Emergence of the B.1.214.2 SARS-CoV-2 lineage with an Omicron-like spike insertion and a unique upper airway immune signature.

We investigate the emergence, mutation profile, and dissemination of SARS-CoV-2 lineage B.1.214.2, first identified in Belgium in January 2021. This variant, featuring a 3-amino acid insertion in the spike protein similar to the Omicron variant, was speculated to enhance transmissibility or immune evasion. Initially detected in international travelers, it substantially transmitted in Central Africa, Belgium, Switzerland, and France, peaking in April 2021. Our travel-aware phylogeographic analysis, incorporating travel history, estimated the origin to the Republic of the Congo, with primary European entry through France and Belgium, and multiple smaller introductions during the epidemic. We correlate its spread with human travel patterns and air passenger data. Further, upon reviewing national reports of SARS-CoV-2 outbreaks in Belgian nursing homes, we found this strain caused moderately severe outcomes (8.7% case fatality ratio). A distinct nasopharyngeal immune response was observed in elderly patients, characterized by 80% unique signatures, higher B- and T-cell activation, increased type I IFN signaling, and reduced NK, Th17, and complement system activation, compared to similar outbreaks. This unique immune response may explain the variant's epidemiological behavior and underscores the need for nasal vaccine strategies against emerging variants.

A core proteome profile unites mouse models and patients in Alzheimer disease.

Levites et al. demonstrate that mouse models of Alzheimer disease (AD), exhibiting amyloid-beta (Αß) plaque formation, share Αß responsome proteins with humans. Their work underscores the value of these models in studying Αß aggregation, cellular vulnerability, and early-stage AD pathology.

An end-to-end approach for single-cell infrared absorption spectroscopy of bacterial inclusion bodies: from AFM-IR measurement to data interpretation of large sample sets.

BACKGROUND: Inclusion bodies (IBs) are well-known subcellular structures in bacteria where protein aggregates are collected. Various methods have probed their structure, but single-cell spectroscopy remains challenging. Atomic Force Microscopy-based Infrared Spectroscopy (AFM-IR) is a novel technology with high potential for the characterisation of biomaterials such as IBs. RESULTS: We present a detailed investigation using AFM-IR, revealing the substructure of IBs and their variation at the single-cell level, including a rigorous optimisation of data collection parameters and addressing issues such as laser power, pulse frequency, and sample drift. An analysis pipeline was developed tailored to AFM-IR image data, allowing high-throughput, label-free imaging of more than 3500 IBs in 12,000 bacterial cells. We examined IBs generated in Escherichia coli under different stress conditions. Dimensionality reduction analysis of the resulting spectra suggested distinct clustering of stress conditions, aligning with the nature and severity of the applied stresses. Correlation analyses revealed intricate relationships between the physical and morphological properties of IBs. CONCLUSIONS: Our study highlights the power and limitations of AFM-IR, revealing structural heterogeneity within and between IBs. We show that it is possible to perform quantitative analyses of AFM-IR maps over a large collection of different samples and determine how to control for various technical artefacts.

Central nervous system tumours and occupational ionising radiation exposure: a nested case-control study among the ORICAMs cohort of healthcare workers in France.

