bluuebunny/crossref_metadata_2025 huggingface dataset:
Hedgehog signaling controls cytotoxic T cell migration in the tumour microenvironment
Authors: Kapeni Chrysa , O’Brien Louise , Sabirova Dilyara , Cast Oliver , Carbonaro Valentina , Clark-Leonard Stephen , Beke Flavio , McDonald Sarah , Fife Kate , de la Roche Maike
Published: August 2024

Cytotoxic T lymphocytes effectively eliminate cancer cells. Their abundance in the tumour microenvironment is one of the strongest pan-cancer predictors of clinical response. Here, we show that Hedgehog (Hh) signaling regulates T cell migration into tumours.

Using conditional knockout mouse models of central Hh signaling componentsIhh,SmoandGli1in CD8 T cells, we show thatSmodeletion greatly impairs the anti-tumour responsein vivodue to diminished CD8 T cell migration into the tumour microenvironment. The migration defect is mediated exclusively by Smo, both inin vivocancer models andin vitromigration assays. This effect is independent of the canonical Hh pathway and relies on the GPCR function of Smo to regulate the migration of murine and human CD8 T cells via RhoA.

Hh signaling is critical during embryonic development and adult stem cell homeostasis, but is also amplified in multiple cancer types. Hh inhibitors targeting SMO have been clinically-approved and shown efficacy in the treatment of Hh-driven basal cell carcinoma and medulloblastoma but have failed in clinical trials in other solid cancers with upregulated Hh signaling. We demonstrate that SMO inhibitors specifically decrease CD8 T cell migration into the tumour microenvironment, both in murine cancer models and resected BCCs from patients treated with the SMO inhibitor vismodegib, providing the first mechanistic explanation as to why Hh inhibitors have failed in solid cancers.

Our data establishes a novel link between Hh inhibitionin vivoand the anti-tumour immune response and reveals a fundamental mechanism controlling T cell migration. The work provides the basis for improved Hh targeting approaches in the clinic and new entry points into enhancing migration in T cell therapies.
Iron chelation by oral deferoxamine treatment decreased brain iron and iron signaling proteins
Authors: Thorwald Max A. , Godoy-Lugo Jose A. , Santa Maria Naomi , Chakhoyan Ararat , O'Day Peggy , Jacobs Russell , Zlokovic Berislav , Finch Caleb
Published: August 2024

Background: Deferoxamine (DFO) and other iron chelators are clinically used for cancer and stroke. They may also be useful for Alzheimers disease (AD) to diminish iron from microbleeds. DFO may also stimulate antioxidant membrane repair which is impaired during AD. DFO, and other chelators do enter the brain despite some contrary reports. Objective: Low dose, oral DFO was given in lab chow to wildtype (WT) C57BL/6 mice to evaluate potential impact on iron levels, iron-signaling and storage proteins, and amyloid precursor protein (APP) and processing enzymes. Young WT mice do not have microbleeds or disrupted blood-brain barrier of AD mice. Methods: Iron was measured by MRI and chemically after two weeks of dietary DFO. Cerebral cortex was examined for changes in iron metabolism, antioxidant signaling, and APP processing by Western blot. Results: DFO decreased brain iron by 18% (MRI) and decreased seven major proteins that mediate iron metabolism by at least 25%. The iron storage proteins ferritin light and heavy chain decreased by at least 30%. APP and secretase enzymes also decreased by 30%. Conclusions: WT mice respond to DFO with decreased APP, amyloid processing enzymes, and antioxidant repair. Potential DFO treatment for early-stage AD by DFO should consider the benefits of lowered APP and secretase enzymes.
Conservative taxonomy and quality assessment of giant virus genomes with GVClass
Authors: Pitot Thomas M , Brůna Tomàš , Schulz Frederik
Published: August 2024

Background

Large double-stranded DNA viruses of the phylum Nucleocytoviricota (Giant viruses; GVs) include the largest known viruses, both in terms of capsid and genome size and are associated with a wide range of eukaryotic hosts. The ones able to infect protists and algae have been shown to be the dominant orders of GVs in the environmental samples. These viruses encode for genes that may have significantly impacted biogeochemical cycling and host genome evolution. While GVs are frequently found in environmental sequence data, their large and complex genomes, composed of genes acquired from various cellular lineages, pose challenges for their identification and taxonomic classification.

