De novo and inherited dominant variants in U4 and U6 snRNAs cause retinitis pigmentosa.

The U4 small nuclear RNA (snRNA) forms a duplex with the U6 snRNA and, together with U5 and ∼30 proteins, is part of the U4/U6.U5 tri-snRNP complex, located at the core of the major spliceosome. Recently, recurrent variants in the U4 RNA, transcribed from the gene, and in at least two other genes were discovered to cause neurodevelopmental disorder.

HiFi long-read genomes for difficult-to-detect, clinically relevant variants.

Clinical short-read exome and genome sequencing approaches have positively impacted diagnostic testing for rare diseases. Yet, technical limitations associated with short reads challenge their use for the detection of disease-associated variation in complex regions of the genome. Long-read sequencing (LRS) technologies may overcome these challenges, potentially qualifying as a first-tier test for all rare diseases.

XCMS-METLIN: data-driven metabolite, lipid, and chemical analysis.

In this Correspondence, G. Siuzdak and colleagues present XCMS-METLIN, an extensive resource for metabolomics, lipidomics, and chemical analysis. [Image: see text]

Immune suppression by human thymus-derived effector Tregs relies on glucose/lactate-fueled fatty acid synthesis.

Regulatory T cells (Tregs) suppress pro-inflammatory conventional T cell (Tconv) responses. As lipids impact cell signaling and function, we compare the lipid composition of CD4 thymus-derived (t)Tregs and Tconvs. Lipidomics reveal constitutive enrichment of neutral lipids in Tconvs and phospholipids in tTregs.

The Neurolipid Atlas: a lipidomics resource for neurodegenerative diseases uncovers cholesterol as a regulator of astrocyte reactivity impaired by ApoE4.

Lipid changes in the brain have been implicated in many neurodegenerative diseases including Alzheimer's Disease (AD), Parkinson's disease and Amyotrophic Lateral Sclerosis. To facilitate comparative lipidomic research across brain-diseases we established a data commons named the Neurolipid Atlas, that we have pre-populated with novel human, mouse and isogenic induced pluripotent stem cell (iPSC)-derived lipidomics data for different brain diseases. We show that iPSC-derived neurons, microglia and astrocytes display distinct lipid profiles that recapitulate lipotypes.

Unravelling undiagnosed rare disease cases by HiFi long-read genome sequencing.

Solve-RD is a pan-European rare disease (RD) research program that aims to identify disease-causing genetic variants in previously undiagnosed RD families. We utilised 10-fold coverage HiFi long-read sequencing (LRS) for detecting causative structural variants (SVs), single nucleotide variants (SNVs), insertion-deletions (InDels), and short tandem repeat (STR) expansions in extensively studied RD families without clear molecular diagnoses. Our cohort includes 293 individuals from 114 genetically undiagnosed RD families selected by European Rare Disease Network (ERN) experts.

Future of Dutch NGS-Based Newborn Screening: Exploring the Technical Possibilities and Assessment of a Variant Classification Strategy.

In this study, we compare next-generation sequencing (NGS) approaches (targeted panel (tNGS), whole exome sequencing (WES), and whole genome sequencing (WGS)) for application in newborn screening (NBS). DNA was extracted from dried blood spots (DBS) from 50 patients with genetically confirmed inherited metabolic disorders (IMDs) and 50 control samples. One hundred IMD-related genes were analyzed.

Tandem Mass Spectrometry across Platforms.

PubChem serves as a comprehensive repository, housing over 100 million unique chemical structures representing the breadth of our chemical knowledge across numerous fields including metabolism, pharmaceuticals, toxicology, cosmetics, agriculture, and many more. Rapid identification of these small molecules increasingly relies on electrospray ionization (ESI) paired with tandem mass spectrometry (MS/MS), particularly by comparison to genuine standard MS/MS data sets. Despite its widespread application, achieving consistency in MS/MS data across various analytical platforms remains an unaddressed concern.

Spatially and temporally resolved metabolome of the human oral cavity.

Saliva is a complex bodily fluid composed of secretions by major and minor salivary glands. Salivary glands and their secretions are known to be unevenly distributed in the human oral cavity. Moreover, saliva flow rate and composition vary across locations and time of the day.

A Clinically Selected Staphylococcus aureus Mutant Survives Daptomycin Treatment by Reducing Binding of the Antibiotic and Adapting a Rod-Shaped Morphology.

Daptomycin is a last-resort antibiotic used for the treatment of infections caused by Gram-positive antibiotic-resistant bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA). Treatment failure is commonly linked to accumulation of point mutations; however, the contribution of single mutations to resistance and the mechanisms underlying resistance remain incompletely understood. Here, we show that a single nucleotide polymorphism (SNP) selected during daptomycin therapy inactivates the highly conserved ClpP protease and is causing reduced susceptibility of MRSA to daptomycin, vancomycin, and β-lactam antibiotics as well as decreased expression of virulence factors.

Comprehensive de novo mutation discovery with HiFi long-read sequencing.

Background: Long-read sequencing (LRS) techniques have been very successful in identifying structural variants (SVs). However, the high error rate of LRS made the detection of small variants (substitutions and short indels < 20 bp) more challenging. The introduction of PacBio HiFi sequencing makes LRS also suited for detecting small variation.