OBJECTIVE: This study aimed at investigating the relationship between occupational exposure to external ionising radiation and central nervous system (CNS) tumours mortality in healthcare workers working in France. DESIGN AND SETTING: The Occupational Radiation-Induced Cancer in Medical staff (ORICAMs) nested case-control study was conducted based on the dosimetric records of the national register of occupational dosimetry (Système d'information de la surveillance de l'exposition aux rayonnements ionisants). PARTICIPANTS AND METHODS: 33 CNS tumour deaths occurred between 2002 and 2012 among the ORICAMs cohort composed of 164 015 healthcare workers. Each case was matched to five controls alive at the time of the corresponding case's death, based on sex, year of birth, date of enrolment in the cohort and duration of follow-up. All participants were badge monitored for external radiation exposure, expressed in Hp(10). Conditional logistic regression was used to analyse the dose-response relationship between radiation dose and CNS mortality. RESULTS: Cases were exposed to a mean cumulative career radiation dose of 5.8±13.7 (max: 54.3) millisievert (mSv) compared with 4.1±15.2 (142.2) mSv for controls. No statistically significant association was found between CNS tumour mortality and cumulative whole-body career dose (OR=1.00, 95% CI 0.98 to 1.03), duration of exposure (OR=1.03; 95% CI 0.95 to 1.12) or age at first exposure (OR=0.98; 95% CI 0.91 to 1.06). CONCLUSION: We found no evidence of an association between external radiation exposure and CNS tumour risk in healthcare workers. Limitations of the study include low statistical power and short duration of follow-up.

Pharmacological modulation of septins restores calcium homeostasis and is neuroprotective in models of Alzheimer's disease.

Abnormal calcium signaling is a central pathological component of Alzheimer's disease (AD). Here, we describe the identification of a class of compounds called ReS19-T, which are able to restore calcium homeostasis in cell-based models of tau pathology. Aberrant tau accumulation leads to uncontrolled activation of store-operated calcium channels (SOCCs) by remodeling septin filaments at the cell cortex. Binding of ReS19-T to septins restores filament assembly in the disease state and restrains calcium entry through SOCCs. In amyloid-ß and tau-driven mouse models of disease, ReS19-T agents restored synaptic plasticity, normalized brain network activity, and attenuated the development of both amyloid-ß and tau pathology. Our findings identify the septin cytoskeleton as a potential therapeutic target for the development of disease-modifying AD treatments.

Mechanistic insights into the adaptive evolvability of spore heat resistance in Bacillus cereus sensu lato.

Wet heat treatment is a commonly applied method in the food and medical industries for the inactivation of microorganisms, and bacterial spores in particular. While many studies have delved into the mechanisms underlying wet heat killing and spore resistance, little attention has so far been dedicated to the capacity of spore-forming bacteria to tune their resistance through adaptive evolution. Nevertheless, a recent study from our group revealed that a psychrotrophic strain of the Bacillus cereus sensu lato group (i.e. Bacillus weihenstephanensis LMG 18989) could readily and reproducibly evolve to acquire enhanced spore wet heat resistance without compromising its vegetative cell growth ability at low temperatures. In the current study, we demonstrate that another B. cereus strain (i.e. the mesophilic B. cereus sensu stricto ATCC 14579) can acquire significantly increased spore wet heat resistance as well, and we subjected both the previously and currently obtained mutants to whole genome sequencing. This revealed that five out of six mutants were affected in genes encoding regulators of the spore coat and exosporium pathway (i.e. spoIVFB, sigK and gerE), with three of them being affected in gerE. A synthetically constructed ATCC 14579 ΔgerE mutant likewise yielded spores with increased wet heat resistance, and incurred a compromised spore coat and exosporium. Further investigation revealed significantly increased spore DPA levels and core dehydration as the likely causes for the observed enhanced spore wet heat resistance. Interestingly, deletion of gerE in Bacillus subtilis 168 did not impose increased spore wet heat resistance, underscoring potentially different adaptive evolutionary paths in B. cereus and B. subtilis.

CORDAX web server: an online platform for the prediction and 3D visualization of aggregation motifs in protein sequences.