Results

We present GVClass, a tool that identifies giant viruses in sequence data and provides taxonomic assignments, and estimates for genome completeness and contamination. GVClass performs gene calling optimized for giant viruses and utilizes a conservative approach based on consensus single protein phylogenies for robust taxonomic assignments. The genes used for classification represent highly conserved giant virus orthologous groups and low copy number cellular and viral panorthologs. In our benchmarking, GVClass demonstrated high quality and accurate taxonomic assignment of giant virus sequences. GVClass showed high to very high precision, with over 90% of tested instances correctly predicted at the genus level and near-perfect prediction (>99%) at higher taxonomic ranks (family, order, class).

Conclusion

In the light of rapidly increasing amounts of sequence data and associated metagenome-assembled genomes, GVClass provides a conservative approach to identify, classify and quality-check giant virus genomes, which with other methods often remained unassigned or misclassified using other methods. GVClass has already been used through viral meta-analysis and to benchmark the viral sequences detection pipeline geNomad. The standalone version is freely available and it has been integrated in the Integrated Microbial Genomes / Virus database (IMG/VR), offering the opportunity to upload user data for giant virus classification.
Molecular simulations of enzymatic phosphorylation of disordered proteins and their condensates
Authors: Zippo Emanuele , Dormann Dorothee , Speck Thomas , Stelzl Lukas
Published: August 2024

Understanding the condensation and aggregation of intrinsically disordered proteins in a non-equilibrium environment is crucial for unraveling many biological processes. Active enzymes catalyse many processes by consuming chemical fuels such as ATP. Enzymes called kinases phosphorylate disordered regions of proteins and thus profoundly affect their properties and interactions. Protein phosphorylation is implicated in neurodegenerative diseases and may modulate pathogenesis. However, how protein sequence and molecular recognition of a disordered protein by kinases determine phosphorylation patterns is not understood. In principle, molecular dynamics simulations hold the promise to resolve how phosphorylation affects disordered proteins and their assemblies. In practice, chemically-detailed simulations of enzymatic reactions and the dynamics of enzymes are highly challenging, in particular it is difficult to verify whether implementations of driven simulations are thermodynamically consistent. We can now address this problem with residue-level coarse-grained molecular dynamics simulations, integrating Metropolis Monte Carlo steps to model chemical reactions. Importantly, we show how to verify by Markov-state modeling that the realisation of a non-equilibrium steady state satisfies local-detailed balance. We investigate TDP-43 phosphorylation by the kinase CK1δ in simulations, examining patterns of phosphorylation and assessing its preventive role in chain aggregation, which may be a cytoprotective mechanism in neurodegenerative diseases. We find that the degree of residue phosphorylation is determined by sequence preference and charges, rather than by the position in the chain. The phosphorylation frequency is also affected by the phosphorylation patterns, since the interactions between CK1δ and TDP-43 actively change after each reaction. For TDP-43, our simulations show condensates dissolution through phosphorylation with kinases binding to the condensates and phosphorylating TDP-43 in the condensates.
Removal of NHS-labelling by-products in Proteomic Samples
Authors: Demyanenko Yana , Giltrap Andrew M. , Davis Benjamin G. , Mohammed Shabaz
Published: August 2024

N-Hydroxysuccinimide (NHS) ester chemistry is used extensively across proteomics sample preparation. One of its increasingly prevalent applications is in isobaric reagent-based quantitation such as the iTRAQ (isobaric tags for relative and absolute quantitation) and TMT (tandem mass tag) approaches. In these methods, labelling on the primary amines of lysine residues and N-termini of tryptic peptides via amide formation (N-derivatives) from corresponding NHS ester reagents is the intended reactive outcome. However, the role of NHS esters as activated carboxyls can also drive the formation of serine-, tyrosine-, and threonine- derived esters (O-derivatives). These O-derivative peptides are typically classed as over-labelled and are disregarded for quantification, leading to loss of information and hence potential sensitivity. Their presence also unnecessarily increases sample complexity, which reduces the overall identification rates. One common approach for removing these unwanted labelling events has involved a quench with hydroxylamine. We show here that this approach is not fully efficient and can still leave substantial levels of unwanted over-labelled peptides. Through systematic screening of nucleophilic aminolysis reagents and reaction conditions, we have now developed a robust method to efficiently remove over-labelled peptides. The new method reduces the proportion of over-labelled peptides in the sample to less than 1% without affecting the labelling rate or introducing other modifications, leading to superior identification rates and quantitation precision.
Blastocoel expansion and AMOT degradation cooperatively promote YAP nuclear localization during epiblast formation
Authors: Maeda Hinako , Sasaki Hiroshi
Published: August 2024

