The STRIPAK complex is required for radial sorting and laminin receptor expression in Schwann cells.

During peripheral nervous system development, Schwann cells undergo Rac1-dependent cytoskeletal reorganization as they insert cytoplasmic extensions into axon bundles to radially sort, ensheath, and myelinate individual axons. However, our understanding of the direct effectors targeted by Rac1 is limited. Here, we demonstrate that striatin-3 and MOB4 are novel Rac1 interactors.

Combining Data Independent Acquisition With Spike-In SILAC (DIA-SiS) Improves Proteome Coverage and Quantification.

Data-independent acquisition (DIA) is increasingly preferred over data-dependent acquisition due to its higher throughput and fewer missing values. Whereas data-dependent acquisition often uses stable isotope labeling to improve quantification, DIA mostly relies on label-free approaches. Efforts to integrate DIA with isotope labeling include chemical methods like mass differential tags for relative and absolute quantification and dimethyl labeling, which, while effective, complicate sample preparation.

Pathogenic mutations of human phosphorylation sites affect protein-protein interactions.

Despite their lack of a defined 3D structure, intrinsically disordered regions (IDRs) of proteins play important biological roles. Many IDRs contain short linear motifs (SLiMs) that mediate protein-protein interactions (PPIs), which can be regulated by post-translational modifications like phosphorylation. 20% of pathogenic missense mutations are found in IDRs, and understanding how such mutations affect PPIs is essential for unraveling disease mechanisms.

ADAM19 cleaves the PTH receptor and associates with brachydactyly type E.

Brachydactyly type E (BDE), shortened metacarpals, metatarsals, cone-shaped epiphyses, and short stature commonly occurs as a sole phenotype. Parathyroid hormone-like protein (PTHrP) has been shown to be responsible in all forms to date, either directly or indirectly. We used linkage and then whole genome sequencing in a small pedigree, to elucidate BDE and identified a truncated disintegrin-and-metalloproteinase-19 (ADAM19) allele in all affected family members, but not in nonaffected persons.

Cryo-EM captures early ribosome assembly in action.

Ribosome biogenesis is a fundamental multi-step cellular process in all domains of life that involves the production, processing, folding, and modification of ribosomal RNAs (rRNAs) and ribosomal proteins. To obtain insights into the still unexplored early assembly phase of the bacterial 50S subunit, we exploited a minimal in vitro reconstitution system using purified ribosomal components and scalable reaction conditions. Time-limited assembly assays combined with cryo-EM analysis visualizes the structurally complex assembly pathway starting with a particle consisting of ordered density for only ~500 nucleotides of 23S rRNA domain I and three ribosomal proteins.

Monitoring mitochondrial translation by pulse SILAC.

Mitochondrial ribosomes are specialized to translate the 13 membrane proteins encoded in the mitochondrial genome, which shapes the oxidative phosphorylation complexes essential for cellular energy metabolism. Despite the importance of mitochondrial translation (MT) control, it is challenging to identify and quantify the mitochondrial-encoded proteins because of their hydrophobic nature and low abundance. Here, we introduce a mass spectrometry-based proteomic method that combines biochemical isolation of mitochondria with pulse stable isotope labeling by amino acids in cell culture.

Combining Metabolic Pulse Labeling and Quantitative Proteomics to Monitor Protein Synthesis Upon Viral Infection.

Viruses like influenza A virus (IAV) hijack host cells in order to replicate. To actively and abundantly synthesize viral proteins, they reprogram the cellular transcriptional and translational landscape. Here, we present a proteomic approach that allows us to quantify the differences in host and viral protein synthesis comparatively for different strains of IAV.

A critical period of translational control during brain development at codon resolution.

Translation modulates the timing and amplification of gene expression after transcription. Brain development requires uniquely complex gene expression patterns, but large-scale measurements of translation directly in the prenatal brain are lacking. We measure the reactants, synthesis and products of mRNA translation spanning mouse neocortex neurogenesis, and discover a transient window of dynamic regulation at mid-gestation.

A Novel Gene Controls a New Structure: PiggyBac Transposable Element-Derived 1, Unique to Mammals, Controls Mammal-Specific Neuronal Paraspeckles.

Although new genes can arrive from modes other than duplication, few examples are well characterized. Given high expression in some human brain subregions and a putative link to psychological disorders [e.g.

Therapeutic targeting of ATR in alveolar rhabdomyosarcoma.

Despite advances in multi-modal treatment approaches, clinical outcomes of patients suffering from PAX3-FOXO1 fusion oncogene-expressing alveolar rhabdomyosarcoma (ARMS) remain dismal. Here we show that PAX3-FOXO1-expressing ARMS cells are sensitive to pharmacological ataxia telangiectasia and Rad3 related protein (ATR) inhibition. Expression of PAX3-FOXO1 in muscle progenitor cells is not only sufficient to increase sensitivity to ATR inhibition, but PAX3-FOXO1-expressing rhabdomyosarcoma cells also exhibit increased sensitivity to structurally diverse inhibitors of ATR.

Compound Interaction Screen on a Photoactivatable Cellulose Membrane (CISCM) Identifies Drug Targets.

Identifying the protein targets of drugs is an important but tedious process. Existing proteomic approaches enable unbiased target identification but lack the throughput needed to screen larger compound libraries. Here, we present a compound interaction screen on a photoactivatable cellulose membrane (CISCM) that enables target identification of several drugs in parallel.