MOTIVATION: Proteins, the molecular workhorses of biological systems, execute a multitude of critical functions dictated by their precise three-dimensional structures. In a complex and dynamic cellular environment, proteins can undergo misfolding, leading to the formation of aggregates that take up various forms, including amorphous and ordered aggregation in the shape of amyloid fibrils. This phenomenon is closely linked to a spectrum of widespread debilitating pathologies, such as Alzheimer's disease, Parkinson's disease, type-II diabetes, and several other proteinopathies, but also hampers the engineering of soluble agents, as in the case of antibody development. As such, the accurate prediction of aggregation propensity within protein sequences has become pivotal due to profound implications in understanding disease mechanisms, as well as in improving biotechnological and therapeutic applications. RESULTS: We previously developed Cordax, a structure-based predictor that utilizes logistic regression to detect aggregation motifs in protein sequences based on their structural complementarity to the amyloid cross-beta architecture. Here, we present a dedicated web server interface for Cordax. This online platform combines several features including detailed scoring of sequence aggregation propensity, as well as 3D visualization with several customization options for topology models of the structural cores formed by predicted aggregation motifs. In addition, information is provided on experimentally determined aggregation-prone regions that exhibit sequence similarity to predicted motifs, scores, and links to other predictor outputs, as well as simultaneous predictions of relevant sequence propensities, such as solubility, hydrophobicity, and secondary structure propensity. AVAILABILITY AND IMPLEMENTATION: The Cordax webserver is freely accessible at https://cordax.switchlab.org/.

Radiation protection in a cohort of healthcare workers: knowledge, attitude, practices, feelings and IR-exposure in French hospitals.

The number of healthcare workers occupationally exposed to ionizing radiation (IR) is increasing every year. As health effects from exposure to low doses IR have been reported, radiation protection (RP) in the context of occupational activities is a major concern. This study aims to assess the compliance of healthcare workers with RP policies, according to their registered cumulative dose, profession, and perception of radiation self-exposure and associated risk. Every healthcare worker from one of the participating hospitals in France with at least one dosimetric record for each year 2009, 2014, and 2019 in the SISERI registry was included and invited to complete an online questionnaire including information on the worker's occupational exposure, perception of IR-exposure risk and RP general knowledge. Hp(10) doses were provided by the SISERI system. Multivariate logistic regressions were used. Dosimeter wearing and RP practices compliance were strongly associated with 'feeling of being IR-exposed' (OR = 3.69, CI95% 2.04-6.66; OR = 4.60, CI95% 2.28-9.30, respectively). However, none of these factors was associated with RP training courses attendance. The main reason given for non-compliance is unsuitability or insufficient numbers of RP devices. This study provided useful information for RP policies. Making exposed workers aware of their own IR-exposure seems to be a key element to address in RP training courses. This type of questionnaire should be introduced into larger epidemiological studies. Dosimeter wearing and RP practices compliance are associated to feeling being IR-exposed. RP training courses should reinforce workers' awareness of their exposure to IR.

Integrating physics in deep learning algorithms: a force field as a PyTorch module.

MOTIVATION: Deep learning algorithms applied to structural biology often struggle to converge to meaningful solutions when limited data is available, since they are required to learn complex physical rules from examples. State-of-the-art force-fields, however, cannot interface with deep learning algorithms due to their implementation. RESULTS: We present MadraX, a forcefield implemented as a differentiable PyTorch module, able to interact with deep learning algorithms in an end-to-end fashion. AVAILABILITY AND IMPLEMENTATION: MadraX documentation, together with tutorials and installation guide, is available at madrax.readthedocs.io.

Local structural preferences in shaping tau amyloid polymorphism.

Tauopathies encompass a group of neurodegenerative disorders characterised by diverse tau amyloid fibril structures. The persistence of polymorphism across tauopathies suggests that distinct pathological conditions dictate the adopted polymorph for each disease. However, the extent to which intrinsic structural tendencies of tau amyloid cores contribute to fibril polymorphism remains uncertain. Using a combination of experimental approaches, we here identify a new amyloidogenic motif, PAM4 (Polymorphic Amyloid Motif of Repeat 4), as a significant contributor to tau polymorphism. Calculation of per-residue contributions to the stability of the fibril cores of different pathologic tau structures suggests that PAM4 plays a central role in preserving structural integrity across amyloid polymorphs. Consistent with this, cryo-EM structural analysis of fibrils formed from a synthetic PAM4 peptide shows that the sequence adopts alternative structures that closely correspond to distinct disease-associated tau strains. Furthermore, in-cell experiments revealed that PAM4 deletion hampers the cellular seeding efficiency of tau aggregates extracted from Alzheimer's disease, corticobasal degeneration, and progressive supranuclear palsy patients, underscoring PAM4's pivotal role in these tauopathies. Together, our results highlight the importance of the intrinsic structural propensity of amyloid core segments to determine the structure of tau in cells, and in propagating amyloid structures in disease.