The epiblast is a pluripotent cell population formed in the late blastocyst stage of preimplantation embryos. During the process of epiblast formation from the inner cell mass (ICM) of the early blastocyst, activation of the Hippo pathway transcription factor TEAD by the nuclear translocation of the coactivator protein YAP is required for the robust expression of pluripotency factors. However, the mechanisms that alter YAP localization during epiblast formation remain unknown. Here, we reveal two such mechanisms. Expansion of the blastocoel promotes nuclear YAP localization by increasing cytoplasmic F-actin and reducing YAP phosphorylation. Additionally, cell differentiation regulates YAP. Expression of the junctional Hippo component, AMOT, gradually decreases during epiblast formation through a tankyrase-mediated degradation. SOX2 expression in the ICM is necessary for the reduction of AMOT and YAP phosphorylation. These two mechanisms function in parallel. Thus, the blastocoel-F-actin and SOX2-AMOT axes cooperatively suppress YAP phosphorylation and promote YAP nuclear localization during epiblast formation. The cooperation of these two distinct mechanisms likely contributes to the robustness of epiblast cell differentiation.
AVJ16 inhibits the RNA binding protein IGF2BP1 in lung adenocarcinomas and prevents tumor growth in mice
Authors: Wallis Nadav , Gershon Tehila , Shaaby Sojod , Oberman Froma , Grunewald Myriam , Duran Deborah , Singh Amandeep , Vainer Gilad , Spiegelman Vladimir S. , Sharma Arun K. , Yisraeli Joel K.
Published: August 2024

IGF2BP1 is an oncofoetal RNA binding protein that is expressed in many tumors. We have recently described a small molecule inhibitor of IGF2BP1, termed AVJ16, that prevents binding of the protein to its RNA targets by directly associating with the protein. Here, using a multi-omics approach, we have analyzed the effects of this inhibition on RNA binding, RNA expression, and protein expression. AVJ16 treatment downregulates RNAs encoding members of several pro-oncogenic signaling pathways, including Hedgehog, Wnt, and PI3K-Akt, and there is a strong correlation between IGF2BP1 RNA binding, RNA expression, and protein expression. AVJ16 treatment of lung adenocarcinoma (LUAD) cells in culture causes a strong reduction in proliferation, colony formation, invasion, and spheroid growth while enhancing apoptosis and cell death. All of these effects are limited to cells expressing IGF2BP1. LUAD cells treated with AVJ16 show a pronounced reduction in vital dye efflux, often correlated with enhanced chemosensitivity. In syngeneic LUAD xenografts in mice, IP injection of AVJ16 prevents tumor growth, and incubation with AVJ16 induces cell death in human organoids derived from IGF2BP1-expressing LUADs but not from healthy lung tissue. These results suggest that AVJ16 is a promising candidate for mono- and/or adjuvant therapy directed against tumors expressing IGF2BP1.
A genetically engineered therapeutic lectin inhibits human influenza A virus infection and sustains robust virus-specific CD8 T cell expansion
Authors: Yu Meng , Lin Ang , Baharom Faezzah , Li Shuijie , Legendre Maureen , Covés-Datson Evelyn , Sohlberg Ebba , Schlisio Susanne , Loré Karin , Markovitz David M. , Smed-Sörensen Anna
Published: August 2024

Native banana lectin (BanLec) is antiviral but highly mitogenic, which limits its therapeutic value. In contrast, the genetically engineered H84T BanLec (H84T) is not mitogenic but remains effective against influenza A virus (IAV) infection in mouse models. However, the potency and effect of H84T on human immune cells and IAV-specific immune responses is undetermined. We found that H84T efficiently inhibited IAV replication in human dendritic cells (DCs) from blood and tonsils, which preserved DC viability and allowed acquisition and presentation of viral antigen. Consequently, H84T-treated DCs initiated effective expansion of IAV-specific CD8 T cells. Furthermore, H84T preserved the capacity of IAV-exposed DCs to present a second non-IAV antigen and induce robust CD8 T cell expansion. This supports H84T as a potent antiviral in humans as it effectively inhibits IAV infection without disrupting DC function, and preserves induction of antigen-specific adaptive immune responses against diverse antigens, which likely is clinically beneficial.
Unlocking Protein Evolution Insights: Efficient and Interpretable Mutational Effect Predictions with GEMME
Authors: Carbone A. , Laine E. , Lombardi G.
Published: August 2024