Qualitative Assessment of Titanium versus Carbon Fiber/Polyetheretherketone Pedicle Screw-Related Artifacts: A Cadaveric Study.

Background: Dorsal instrumentation and decompression are the mainstays of spinal tumor treatment. Replacing titanium screws with carbon fiber-reinforced polyetheretherketone (CFRP) screws can reduce imaging artifacts on neural structures and perturbations of radiation dose. Further reduction of metal content in such screws might enhance the benefit.

In Vitro Kinase-to-Phosphosite Database (iKiP-DB) Predicts Kinase Activity in Phosphoproteomic Datasets.

Phosphoproteomics routinely quantifies changes in the levels of thousands of phosphorylation sites, but functional analysis of such data remains a major challenge. While databases like PhosphoSitePlus contain information about many phosphorylation sites, the vast majority of known sites is not assigned to any protein kinase. Assigning changes in the phosphoproteome to the activity of individual kinases therefore remains a key challenge.

Physiological Functions of Intracellular Protein Degradation.

While cellular proteins were initially thought to be stable, research over the last decades has firmly established that intracellular protein degradation is an active and highly regulated process: Lysosomal, proteasomal, and mitochondrial degradation systems were identified and found to be involved in a staggering number of biological functions. Here, we provide a global overview of the diverse roles of cellular protein degradation using seven categories: homeostasis, regulation, quality control, stoichiometry control, proteome remodeling, immune surveillance, and baseline turnover. Using selected examples, we outline how proteins are degraded and why this is functionally relevant.

HDLBP binds ER-targeted mRNAs by multivalent interactions to promote protein synthesis of transmembrane and secreted proteins.

The biological role of RNA-binding proteins in the secretory pathway is not well established. Here, we describe that human HDLBP/Vigilin directly interacts with more than 80% of ER-localized mRNAs. PAR-CLIP analysis reveals that these transcripts represent high affinity HDLBP substrates and are specifically bound in their coding sequences (CDS), in contrast to CDS/3'UTR-bound cytosolic mRNAs.

GIMAP6 regulates autophagy, immune competence, and inflammation in mice and humans.

Inborn errors of immunity (IEIs) unveil regulatory pathways of human immunity. We describe a new IEI caused by mutations in the GTPase of the immune-associated protein 6 (GIMAP6) gene in patients with infections, lymphoproliferation, autoimmunity, and multiorgan vasculitis. Patients and Gimap6-/- mice show defects in autophagy, redox regulation, and polyunsaturated fatty acid (PUFA)-containing lipids.

MYCN mediates cysteine addiction and sensitizes neuroblastoma to ferroptosis.

Aberrant expression of MYC transcription factor family members predicts poor clinical outcome in many human cancers. Oncogenic MYC profoundly alters metabolism and mediates an antioxidant response to maintain redox balance. Here we show that MYCN induces massive lipid peroxidation on depletion of cysteine, the rate-limiting amino acid for glutathione (GSH) biosynthesis, and sensitizes cells to ferroptosis, an oxidative, non-apoptotic and iron-dependent type of cell death.

Proteome-wide quantitative RNA-interactome capture identifies phosphorylation sites with regulatory potential in RBM20.

Cellular mRNA-binding proteins (mRBPs) are major posttranscriptional regulators of gene expression. Although many posttranslational modification sites in mRBPs have been identified, little is known about how these modifications regulate mRBP function. Here, we developed quantitative RNA-interactome capture (qRIC) to quantify the fraction of mRBPs pulled down with polyadenylated mRNAs.

Continuous Intraarterial Nimodipine Infusion for the Treatment of Delayed Cerebral Ischemia After Aneurysmal Subarachnoid Hemorrhage: A Retrospective, Single-Center Cohort Trial.

Background: Delayed cerebral ischemia (DCI) occurs after aneurysmal subarachnoid hemorrhage (aSAH). Continuous intraarterial nimodipine infusion (CIAN) is a promising approach in patients with intracranial large vessel vasospasm (LVV). The objective of this retrospective single-center cohort study was to evaluate the outcome in aSAH-patients treated with CIAN.

SARS-CoV-2 infection triggers profibrotic macrophage responses and lung fibrosis.

COVID-19-induced "acute respiratory distress syndrome" (ARDS) is associated with prolonged respiratory failure and high mortality, but the mechanistic basis of lung injury remains incompletely understood. Here, we analyze pulmonary immune responses and lung pathology in two cohorts of patients with COVID-19 ARDS using functional single-cell genomics, immunohistology, and electron microscopy. We describe an accumulation of CD163-expressing monocyte-derived macrophages that acquired a profibrotic transcriptional phenotype during COVID-19 ARDS.

Neuroblastoma signalling models unveil combination therapies targeting feedback-mediated resistance.

Very high risk neuroblastoma is characterised by increased MAPK signalling, and targeting MAPK signalling is a promising therapeutic strategy. We used a deeply characterised panel of neuroblastoma cell lines and found that the sensitivity to MEK inhibitors varied drastically between these cell lines. By generating quantitative perturbation data and mathematical modelling, we determined potential resistance mechanisms.

An Introduction to Advanced Targeted Acquisition Methods.

Targeted proteomics via selected reaction monitoring (SRM) or parallel reaction monitoring (PRM) enables fast and sensitive detection of a preselected set of target peptides. However, the number of peptides that can be monitored in conventional targeting methods is usually rather small. Recently, a series of methods has been described that employ intelligent acquisition strategies to increase the efficiency of mass spectrometers to detect target peptides.