabilized HIF-1α and led to its nuclear localization, IH did not activate HIF-1α and the expression of its target genes. Using RNA-sequencing, we evaluated transcriptional responses of ECs to hypoxia. SH induced the expression of HIF-1α and hypoxia response genes, while IH affected cell-cycle regulation genes. A cytoscape protein-protein interaction network for EC response to hypoxia was created with differentially expressed genes. The network comprises cell-cycle regulation, inflammatory signaling via NF-κB and response to VEGF stimulus subnetworks on which SH and IH had distinct activities. As OSA is associated with elevated catecholamines, we investigated the effect of epinephrine on the EC response to SH and IH. Transcriptomic responses under IH and epinephrine revealed protein-protein interaction networks emphasizing distinct subnetworks, including cytokine-mediated TNFα signaling via NF-κB, Wnt/LRP/DKK signaling and cell cycle regulation. This study reveals differential transcriptomic responses under SH and IH characterised by HIF-1α transcriptional response induced only by SH, but not by IH. The study also features the potential molecular events that may occur at the vascular level in OSA.

Human cerumen analysis is an innovative and non-invasive trend in diagnosing diseases. Recently, new cerumen volatile-based methods using binary (volatile presence/absence) and semiquantitative (volatile intensity) data approaches have shown great potential in detecting biomarkers for cancer, chronic and rare diseases, and xenobiotic exposures. However, to date, the impacts of demographic factors such as body mass index (BMI), sex, age, and ethnicity/race in cerumen data have not been widely described, which can hamper interpretation in biomarker discovery investigations. This study examined the effects of such factors in cerumen, defining the baseline volatile organic metabolites (VOMs) across different physiological groups. Cerumen samples from seventy volunteers were analyzed using headspace/gas chromatography-mass spectrometry (HS/GC-MS) and multivariate statistical analysis using binary and semiquantitative data approaches. In the binary data approach, several VOMs exhibited patterns of high occurrence in some specific demographic groups. However, no pattern of discrimination that could be attributed to demographic factors was observed. In the semiquantitative approach, the relative abundance of cerumen VOMs was more impacted by sex and BMI than age and ethnicity/race. In summary, we describe how cerumen VOM occurrence and abundance are affected by patient phenotype, which can pave the way for more personalized medicine in future cerumen volatile-based methods.

Pancreatic β-cells are composed of different subtypes that play a key role in the control of insulin secretion and thereby control glucose homeostasis. In vitro differentiation of human induced pluripotent stem cells (hiPSCs) into 3D spheroids leads to the generation of β-cell subtypes and thus to the development of islet-like structures. Using this cutting-edge cell model, the aim of the study was to decipher the signaling signature that underlines β-cell subtypes, with a focus on the search for the activity of motifs of important transcription regulators (TRs). The investigation was performed using data from previous single-cell sequencing analysis introduced into the integrated system for motif activity response analysis (ISMARA) of transcription regulators. We extracted the matrix of important TRs activated in the β-cell subpopulation and bi-hormonal-like β-cells. Based on these TRs and their targets, we built specific regulatory networks for main cell subpopulations. Our data confirmed the transcriptomic heterogeneity of the β-cell subtype lineage and suggested a mechanism that could account for the differentiation of β-cell subtypes during pancreas development. We do believe that our findings could be instrumental for understanding the mechanisms that affect the balance of β-cell subtypes, leading to impaired insulin secretion in type 2 diabetes.

Forensic science, an interdisciplinary field encompassing the collection, examination, and presentation of evidence in legal proceedings, has recently embraced lipidomics as a valuable tool. Lipidomics, a subfield of metabolomics, specializes in the analysis of lipid structures and functions, offering insights into biological processes that can aid forensic investigations. While not a substitute for DNA analysis in personal identification, lipidomics complements this technique by focusing on small biological molecules, with distinct sample requirements. This review comprehensively explores the current applications of lipidomics in forensic science. The review commences with an introduction to the concept and historical background of lipidomics, subsequently delving into its utilization in diverse areas such as drug analysis, ethyl alcohol and substitute assessment, latent fingermark detection, fire debris analysis, and seafood authentication. By showcasing the various biological materials and methods employed, this review underscores the potential of lipidomics as a powerful adjunct in forensic investigations.