The Global Epistatic Model for predicting Mutational Effects (GEMME) is a computational method that reconstructs protein mutational landscapes using sequence data alone. In this article, we delve into the broader biological questions that GEMME can address beyond mere landscape reconstruction. We provide several examples to guide users in maximizing the utility of GEMME. Additionally, we discuss recent advancements that enhance GEMME’s predictive accuracy and extend its capabilities by integrating sequence data with structural information and allele frequency data, when available. These innovations enable GEMME to answer new biological questions and offer deeper insights into protein evolution and function.
One pot RNA:DNA assembly for ribosomal RNA detection of pathogenic bacteria with single-molecule sensitivity
Authors: Boskovic Filip N , Sandler Sarah , Brauburger Simon , Shui Yuan , Kumar Bhavik , Pereira Dias Joana , Naydenova Plamena , Zhu Jinbo , Baker Stephen , Keyser Ulrich Felix
Published: August 2024

Effective treatment of infectious diseases hinges on timely and accurate diagnosis. Current methods face challenges in distinguishing between pathogens that cause a similar symptom and where appropriate treatment may be hindered by antimicrobial resistance (AMR). Here, we introduce a PCR-free system that identifies and quantifies ribosomal RNA transcripts (rRNA) in pathogenic bacteria instead of DNA-encoding rRNA genes. Our method leverages the potential of >1 million rRNAs found in replicating bacterial cells in comparison to the limited number of rRNA DNA copies per cell. We combined nanopores with RNA nanotechnology to identify rRNA from bacteria and other cells. We developed a simultaneous protocol for rRNA isolation, assembly and enrichment with assembled RNA identifiers (IDs), which prevents heat degradation and eliminates background RNA. Our method detects multiple bacterial species, including AMR variants of Salmonella enterica and Acinetobacter baumannii, and coinfection. This approach offers unmatched specificity, with the ability to identify a single bacterium without amplification. The integration of CRISPR-dCas9 binding to RNA IDs leads to serovar-specific identification and opens new avenues in identifying pathogens up to their variants using abundant rRNAs.
The evolution of signalling and monitoring in plant-fungal networks
Authors: Scott Thomas W. , Kiers E. Toby , West Stuart A.
Published: August 2024

Experiments have shown that when one plant is attacked by a pathogen or herbivore, this can lead to other plants connected to the same mycorrhizal network upregulating their defence mechanisms. It has been hypothesised that this represents signalling, with attacked plants producing a signal to warn other plants of impending harm. We examined the evolutionary plausibility of this and other hypotheses theoretically. We found that the evolution of plant signalling about an attack requires restrictive conditions, and so will rarely be evolutionarily stable. The problem is that signalling about an attack provides a benefit to competing neighbours, even if they are kin, and so reduces the relative fitness of signalling plants. Indeed, selection is often more likely to push plant behaviour in the opposite direction – with plants signalling dishonestly about an attack that has not occurred, or by suppressing a cue that they have been attacked. Instead, we show that there are two viable alternatives that could explain the empirical data: (1) the process of being attacked leads to a cue (information about the attack) which is too costly for the attacked plant to fully suppress; (2) mycorrhizal fungi monitor their host plants, detect when they are attacked, and then the fungi signal this information to warn other plants in their network. Our results suggest the empirical work that would be required to distinguish between these possibilities.

Significance statement

Experiments have shown that when one plant is attacked by a herbivore, this can lead to other plants connected to the same mycorrhizal network upregulating their defence mechanisms. It has been hypothesised that this represents signalling, with attacked plants producing a signal to warn other plants of impending harm. We found theoretically that plant warning signals are rarely evolutionarily stable. Instead, we identify two viable alternatives that could explain the empirical data: (1) being attacked leads to a cue (information about the attack) which is too costly for the attacked plant to suppress; (2) mycorrhizal fungi monitor their host plants, detect when they are attacked, and then the fungi signal this information to warn other plants in their network.
Fate specification triggers a positive feedback loop of TEAD-YAP and NANOG to promote epiblast formation in preimplantation embryos
Authors: Hirono Naoki , Hashimoto Masakazu , Maeda Hinako , Shimojo Hiromi , Sasaki Hiroshi
Published: August 2024