N-glycosylation as a eukaryotic protective mechanism against protein aggregation.

The tendency for proteins to form aggregates is an inherent part of every proteome and arises from the self-assembly of short protein segments called aggregation-prone regions (APRs). While posttranslational modifications (PTMs) have been implicated in modulating protein aggregation, their direct role in APRs remains poorly understood. In this study, we used a combination of proteome-wide computational analyses and biophysical techniques to investigate the potential involvement of PTMs in aggregation regulation. Our findings reveal that while most PTM types are disfavored near APRs, N-glycosylation is enriched and evolutionarily selected, especially in proteins prone to misfolding. Experimentally, we show that N-glycosylation inhibits the aggregation of peptides in vitro through steric hindrance. Moreover, mining existing proteomics data, we find that the loss of N-glycans at the flanks of APRs leads to specific protein aggregation in Neuro2a cells. Our findings indicate that, among its many molecular functions, N-glycosylation directly prevents protein aggregation in higher eukaryotes.

LRRC37B is a human modifier of voltage-gated sodium channels and axon excitability in cortical neurons.

The enhanced cognitive abilities characterizing the human species result from specialized features of neurons and circuits. Here, we report that the hominid-specific gene LRRC37B encodes a receptor expressed in human cortical pyramidal neurons (CPNs) and selectively localized to the axon initial segment (AIS), the subcellular compartment triggering action potentials. Ectopic expression of LRRC37B in mouse CPNs in vivo leads to reduced intrinsic excitability, a distinctive feature of some classes of human CPNs. Molecularly, LRRC37B binds to the secreted ligand FGF13A and to the voltage-gated sodium channel (Nav) ß-subunit SCN1B. LRRC37B concentrates inhibitory effects of FGF13A on Nav channel function, thereby reducing excitability, specifically at the AIS level. Electrophysiological recordings in adult human cortical slices reveal lower neuronal excitability in human CPNs expressing LRRC37B. LRRC37B thus acts as a species-specific modifier of human neuron excitability, linking human genome and cell evolution, with important implications for human brain function and diseases.

Mechanisms and pathology of protein misfolding and aggregation.

Despite advances in machine learning-based protein structure prediction, we are still far from fully understanding how proteins fold into their native conformation. The conventional notion that polypeptides fold spontaneously to their biologically active states has gradually been replaced by our understanding that cellular protein folding often requires context-dependent guidance from molecular chaperones in order to avoid misfolding. Misfolded proteins can aggregate into larger structures, such as amyloid fibrils, which perpetuate the misfolding process, creating a self-reinforcing cascade. A surge in amyloid fibril structures has deepened our comprehension of how a single polypeptide sequence can exhibit multiple amyloid conformations, known as polymorphism. The assembly of these polymorphs is not a random process but is influenced by the specific conditions and tissues in which they originate. This observation suggests that, similar to the folding of native proteins, the kinetics of pathological amyloid assembly are modulated by interactions specific to cells and tissues. Here, we review the current understanding of how intrinsic protein conformational propensities are modulated by physiological and pathological interactions in the cell to shape protein misfolding and aggregation pathology.

TDP-43 pathology is associated with increased tau burdens and seeding.