High-Throughput Venomics.

In this study, we present high-throughput (HT) venomics, a novel analytical strategy capable of performing a full proteomic analysis of a snake venom within 3 days. This methodology comprises a combination of RP-HPLC-nanofractionation analytics, mass spectrometry analysis, automated in-solution tryptic digestion, and high-throughput proteomics. In-house written scripts were developed to process all the obtained proteomics data by first compiling all Mascot search results for a single venom into a single Excel sheet.

Membrane lipid remodeling modulates γ-secretase processivity.

Imbalances in the amounts of amyloid-β peptides (Aβ) generated by the membrane proteases β- and γ-secretase are considered as a trigger of Alzheimer's disease (AD). Cell-free studies of γ-secretase have shown that increasing membrane thickness modulates Aβ generation but it has remained unclear if these effects are translatable to cells. Here we show that the very long-chain fatty acid erucic acid (EA) triggers acyl chain remodeling in AD cell models, resulting in substantial lipidome alterations which included increased esterification of EA in membrane lipids.

Urinary Metabolic Profiling in Volunteers Undergoing Malaria Challenge in Gabon.

The interaction of malaria parasites with their human host is extensively studied, yet only few studies reported how infection affects urinary metabolite profiles and how this is associated with immunity. We present a longitudinal study of the urinary metabolic profiles of twenty healthy Africans with lifelong exposure to malaria and five malaria-naïve Europeans, who were all challenged with direct venous inoculation of live sporozoïtes (PfSPZ) and followed up until they developed symptoms or became thick blood smear positive (TBS). Urine samples were collected before and at 2, 5, 9 and 11 days post challenge and were analysed.

Diagnostic analysis of the highly complex OPN1LW/OPN1MW gene cluster using long-read sequencing and MLPA.

Pathogenic variants in the OPN1LW/OPN1MW gene cluster are causal for a range of mild to severe visual impairments with color deficiencies. The widely utilized short-read next-generation sequencing (NGS) is inappropriate for the analysis of the OPN1LW/OPN1MW gene cluster and many patients with pathogenic variants stay underdiagnosed. A diagnostic genetic assay was developed for the OPN1LW/OPN1MW gene cluster, consisting of copy number analysis via multiplex ligation-dependent probe amplification and sequence analysis via long-read circular consensus sequencing.

The performance of genome sequencing as a first-tier test for neurodevelopmental disorders.

Genome sequencing (GS) can identify novel diagnoses for patients who remain undiagnosed after routine diagnostic procedures. We tested whether GS is a better first-tier genetic diagnostic test than current standard of care (SOC) by assessing the technical and clinical validity of GS for patients with neurodevelopmental disorders (NDD). We performed both GS and exome sequencing in 150 consecutive NDD patient-parent trios.

Generating Heterokaryotic Cells via Bacterial Cell-Cell Fusion.

Fusion of cells is an important and common biological process that leads to the mixing of cellular contents and the formation of multinuclear cells. Cell fusion occurs when distinct membranes are brought into proximity of one another and merge to become one. Fusion holds promise for biotechnological innovations, for instance, for the discovery of urgently needed new antibiotics.

Neutral Loss Mass Spectral Data Enhances Molecular Similarity Analysis in METLIN.

Neutral loss (NL) spectral data presents a mirror of MS data and is a valuable yet largely untapped resource for molecular discovery and similarity analysis. Tandem mass spectrometry (MS) data is effective for the identification of known molecules and the putative identification of novel, previously uncharacterized molecules (unknowns). Yet, MS data alone is limited in characterizing structurally related molecules.

Long-read technologies identify a hidden inverted duplication in a family with choroideremia.

The lack of molecular diagnoses in rare genetic diseases can be explained by limitations of current standard genomic technologies. Upcoming long-read techniques have complementary strengths to overcome these limitations, with a particular strength in identifying structural variants. By using optical genome mapping and long-read sequencing, we aimed to identify the pathogenic variant in a large family with X-linked choroideremia.

Spatially resolved sampling of the human oral cavity for metabolic profiling.

Here, we present a spatially resolved sampling protocol for the oral human cavity aimed at untargeted metabolomics. We describe the spatial collection of salivary biospecimens, their preparation, and subsequent mass-spectrometry-based untargeted metabolomics analysis. Our protocol avoids complex procedures generally required for gland-specific saliva collection.

Proteomics with Enhanced In-Source Fragmentation/Annotation: Applying XCMS-EISA Informatics and Q-MRM High-Sensitivity Quantification.

Enhanced in-source fragmentation/annotation (EISA) has recently been shown to produce fragment ions that match tandem mass spectrometry data across a wide range of small molecules. EISA has been developed to facilitate data-dependent acquisition (DDA), data-independent acquisiton (DIA), and multiple-reaction monitoring (MRM), enabling molecular identifications in untargeted metabolomics and targeted quantitative single-quadrupole MRM (Q-MRM) analyses. Here, EISA has been applied to peptide-based proteomic analysis using optimized in-source fragmentation to generate fragmentation patterns for a mixture of 38 peptides, which were comparable to the b- and y-type fragment ions typically observed in tandem MS experiments.