Lentiviral vectors (LV) are emerging tools for genetic therapies and novel cancer treatments. While effective, LV-based therapies have extremely large costs associated with their manufacturing and delivery. LV technology descends from human immunodeficiency virus (HIV), whose lipid envelope has been previously measured and shown to have a direct impact on its transduction efficiency. We developed a rapid, robust, and sensitive untargeted lipidomics pipeline to analyze novel LV biotherapeutic products and demonstrate its utility on HEK 293T packaging cells and concentrated culture media containing LV. The impact of 48 hours of LV production on the lipidome of HEK 293T cells was measured and compared to the expression of vesicular stomatitis virus G protein (VSV G) over the same timeframe. 151 lipids were identified in HEK 293T packaging cells, 84 of which had fold changes with FDR-corrected P < 0.05 compared to HEK 293T treated with media. It was found that fold changes with FDR-adjusted P < 0.05 after VSV G expression and LV production were highly correlated (R² = 0.89). Concentrating LV in culture media led to the identification of 102 lipids, half of which were determined to be unique LV virion lipids after subtracting the media lipidome. Our approach can be readily used to study the lipid dynamics of large-scale LV production and be rapidly translated into targeted methods to quantify individual lipid components or applied to other viral vector platforms.

Coupling size exclusion chromatography (SEC) with mass spectrometry-based proteomics enables investigating protein complexes, with degradomic profiling providing deeper insights into complex-associated proteolytic processing and retaining of cleavage products. This study aims to map protein complex formation upon inflammasome activation in bone marrow-derived dendritic cells (BMDCs) from gasdermin D-deficient mice, focusing on proteolytic enzymes and truncated proteins in higher molecular weight complexes. Cultured BMDCs were primed with LPS and subsequently treated with nigericin or Val-boroPro (VbP). SEC-fractionated proteins were TMT-labelled and analyzed via liquid chromatography-tandem mass spectrometry (LC-MS/MS). We identified 6862 proteins and 70 802 peptides, including 14 714 semi-tryptic peptides indicating elevated endogenous proteolytic processing. The sequence motif of numerous cleavage sites maps to caspase-like activity. Inflammasome activation was corroborated by elevated levels of apoptosis-associated speck-like protein containing a CARD (ASC) in higher molecular weight (MW) fractions and increased IL-1β levels in low MW fractions upon nigericin or VbP treatment. The majority of truncated cleavage products remained within their corresponding, higher MW protein complexes while caspase-specific cleavage products of Rho-associated protein kinase 1, gelsolin, and AP-2 complex subunit alpha-2 dissociated to lower MW fractions. SEC profiles identified 174 proteases, with cell surface proteases forming high MW complexes, including ADAMs and DPP4 but not MMP14. VbP treatment led to the accumulation of ISG15 in low MW fractions while RNA polymerase II coactivator p15 shifted to higher MW fractions. This study demonstrates that SEC-coupled proteomics and degradomic profiling offer unique insights into protein complex dynamics and proteolytic processes upon inflammasome activation.

Lipid metabolism is recognised as being central to growth, disease and health. Lipids, therefore, have an important place in current research on globally significant topics such as food security and biodiversity loss. However, answering questions in these important fields of research requires not only identification and measurement of lipids in a wider variety of sample types than ever before, but also hypothesis-driven analysis of the resulting ‘big data’. We present a novel pipeline that can collect data from a wide range of biological sample types, taking 1 000 000 lipid measurements per 384 well plate, and analyse the data systemically. We provide evidence of the power of the tool through proof-of-principle studies using edible fish (mackerel, bream, seabass) and colonies of Bombus terrestris. Bee colonies were found to be more like mini-ecosystems and there was evidence for considerable changes in lipid metabolism in bees through key developmental stages. This is the first report of either high throughput LCMS lipidomics or systemic analysis in individuals, colonies and ecosystems. This novel approach provides new opportunities to analyse metabolic systems at different scales at a level of detail not previously feasible, to answer research questions about societally important topics.

We would like to take this opportunity to thank all of Molecular Omics’ reviewers for helping to preserve quality and integrity in chemical science literature. We would also particularly like to highlight the Outstanding Reviewers for Molecular Omics in 2023.

The natural product 9-methoxystrobilurin G (9MG) from Favolaschia spp basidiomycetes is a potent and selective antimalarial. The mechanism of action of 9MG is unknown. We induced 9MG resistance in Plasmodium falciparum 3D7 and Dd2 strains and identified mutations associated with resistance by genome sequencing. All 9MG-resistant clones possessed missense mutations in the cytochrome b (CYTB) gene, a key component of mitochondrial complex III. The mutations map to the quinol oxidation site of CYTB, which is also the target of antimalarials such as atovaquone. In a complementary approach to identify protein targets of 9MG, a photoactivatable derivative of 9MG was synthesized and applied in chemoproteomic-based target profiling. Three components of mitochondrial complex III (QCR7, QCR9, and COX15) were specifically enriched consistent with 9MG targeting CYTB and complex III function in P. falciparum. Inhibition of complex III activity by 9MG was confirmed by ubiquinone cytochrome c reductase assay using P. falciparum extract. The findings from this study may be useful for developing novel antimalarials targeting CYTB.