In preimplantation embryos, epiblast (EPI) fate specification from the inner cell mass is controlled by the segregation of NANOG and GATA6 expression. TEAD-YAP interaction is activated during EPI formation, and is required for pluripotency factor expression. These events occur asynchronously with similar timing during EPI formation, and their relationship remains elusive. Here, we examined the relationship between NANOG-GATA6 and TEAD-YAP. The nuclear accumulation of YAP takes place only in EPI-specified cells, and a positive feedback loop operates between NANOG and TEAD-YAP. The effects of TEAD-YAP on SOX2 upregulation in EPI-specified cells are likely indirect. EPI fate specification also alters the response of Nanog, Sox2 and Cdx2 to TEAD-YAP. These results suggest that EPI-fate specification alters the transcriptional network from the morula-like to the EPI-specified state and activates TEAD-YAP to trigger a positive feedback loop with NANOG, which stabilizes the EPI fate. The coordinated occurrence of these processes in individual cells likely supports proper EPI formation under the condition of asynchronous EPI-fate specification.
Formation of giant ER sheets by pentadecanoic acid causes lipotoxicity in fission yeast
Authors: Hoshikawa Yojiro , Shirota Natsuho , Tsugawa Hiroshi , Kimura Satoshi , Matsuyama Akihisa , Yashiroda Yoko , Kakeya Hideaki , Arita Makoto , Iizumi Reiko , Yoshida Minoru , Nishimura Shinichi
Published: August 2024

Excess amounts of saturated fatty acids are toxic to organisms, a condition termed lipotoxicity, which is often accompanied by pleiotropic cellular and tissue dysfunction. Here we show that pentadecanoic acid (C15:0) exerts toxicity on the fission yeastSchizosaccharomyces pombeby generating an aberrantly planar endoplasmic reticulum (ER) structure, which we named a “giant ER sheet.” Untargeted lipidomics revealed that C15:0 is incorporated into complex lipids depending on an acyl-CoA ligase Lcf1 and an acyl-CoA transferase Slc1, thereby increasing the saturation level of the acyl chains. The toxicity and giant ER sheet formation were abolished by deleting Lcf1 or Slc1, indicating that the incorporation of C15:0 into glycerophospholipids causes giant ER sheet formation. The giant ER sheets disrupted the correct migration of Mid1, a protein determining the cell division site, and physically blocked septum formation, hindering correct cell separation. Our results suggest that the ER is the primary site targeted by saturated fatty acids, leading to lipotoxicity.
Myosin forces elicit an F-actin structural landscape that mediates mechanosensitive protein recognition
Authors: Carl Ayala G. , Reynolds Matthew J. , Gurel Pinar S. , Phua Donovan Y.Z. , Sun Xiaoyu , Mei Lin , Hamilton Keith , Takagi Yasuharu , Noble Alex J. , Sellers James R. , Alushin Gregory M.
Published: August 2024

Cells mechanically interface with their surroundings through cytoskeleton-linked adhesions, allowing them to sense physical cues that instruct development and drive diseases such as cancer. Contractile forces generated by myosin motor proteins mediate these mechanical signal transduction processes through unclear protein structural mechanisms. Here, we show that myosin forces elicit structural changes in actin filaments (F-actin) that modulate binding by the mechanosensitive adhesion protein α-catenin. Using correlative cryo-fluorescence microscopy and cryo-electron tomography, we identify F-actin featuring domains of nanoscale oscillating curvature at cytoskeleton-adhesion interfaces enriched in zyxin, a marker of actin-myosin generated traction forces. We next introduce a reconstitution system for visualizing F-actin in the presence of myosin forces with cryo-electron microscopy, which reveals morphologically similar superhelical F-actin spirals. In simulations, transient forces mimicking tugging and release of filaments by motors produce spirals, supporting a mechanistic link to myosin’s ATPase mechanochemical cycle. Three-dimensional reconstruction of spirals uncovers extensive asymmetric remodeling of F-actin’s helical lattice. This is recognized by α-catenin, which cooperatively binds along individual strands, preferentially engaging interfaces featuring extended inter-subunit distances while simultaneously suppressing rotational deviations to regularize the lattice. Collectively, we find that myosin forces can deform F-actin, generating a conformational landscape that is detected and reciprocally modulated by a mechanosensitive protein, providing a direct structural glimpse at active force transduction through the cytoskeleton.
Integrative genomics approach identifies glial transcriptomic dysregulation and risk in the cortex of individuals with Alcohol Use Disorder
Authors: Warden Anna S. , Salem Nihal A. , Brenner Eric , Sutherland Greg T. , Stevens Julia , Kapoor Manav , Goate Alison M. , Dayne Mayfield R.
Published: August 2024

