European Molecular Biology Laboratory

Hydrogen embrittlement is known to adversely affect the mechanical properties of low-carbon steels used for pipelines and pressure vessels, leading to accelerated crack growth and lowered fracture toughness. To overcome the limitations of surface-based analysis, this study employs X-ray micro-computed tomography (µ-CT) to provide a comprehensive 3D evaluation of the crack evolution. This approach is used to assess hydrogen-assisted crack growth in P355NH compact tension samples from previous fracture mechanical tests and enables a precise quantification of the internal crack path and the crack tip opening angle (CTOA) across the entire specimen thickness as well as the local damage morphology. By integrating these spatial parameters, a deeper understanding of the impact of hydrogen on local fracture mechanisms is achieved, revealing insights that have remained hidden in previous two-dimensional microscopy observations. For instance, µ-CT results clearly demonstrate that the hydrogen-assisted crack propagation is associated with increased void formation and secondary cracking in vicinity of the crack tip. However, it is proposed that the results are superimposed with continuous hydrogen desorption, which implies a need for in situ charging during mechanical loading and an analysis of the hydrogen concentration profile. Both will be the scope of further studies.

Dormancy is a survival strategy employed by all domains of life to withstand prolonged nutrient deprivation and environmental stress that is marked by a global shutdown of protein synthesis. However, the molecular mechanisms driving ribosome inactivation and reactivation during and after dormancy in eukaryotes remain poorly understood. Here, we identify SNOR, a novel SBDS-like ribosome-associated factor in Schizosaccharomyces pombe, that is upregulated and associates with ribosomes during induced dormancy triggered by glucose depletion. SNOR contributes to protein synthesis repression by binding the ribosome to probe the peptidyl transferase center (PTC), block tRNA-binding sites, and cap the polypeptide exit tunnel (PET). Importantly, we show that SNOR is essential for the restart of protein synthesis upon glucose reintroduction and exit from dormancy. SNOR is evolutionarily conserved and specifically upregulated in response to glucose stress in fungi. These findings reveal a previously unrecognized ribosome-associated factor that links glucose stress and cellular dormancy to surveillance of protein synthesis and highlight the power of in situ structural biology to uncover stress-responsive regulators of translation.

RNA structure starts forming cotranscriptionally as the nascent RNA emerges from the RNA polymerase and is dynamically modulated by cellular factors. How individual RNA conformations, out of an ensemble of RNA molecules, relate to function is not well understood. Here, developing multicolor single-molecule fluorescence microscopy experiments, we track in real time nascent RNA structure formation, functionally characterizing up to eight different types of RNA molecules. We find that ribosomal proteins, RNA modification enzymes or antisense oligonucleotides specifically modulate a subset of the RNA folding classes. For example, we provide direct evidence that increased local RNA accessibility at specific sites correlates with the chaperoning activity of ribosomal proteins during ribosome assembly. These experiments provide a general framework to study how dynamic RNA folding, and misfolding, relates to function.

Access to cross-border clinical trials may represent the sole therapeutic option for children living with rare diseases for which no approved medicines exist. Many children are excluded from participation in trials due to language restrictions. There are insufficient comprehensive analyses of the experiences and preferences of parents across Europe concerning participation and exclusion of their child in international clinical trials, particularly regarding language support during enrollment in cross-border clinical research studies. An anonymous online survey was designed and translated into 22 official European languages to collect data from parents of children living with a disease across Europe. The survey included five sections: (1) sociodemographic information; (2) experience participating in a clinical trial; (3) experience in cases where the patient was unable to take part in a study abroad; (4) experience participating in a clinical trial abroad; and (5) preferences regarding decentralized trial options. 1,436 responses were analyzed from parents across 34 European countries. 55.7% of the parents reported being able to communicate in English. 10.7% had prior clinical trial experience, of whom 30.1% traveled abroad to enable their child to participate. Among those reporting being excluded from cross-border trials, 34.7% cited language barriers or country of residence as the reason. Most families expressed a strong willingness to accept decentralized trial options, regardless of where the study may be conducted. Accommodating language translation to permit participation in a clinical trial abroad is feasible. While a significant percentage of caregivers of pediatric patients in Europe could communicate in English, approximately one-third of those excluded from clinical trials cited language barriers or country of residence as the reason. When translation was required, the most commonly offered solution was the use of professional interpreters, an accommodation that could enable broader patient participation in essential research.

Highly pathogenic avian influenza viruses (HPAIVs) derive from H5 and H7 low pathogenic avian influenza viruses (LPAIVs). Although insertion of a furin-cleavable multibasic cleavage site (MBCS) in the hemagglutinin gene was identified decades ago as the genetic basis for the LPAIV-to-HPAIV transition, the mechanisms underlying the occurrence of insertion are unknown. Here, we show that transient H5 RNA structures, predicted to trap the influenza virus polymerase on purine-rich sequences, drive nucleotide insertions, providing empirical evidence of RNA structure involvement in MBCS acquisition. Introduction of H5-like sequences and structures into an H6 hemagglutinin resulted in MBCS-yielding insertions. Our results show that nucleotide insertions that underlie H5 HPAIV emergence result from an RNA structure-driven diversity-generating mechanism, which could also occur in other RNA viruses.

