QSample: An Automated System for Rapid Monitoring of Quality Indicators in Proteomics Samples.

Mass spectrometry-based proteomics is an essential technique in contemporary biomedicine, offering quantitative, sensitive, and rapid analysis of proteomes. Recent advancements in mass spectrometry have enabled the acquisition of data from increasingly large-scale experiments, often conducted in core facilities and research infrastructures. While automated tools exist to assess instrument performance using predefined control samples, the analysis of experimental samples typically occurs postacquisition, which can delay decision-making and lead to potential data integrity issues.

Claudin-6 mediates an embryonic stem cell-driven antitumor response against breast cancer.

Stem cells are emerging sources of antigens for cancer immunotherapy. Here, we used TNG-A mouse embryonic stem cells to trigger an anticancer response against tumors derived from E0771 mouse breast cancer cells, possibly mediated by the mouse immune system. TNG-A cells were cultured in the presence or absence of two specific kinase inhibitors.

LAP1 Interactome Profiling Provides New Insights into LAP1's Physiological Functions.

The nuclear envelope (NE), a protective membrane bordering the nucleus, is composed of highly specialized proteins that are indispensable for normal cellular activity. Lamina-associated polypeptide 1 (LAP1) is a NE protein whose functions are just beginning to be unveiled. The fact that mutations causing LAP1 deficiency are extremely rare and pathogenic is indicative of its paramount importance to preserving human health, anticipating that LAP1 might have a multifaceted role in the cell.

Spectral libraries from nucleobases and deoxyribonucleosides facilitate the identification of ribonucleosides by nano-flow liquid chromatography-tandem mass spectrometry.

Rationale: The study addresses the challenge of identifying RNA post-transcriptional modifications when commercial standards are not available to generate reference spectral libraries. It proposes employing homologous nucleobases and deoxyribonucleosides as alternative reference spectral libraries to aid in identifying modified ribonucleosides and distinguishing them from their positional isomers when the standards are unavailable.

Methods: Complete sets of ribonucleoside, deoxyribonucleoside and nucleobase standards were analyzed using high-performance nano-flow liquid chromatography coupled to an Orbitrap Eclipse Tribrid mass spectrometer.

Expression of correlates with breast cancer aggressiveness and protects breast cancer cells from chemotherapy.

The recently discovered human lncRNA is induced after DNA damage in a p53-dependent manner. It plays a critical role in the maintenance of genomic stability through interaction with Pumilio proteins, limiting the repression of their target mRNAs. Therefore, inactivation causes chromosomal instability and aneuploidy, which contributes to the accumulation of genetic abnormalities and tumorigenesis.

Loss of dyskerin facilitates the acquisition of metastatic traits by altering the mevalonate pathway.

The initial dissemination of cancer cells from many primary tumors implies intravasation to lymphatic nodes or blood vessels. To investigate the mechanisms involved, we analyzed the expression of small non-coding RNAs in cutaneous squamous cell carcinoma (cSCC), a prevalent tumor that mainly spreads to lymph nodes. We report the reduced expression of small nucleolar RNAs in primary cSCCs that metastasized when compared to non-metastasizing cSCCs, and the progressive loss of DKC1 (dyskerin, which stabilizes the small nucleolar RNAs) along the metastasis.

High-performance nano-flow liquid chromatography column combined with high- and low-collision energy data-independent acquisition enables targeted and discovery identification of modified ribonucleotides by mass spectrometry.

Over 170 post-transcriptional RNA modifications have been described and are common in all kingdoms of life. These modifications range from methylation to complex chemical structures, with methylation being the most abundant. RNA modifications play a key role in RNA folding and function and their dysregulation in humans has been linked to several diseases such as cancer, metabolic diseases or neurological disorder.

The loss of DHX15 impairs endothelial energy metabolism, lymphatic drainage and tumor metastasis in mice.

DHX15 is a downstream substrate for Akt1, which is involved in key cellular processes affecting vascular biology. Here, we explored the vascular regulatory function of DHX15. Homozygous DHX15 gene deficiency was lethal in mouse and zebrafish embryos.

Covalent Histone Modification by an Electrophilic Derivative of the Anti-HIV Drug Nevirapine.

Nevirapine (NVP), a non-nucleoside reverse transcriptase inhibitor widely used in combined antiretroviral therapy and to prevent mother-to-child transmission of the human immunodeficiency virus type 1, is associated with several adverse side effects. Using 12-mesyloxy-nevirapine, a model electrophile of the reactive metabolites derived from the NVP Phase I metabolite, 12-hydroxy-NVP, we demonstrate that the nucleophilic core and -terminal residues of histones are targets for covalent adduct formation. We identified multiple NVP-modification sites at lysine (e.

QCloud2: An Improved Cloud-based Quality-Control System for Mass-Spectrometry-based Proteomics Laboratories.

QCloud is a cloud-based system to support proteomics laboratories in daily quality assessment using a user-friendly interface, easy setup, and automated data processing. Since its release, QCloud has facilitated automated quality control for proteomics experiments in many laboratories. QCloud provides a quick and effortless evaluation of instrument performance that helps to overcome many analytical challenges derived from clinical and translational research.

The cochaperone CHIP marks Hsp70- and Hsp90-bound substrates for degradation through a very flexible mechanism.

Some molecular chaperones are involved not only in assisting the folding of proteins but also, given appropriate conditions, in their degradation. This is the case for Hsp70 and Hsp90 which, in concert with the cochaperone CHIP, direct their bound substrate to degradation through ubiquitination. We generated complexes between the chaperones (Hsp70 or Hsp90), the cochaperone CHIP and, as substrate, a p53 variant containing the GST protein (p53-TMGST).

Knock-down of AHCY and depletion of adenosine induces DNA damage and cell cycle arrest.