Alcohol use disorder (AUD) is a prevalent neuropsychiatric disorder that is a major global health concern, affecting millions of people worldwide. Past molecular studies of AUD used underpowered single cell analysis or bulk homogenates of postmortem brain tissue, which obscures gene expression changes in specific cell types. Here we performed single nuclei RNA-sequencing analysis of 73 post-mortem samples from individuals with AUD (N=36, Nnuclei= 248,873) and neurotypical controls (N=37, Nnuclei= 210,573) in both sexes across two institutional sites. We identified 32 clusters and found widespread cell type-specific transcriptomic changes across the cortex in AUD, particularly affecting glia. We found the greatest dysregulation in novel microglial and astrocytic subtypes that accounted for the majority of differential gene expression and co-expression modules linked to AUD. Analysis for cell type-specific enrichment of aggregate genetic risk for AUD identified subtypes of microglia and astrocytes as potential key players not only affected by but causally linked to the progression of AUD. These results highlight the importance of cell-type specific molecular changes in AUD and offer opportunities to identify novel targets for treatment.
Steering cell-state and phenotype transitions by causal disentanglement learning
Authors: Zhang Chengming , Chen Zexi , Miao Yuanxiang , Xue Yun , Cai Deyu , Guo Weifeng , Ji hongbin , Aihara Kazuyuki , Chen Luonan
Published: August 2024

Understanding and manipulating cell-state and phenotype transitions is essential for advancing biological research and therapeutic treatment. We introduce CauFinder, an advanced framework designed to accurately identify causal regulators of these transitions and further precisely steer such transitions by integrating causal disentanglement modelling with network control based solely on observed data. By leveraging do-calculus and optimizing information flow metrics, CauFinder can distinguish causal factors from spurious ones, ensuring precise control over desired state transitions. One significant advantage of CauFinder is its ability to identify those variables causally affecting the cell-state/phenotype transitions among all observed variables, both theoretically and computationally, leading to the identification of their master regulators when combined with network control. Consequently, by employing a counterfactual algorithm, CauFinder is able effectively to facilitate desirable state transitions or steer these transitional trajectories/paths by modulating these causal drivers. Beyond its theoretical advantages, CauFinder outperforms existing approaches computationally in both simulated and real-world settings. CauFinder is able to not only reveal natural biological transition processes such as (a) cell differentiation, (b) lung adenocarcinoma (LUAD) to lung squamous cell carcinoma (LUSC) transdifferentiation and (c) drug-sensitive to drug-resistant transitions but also identify the causal regulators of their reverse transition processes, such as (A) cell dedifferentiation, (B) LUSC to LUAD transdifferentiation and (C) drug-resistant to drug-sensitive transitions. These findings highlight its superior ability to causally uncover essential regulatory mechanisms and accurately steer cell-state/phenotype transitions, thus providing novel therapeutic strategies.
Interspecies differences in the transcriptome response of corals to acute heat stress
Authors: Da-Anoy Jeric , Posadas Nino , Conaco Cecilia
Published: August 2024

Rising sea surface temperatures threaten the survival of corals worldwide, with coral bleaching events becoming more commonplace. However, different coral species are known to exhibit variable levels of susceptibility to thermal stress events. To elucidate genetic mechanisms that may underlie these differences, we compared the gene complement of four coral species, Favites colemani, Montipora digitata, Acropora digitifera, and Seriatopora caliendrum, that were previously demonstrated to have differing responses to acute thermal stress. We found that more tolerant species, like F. colemani and M. digitata, possess a greater abundance of antioxidant protein families and chaperones. Under acute thermal stress conditions, only S. caliendrum showed a significant bleaching response, which was accompanied by activation of DNA damage response network and drastic upregulation of stress response genes (SRGs). This suggests that differences in SRG complement, as well as the mechanisms that control SRG expression response, contribute to the ability of corals to maintain stable physiological functions that is required to survive shifts in seawater temperature.
Multipartite complexity of the lichen symbiosis revealed by metagenome and transcriptome analysis ofXanthoria parietina
Authors: Tagirdzhanova Gulnara , Scharnagl Klara , Sahu Neha , Yan Xia , Bucknell Angus , Bentham Adam R. , Jégousse Clara , Ament-Velásquez Sandra Lorena , Onuț-Brännström Ioana , Johannesson Hanna , MacLean Dan , Talbot Nicholas J.
Published: August 2024