Gold’s importance in various industries, the ever-rising demand for electronics, and shrinking gold reserves all underscore the pressing need to develop effective gold recovery methods from waste. In this study, we systematically evaluate several large-pore Zirconium-carboxylate Metal–Organic Frameworks (Zr-MOFs) having slightly varied internal surface functionality (-H, -CH3, -NH2, and -NO2) in gold recovery from acidic, simulated e-waste solutions. We compare two Zr-MOFs constructed from oligophenylene ligands with four MOFs built from structurally analogous piperazine-based ligands, including two unreported frameworks. The piperazine-based frameworks offer a critical sustainability advantage: by replacing the central benzene core with piperazine, they eliminate the need for expensive Pd-catalyzed coupling chemistry required for oligophenylene synthesis. In this work, we demonstrate for the first time that Zr-piperazine MOFs exhibit stability in acidic conditions, significantly surpassing their oligophenylene-based counterparts. One developed Zr-piperazine MOF, functionalized with both amine and methyl groups, substantially outperforms all analogs, displaying the highest stability alongside a gold adsorption capacity of up to 1826 mg/g and rapid adsorption kinetics in a 40 ppm gold solution. X-ray Absorption Near-Edge Structure (XANES) studies reveal distinct gold adsorption mechanisms for primary versus tertiary amines. These findings establish piperazine-based building blocks as promising candidates for designing selective gold adsorbents for e-waste recovery applications.

Prostate cancer (PCa) is a heterogeneous disease, impeding early detection and risk stratification. Liquid biopsies (LBx) enable minimally invasive tumor profiling, but circulating tumor-derived DNA (ctDNA) detection remains difficult, particularly in early-stage PCa. We developed a multimodal LBx approach combining genomic and epigenomic cell-free DNA (cfDNA) features in plasma and urine from newly diagnosed PCa patients to improve early characterization of PCa and risk stratification of aggressive disease. Plasma and urine samples from 55 localized PCa (lPCa) patients, 18 advanced PCa (aPCa) patients, and 36 cancer-free controls were subjected to low-coverage whole-genome sequencing and methylated DNA immunoprecipitation sequencing to assess fragmentation, chromosomal instability, and methylation in cfDNA. This complementary approach yielded a 45% ctDNA detection rate in newly diagnosed PCa. Major differences were observed between aPCa and controls, reflecting increasing signals with tumor progression. Epigenomic cfDNA features differentiated lPCa from aPCa, and ctDNA was detected in 46% of PCa patients with prostate-specific antigen <10 ng/mL, suggesting potential for risk stratification. This study highlights the value of multimodal LBx approaches for early characterization of primary PCa and identification of aggressive disease at initial diagnosis. Integration into clinical workflows could complement diagnostics and support personalized decision-making tailored to patients’ PCa risk profiles.

In plants, differentiation and expansion of some cell types is accompanied by exit from cell division and onset of endoreplication. Here, we describe a protocol to study changes in mRNA levels during endoreplication in Arabidopsis thaliana cotyledons. We provide detailed steps for the isolation of total nuclei and a strategy to sort stained nuclei according to their ploidy level. Finally, we detail procedures for RNA extraction, a housekeeping gene, and a marker gene for qPCR across ploidy.

Lipid transfer proteins (LTPs) maintain the specialized lipid compositions of organellar membranes1,2. In humans, many LTPs are implicated in diseases3, but the cargo and auxiliary lipids that facilitate the transfer of the majority of LTPs remain unknown. Here we combined biochemical, lipidomic and computational methods to systematically characterize LTP-lipid complexes4 and measure how LTP gains of function affect cellular lipidomes. We identified bound lipids for around half of the hundreds of LTPs that we analysed, confirming known ligands and identifying new ones across most LTP families. Gains in LTP function affected the cellular abundance of both their known and newly identified lipid ligands, indicating comparable functional relevance of the two ligand sets. Using structural bioinformatics, we characterized mechanisms that contribute to lipid selectivity and identified preferences based on headgroup or acyl chain. We demonstrate some basic principles of how LTPs mobilize their ligands. They commonly interact with several classes of lipids and exhibit broad but selective preference for particular headgroups and for lipid species with shorter acyl chains that contain one or two unsaturated carbons, suggesting that only subsets of lipid species are efficiently mobilized. The datasets represent a resource for further analysis in different cell types and states, such as those associated with pathologies.

Gut microbiota may contribute to the adiposity-associated disease risk, but human studies reported inconsistent associations of adiposity with gut microbiota composition. We examined associations of body mass index (BMI) with alpha diversity and relative microbial abundance at the phylum and genus taxonomic levels (based on 16S rRNA amplicon sequencing or metagenomics) among 7415 adults from eight European observational studies in a joint federated analysis of harmonized data using DataSHIELD. Higher BMI (per 5 kg/m2) was associated with lower alpha diversity (β: -0.05; 95% CI: -0.07, -0.03) and, on the phylum level, positively associated with Proteobacteria, but neither with Firmicutes nor Bacteroidetes nor their ratio, where high between-study heterogeneity was observed. On the genus level, BMI was inversely associated with the relative abundance of Faecalibacterium of the Firmicutes phylum (β: -0.11; 95% CI: -0.14, -0.07) but positively with the odds of detection of Dorea, Streptococcus, and Clostridium (all three Firmicutes) as well as Collinsella (Actinobacteria). This federated analysis of multiple studies found lower alpha diversity, alongside depleted Faecalibacterium, as well as higher odds of detection of Dorea, Streptococcus, Clostridium, and Collinsella with higher adiposity. By combining data from diverse study populations using harmonized data and statistical methods, our analysis partly overcomes sources of heterogeneity that may explain previously observed inconsistencies.