Recently, functional connections between S-adenosylhomocysteine hydrolase (AHCY) activity and cancer have been reported. As the properties of AHCY include the hydrolysis of S-adenosylhomocysteine and maintenance of the cellular methylation potential, the connection between AHCY and cancer is not obvious. The mechanisms by which AHCY influences the cell cycle or cell proliferation have not yet been confirmed.

Proteomic Profiling of Microtubule Self-organization in M-phase.

Microtubules (MTs) and associated proteins can self-organize into complex structures such as the bipolar spindle, a process in which RanGTP plays a major role. Addition of RanGTP to M-phase egg extracts promotes the nucleation and self-organization of MTs into asters and bipolar-like structures in the absence of centrosomes or chromosomes. We show here that the complex proteome of these RanGTP-induced MT assemblies is similar to that of mitotic spindles.

QCloud: A cloud-based quality control system for mass spectrometry-based proteomics laboratories.

The increasing number of biomedical and translational applications in mass spectrometry-based proteomics poses new analytical challenges and raises the need for automated quality control systems. Despite previous efforts to set standard file formats, data processing workflows and key evaluation parameters for quality control, automated quality control systems are not yet widespread among proteomics laboratories, which limits the acquisition of high-quality results, inter-laboratory comparisons and the assessment of variability of instrumental platforms. Here we present QCloud, a cloud-based system to support proteomics laboratories in daily quality assessment using a user-friendly interface, easy setup, automated data processing and archiving, and unbiased instrument evaluation.

Proteome profiling of clear cell renal cell carcinoma in von Hippel-Lindau patients highlights upregulation of Xaa-Pro aminopeptidase-1, an anti-proliferative and anti-migratory exoprotease.

Patients of the von Hippel-Lindau (VHL) disease frequently develop clear cell renal cell carcinoma (ccRCC). Using archived, formalin-fixed, paraffin-embedded (FFPE) samples, we sought to determine global proteome alterations that distinguish ccRCC tissue from adjacent, non-malignant kidney tissue in VHL-patients. Our quantitative proteomic analysis clearly discriminated tumor and non-malignant tissue.

Proteotoxicity in cardiac amyloidosis: amyloidogenic light chains affect the levels of intracellular proteins in human heart cells.

AL amyloidosis is characterized by widespread deposition of immunoglobulin light chains (LCs) as amyloid fibrils. Cardiac involvement is frequent and leads to life-threatening cardiomyopathy. Besides the tissue alteration caused by fibrils, clinical and experimental evidence indicates that cardiac damage is also caused by proteotoxicity of prefibrillar amyloidogenic species.

Evaluation of different peptide fragmentation types and mass analyzers in data-dependent methods using an Orbitrap Fusion Lumos Tribrid mass spectrometer.

One of the major additions in MS technology has been the irruption of the Orbitrap mass analyzer, which has boosted the proteomics analyses of biological complex samples since its introduction. Here, we took advantage of the capabilities of the new Orbitrap Fusion Lumos Tribrid mass spectrometer to assess the performance of different data-dependent acquisition methods for the identification and quantitation of peptides and phosphopeptides in single-shot analysis of human whole cell lysates. Our study explored the capabilities of tri-hibrid mass spectrometers for (phospho-) peptide identification and quantitation using different gradient lengths, sample amounts, and combinations of different peptide fragmentation types and mass analyzers.

A multicentric study to evaluate the use of relative retention times in targeted proteomics.

Unlabelled: Despite the maturity reached by targeted proteomic strategies, reliable and standardized protocols are urgently needed to enhance reproducibility among different laboratories and analytical platforms, facilitating a more widespread use in biomedical research. To achieve this goal, the use of dimensionless relative retention times (iRT), defined on the basis of peptide standard retention times (RT), has lately emerged as a powerful tool. The robustness, reproducibility and utility of this strategy were examined for the first time in a multicentric setting, involving 28 laboratories that included 24 of the Spanish network of proteomics laboratories (ProteoRed-ISCIII).

Signatures of Evolutionary Adaptation in Quantitative Trait Loci Influencing Trace Element Homeostasis in Liver.

Essential trace elements possess vital functions at molecular, cellular, and physiological levels in health and disease, and they are tightly regulated in the human body. In order to assess variability and potential adaptive evolution of trace element homeostasis, we quantified 18 trace elements in 150 liver samples, together with the expression levels of 90 genes and abundances of 40 proteins involved in their homeostasis. Additionally, we genotyped 169 single nucleotide polymorphism (SNPs) in the same sample set.

Profiling the secretome and extracellular proteome of the potato late blight pathogen Phytophthora infestans.

Oomycetes are filamentous organisms that cause notorious diseases, several of which have a high economic impact. Well known is Phytophthora infestans, the causal agent of potato late blight. Previously, in silico analyses of the genome and transcriptome of P.

Lifelong exercise training modulates cardiac mitochondrial phosphoproteome in rats.

Moderate physical activity has traditionally been associated with the improvement of cardiac function and, consequently, with the extension of life span. Mitochondria play a key role in the adaptation of heart muscle to exercise-related metabolic demands. In order to disclose the molecular mechanisms underlying the beneficial effect of lifelong physical activity in cardiac function, we performed label-free quantitative mass spectrometry-based proteomics of Sprague-Dawley rat heart mitochondrial proteome and phosphoproteome.

Cutting edge proteomics: benchmarking of six commercial trypsins.

Tryptic digestion is an important component of most proteomics experiments, and trypsin is available from many sources with a cost that varies by more than 1000-fold. This high-mass-accuracy LC-MS study benchmarks six commercially available trypsins with respect to autolytic species and sequence specificity. The analysis of autolysis products led to the identification of a number of contaminating proteins and the generation of a list of peptide species that will be present in tryptic digests.