Lichens are composite symbiotic associations of fungi, algae, and bacteria that result in large, anatomically complex organisms adapted to many of the world’s most challenging environments. How such intricate, self-replicating lichen architectures develop from simple microbial components remains unknown because of their recalcitrance to experimental manipulation. Here we report a metagenomic and metatranscriptomic analysis of the lichenXanthoria parietinaat different developmental stages. We identified 168 genomes of symbionts and lichen-associated microbes within a lichen thallus, including representatives of green algae, three different classes of fungi, and 14 bacterial phyla. By analyzing occurrence of individual species across lichen thalli from diverse environments, we defined both substrate-specific and core microbial components of the lichen. Meta-transcriptomic analysis of the principal fungal symbiont from three different developmental stages of a lichen, compared to axenically grown fungus, revealed differential gene expression profiles indicative of lichen-specific transporter functions, specific cell signalling, transcriptional regulation and secondary metabolic capacity. Putative immunity-related proteins and lichen-specific structurally conserved secreted proteins resembling fungal pathogen effectors were also identified, consistent with a role for immunity modulation in lichen morphogenesis.
Transgenerational effects impact the vulnerability of a host-parasitoid system to rising temperatures.
Authors: Bright Natalie L. , Chen Jinlin , Terry Christopher
Published: August 2024

Transgenerational effects, where ancestral experience of environmental conditions influences the performance in subsequent generations, are hypothesised to have substantial consequences for responses to climate change. However, any apparent detriment or advantage these processes generate for a focal species may be counteracted by concurrent effects upon interacting species. Using an experimental Drosophila-parasitoid model system, we determined how the parental thermal environment impacts the performance of both hosts and parasitoids. We found substantial responses in both trophic-levels, with potential evidence for both condition-transfer effects and adaptive transgenerational plasticity. We used these results to parameterise discrete-time simulation models to explore how transgenerational effects of thermal conditions and temporal autocorrelation in temperature are expected to impact the time to extinction for this host-parasitoid system under climate change. The models predicted that transgenerational effects would significantly hasten the time to extinction, largely through a reduction in estimated average performance. Under a simple model, there was an additional hastening of extinction derived from the interaction between the coupled transgenerational effects, although no interactive effects were detectable in an alternative model with greater buffering capacity. Our research demonstrates a new mechanism underlying how tightly-coupled interacting species will respond to climate change and highlights the need to quantify and contextualise thermal transgenerational effects.
Ribosome-inactivation by a class of widely distributed C-tail anchored membrane proteins
Authors: Njenga Robert , Boele Julian , Drepper Friedel , Sinha Kasturica , Marouda Eirini , Huesgen Pitter F. , Blaby-Haas Crysten , Koch Hans-Georg
Published: August 2024

Ribosome hibernation is a commonly used strategy that protects ribosomes under unfavorable conditions and regulates developmental processes. Multiple ribosome-hibernation factors have been identified in all domains of life, but due to their structural diversity and the lack of a common inactivation mechanism, it is currently unknown how many different hibernation factors exist. Here, we show that the YqjD/ElaB/YgaM paralogs, initially discovered as membrane-bound ribosome binding proteins inE. coli,constitute an abundant class of ribosome-hibernating proteins, which are conserved across all proteobacteria and some other bacterial phyla. Our data demonstrate that they inhibitin vitroprotein synthesis by interacting with the 50S ribosomal subunit.In vivocross-linking combined with mass spectrometry reveals their specific interactions with proteins surrounding the ribosomal tunnel exit and even their penetration into the ribosomal tunnel. Thus, YqjD/ElaB/YgaM inhibit translation by blocking the ribosomal tunnel and thus mimic the activity of antimicrobial peptides and macrolide antibiotics.
Anchor Point Engineering on Anticalin Scaffolds for Enhanced Particle Adhesion to CTLA-4 Under Shear Stress
Authors: Sun Yang , Li Jiajun , Vanella Rosario , Liu Haipei , Liu Zhaowei , Nash Michael
Published: August 2024

Achieving high binding strength and efficient delivery of molecular cargo to tumor cells is a significant challenge in cancer immunotherapy. Current research has focused on obtaining antibodies that bind their targets with high affinity (i.e. low equilibrium dissociation constant, KD). However, such an approach neglects the influence of hydrodynamic fluid flow along the surfaces of cells and tissues, which can impart mechanical stress and lead to rapid unbinding of antibodies under flow. Here, we investigated how altering the anchor point within a non-antibody binding scaffold called Anticalin can enhance particle adhesion to the immune checkpoint CTLA-4 on human cells experiencing hydrodynamic shear. By introducing bioorthogonal clickable amino acids into Anticalin at various positions, we investigated the influence of different unbinding geometries on the stability of the Anticalin:(CTLA-4) complex. Both single-molecule AFM and quantitative bead-based adhesion assays on CTLA-4+human cells demonstrate how optimal choice of the anchor point can stabilize the complex and enhance the adhesion strength of receptor-targeted particles. These results demonstrate how anchor point engineering can enhance particle adhesion and delivery to immunotherapy targets.
Improved Zika virus plaque assay using Vero/TMPRSS2 cell line
Authors: Oluwasemowo Olukunle O. , Graham Monica E. , Murugesh Deepa K. , Borucki Monica K.
Published: August 2024