BACKGROUND: Most Alzheimer's Disease (AD) cases also exhibit limbic predominant age-related TDP-43 encephalopathy neuropathological changes (LATE-NC), besides amyloid-ß plaques and neurofibrillary tangles (NFTs) containing hyperphosphorylated tau (p-tau). LATE-NC is characterized by cytoplasmic aggregates positive for pathological TDP-43 and is associated with more severe clinical outcomes in AD, compared to AD cases lacking TDP-43 pathology TDP-43: AD(LATE-NC-). Accumulating evidence suggests that TDP-43 and p-tau interact and exhibit pathological synergy during AD pathogenesis. However, it is not yet fully understood how the presence of TDP-43 affects p-tau aggregation in symptomatic AD. METHODS: In this study, we investigated the impact of TDP-43 proteinopathy on p-tau pathology with different approaches: histologically, in a human post-mortem cohort (n = 98), as well as functionally using a tau biosensor cell line and TDP-43A315T transgenic mice. RESULTS: We found that AD cases with comorbid LATE-NC, AD(LATE-NC+), have increased burdens of pretangles and/or NFTs as well as increased brain levels of p-tau199, compared to AD(LATE-NC-) cases and controls. The burden of TDP-43 pathology was also correlated with the Braak NFT stages. A tau biosensor cell line treated with sarkosyl-insoluble, brain-derived homogenates from AD(LATE-NC+) cases displayed exacerbated p-tau seeding, compared to control and AD(LATE-NC-)-treated cells. Consistently, TDP-43A315T mice injected with AD(LATE-NC+)-derived extracts also exhibited a more severe hippocampal seeding, compared to the remaining experimental groups, albeit no TDP-43 aggregation was observed. CONCLUSIONS: Our findings extend the current knowledge by supporting a functional synergy between TDP-43 and p-tau. We further demonstrate that TDP-43 pathology worsens p-tau aggregation in an indirect manner and increases its seeding potential, probably by increasing p-tau levels. This may ultimately contribute to tau-driven neurotoxicity and cell death. Because most AD cases present with comorbid LATE-NC, this study has an impact on the understanding of TDP-43 and tau pathogenesis in AD and LATE, which account for the majority of dementia cases worldwide. Moreover, it highlights the need for the development of a biomarker that detects TDP-43 during life, in order to properly stratify AD and LATE patients.

Exploiting the aggregation propensity of beta-lactamases to design inhibitors that induce enzyme misfolding.

There is an arms race between beta-lactam antibiotics development and co-evolving beta-lactamases, which provide resistance by breaking down beta-lactam rings. We have observed that certain beta-lactamases tend to aggregate, which persists throughout their evolution under the selective pressure of antibiotics on their active sites. Interestingly, we find that existing beta-lactamase active site inhibitors can act as molecular chaperones, promoting the proper folding of these resistance factors. Therefore, we have created Pept-Ins, synthetic peptides designed to exploit the structural weaknesses of beta-lactamases by causing them to misfold into intracellular inclusion bodies. This approach restores sensitivity to a wide range of beta-lactam antibiotics in resistant clinical isolates, including those with Extended Spectrum variants that pose significant challenges in medical practice. Our findings suggest that targeted aggregation of resistance factors could offer a strategy for identifying molecules that aid in addressing the global antibiotic resistance crisis.

SNPeffect 5.0: large-scale structural phenotyping of protein coding variants extracted from next-generation sequencing data using AlphaFold models.

BACKGROUND: Next-generation sequencing technologies yield large numbers of genetic alterations, of which a subset are missense variants that alter an amino acid in the protein product. These variants can have a potentially destabilizing effect leading to an increased risk of misfolding and aggregation. Multiple software tools exist to predict the effect of single-nucleotide variants on proteins, however, a pipeline integrating these tools while starting from an NGS data output list of variants is lacking. RESULTS: The previous version SNPeffect 4.0 (De Baets in Nucleic Acids Res 40(D1):D935-D939, 2011) provided an online database containing pre-calculated variant effects and low-throughput custom variant analysis. Here, we built an automated and parallelized pipeline that analyzes the impact of missense variants on the aggregation propensity and structural stability of proteins starting from the Variant Call Format as input. The pipeline incorporates the AlphaFold Protein Structure Database to achieve high coverage for structural stability analyses using the FoldX force field. The effect on aggregation-propensity is analyzed using the established predictors TANGO and WALTZ. The pipeline focuses solely on the human proteome and can be used to analyze proteome stability/damage in a given sample based on sequencing results. CONCLUSION: We provide a bioinformatics pipeline that allows structural phenotyping from sequencing data using established stability and aggregation predictors including FoldX, TANGO, and WALTZ; and structural proteome coverage provided by the AlphaFold database. The pipeline and installation guide are freely available for academic users on https://github.com/vibbits/snpeffect and requires a computer cluster.

Amyloid-like p53 as prognostic biomarker in serous ovarian cancer-a study of the OVCAD consortium.

TP53 is the most commonly mutated gene in cancer and has been shown to form amyloid-like aggregates, similar to key proteins in neurodegenerative diseases. Nonetheless, the clinical implications of p53 aggregation remain unclear. Here, we investigated the presence and clinical relevance of p53 aggregates in serous ovarian cancer (OC). Using the p53-Seprion-ELISA, p53 aggregates were detected in 46 out of 81 patients, with a detection rate of 84.3% in patients with missense mutations. High p53 aggregation was associated with prolonged progression-free survival. We found associations of overall survival with p53 aggregates, but they did not reach statistical significance. Interestingly, p53 aggregation was significantly associated with elevated levels of p53 autoantibodies and increased apoptosis, suggesting that high levels of p53 aggregates may trigger an immune response and/or exert a cytotoxic effect. To conclude, for the first time, we demonstrated that p53 aggregates are an independent prognostic marker in serous OC. P53-targeted therapies based on the amount of these aggregates may improve the patient's prognosis.

Sequence-targeted Peptides Divert Functional Bacterial Amyloid Towards Destabilized Aggregates and Reduce Biofilm Formation.

Functional bacterial amyloid provides structural stability in biofilm, making it a promising target for anti-biofilm therapeutics. Fibrils formed by CsgA, the major amyloid component in E. coli are extremely robust and can withstand very harsh conditions. Like other functional amyloids, CsgA contains relatively short aggregation-prone regions (APR) which drive amyloid formation. Here, we demonstrate the use of aggregation-modulating peptides to knock down CsgA protein into aggregates with low stability and altered morphology. Remarkably, these CsgA-peptides also modulate fibrillation of the unrelated functional amyloid protein FapC from Pseudomonas, possibly through recognition of FapC segments with structural and sequence similarity with CsgA. The peptides also reduce the level of biofilm formation in E. coli and P. aeruginosa, demonstrating the potential for selective amyloid targeting to combat bacterial biofilm.

Reduced Levels of Misfolded and Aggregated Mutant p53 by Proteostatic Activation.

In malignant cancer, excessive amounts of mutant p53 often lead to its aggregation, a feature that was recently identified as druggable. Here, we describe that induction of a heat shock-related stress response mediated by Foldlin, a small-molecule tool compound, reduces the protein levels of misfolded/aggregated mutant p53, while contact mutants or wild-type p53 remain largely unaffected. Foldlin also prevented the formation of stress-induced p53 nuclear inclusion bodies. Despite our inability to identify a specific molecular target, Foldlin also reduced protein levels of aggregating SOD1 variants. Finally, by screening a library of 778 FDA-approved compounds for their ability to reduce misfolded mutant p53, we identified the proteasome inhibitor Bortezomib with similar cellular effects as Foldlin. Overall, the induction of a cellular heat shock response seems to be an effective strategy to deal with pathological protein aggregation. It remains to be seen however, how this strategy can be translated to a clinical setting.