Plaque assay is the gold standard for the quantification of viable cytopathic viruses like Zika virus (ZIKV). Some strains of ZIKV produce plaques that are very difficult to accurately visualize and count on the commonly used Vero cell line. From data generated in our lab, we became curious if Vero/TMPRSS2 cells may be a better alternative, therefore we compared the plaque forming units (PFU) of two strains of ZIKV on Vero/TMPRSS2 cells to those produced by Vero cells. We also compared the virus stock titer generated on Vero/TMPRSS2 cells to that generated by the Vero cell line. Although Vero cells generated higher quantity of ZIKV stocks, Vero/TMPRSS2 cells produced plaques with significantly improved morphology and visibility and may therefore be a better alternative to use for performing plaque assays for strains of ZIKV that are more difficult to titer on regular Vero cells.
Crosslinked agarose-gelatine beads as a substrate for investigating biofilms of bacterial pathogens
Authors: Roizman Dan , Herzog Maren , Nath Arpita , Pachaimuthu Nivetha , Hujeirat Ahmad , Kuropka Benno , Rolff Jens , Rodríguez-Rojas Alexandro
Published: August 2024

Treating chronic bacterial infections is challenging due to the formation of biofilms, making bacteria less susceptible to antimicrobials.In vitromodels have limitations in replicating biofilm physiology. To address this problem, we have created a hydrogel substrate that combines crosslinked agarose and gelatine presented as beads, providing stability and resistance to autoclaving. Bacterial pathogens rapidly colonise these biogel beads when submerged in liquid culture. The substrate was tested withEscherichia coli,Pseudomonas aeruginosa, andStaphylococcus aureus, showing more robust biofilm growth than its glass bead counterpart. Additionally, this led to increased virulence factor production and served as a reservoir for biofilm quorum sensing molecules. These features closely resemble clinical situations, suggesting a more accurate representation of biofilm-associated infections than current approaches. This new substrate offers a practical and convenient model for studying biofilms of bacterial pathogens, providing an efficient solution to the research community and holding promise for future breakthroughs.
Challenges in detecting ecological interactions using sedimentary ancient DNA data
Authors: Callahan Fiona Margaret , Li Jacky Kaiyuan , Nielsen Rasmus
Published: August 2024

With increasing availability of ancient and modern environmental DNA technology, whole-community species occurrence and abundance data over time and space is becoming more available. Sedimentary ancient DNA data can be used to infer associations between species, which can generate hypotheses about biotic interactions, a key part of ecosystem function and biodiversity science. Here, we have developed a realistic simulation to evaluate five common methods from different fields for this type of inference. We find that across all methods tested, false discovery rates of inter-species associations are high under simulation conditions where the assumptions of the methods are violated in a variety of ecologically realistic ways. Additionally, we find that for more realistic simulation scenarios, with sample sizes that are currently realistic for this type of data models are typically unable to detect interactions better than random assignment of associations. Different methods perform differentially well depending on the number of taxa in the dataset. Some methods (SPIEC-EASI, SparCC) assume that there are large numbers of taxa in the dataset, and we find that SPIEC-EASI is highly sensitive to this assumption while SparCC is not. Additionally, we find that for many methods, default calibration can result in high false discovery rates. We find that for small numbers of species, no method consistently outperforms logistic and linear regression, indicating a need for further testing and methods development.
Human Replication Protein A complex is a Telomerase Processivity Factor Essential for Telomere Maintenance
Authors: Agrawal Sourav , Lin Xiuhua , Susvirkar Vivek , O'Connor Michael S , Chavez Bianca L , Tholkes Victoria R , Abe Kaitlyn M , He Qixiang , Huang Xuhui , Lim Ci Ji
Published: August 2024

Telomerase is crucial for maintaining telomere length and safeguarding genome stability. In this study, we identified Replication Protein A (RPA) as a novel telomerase processivity factor, functioning alongside the telomerase recruitment factor TPP1-POT1. AlphaFold2 predictions revealed that RPA and TPP1 interact with telomerase at distinct binding sites. Using separation-of-function mutants, we discovered that RPA-mediated telomerase stimulation is indispensable for telomere elongation, while TPP1-POT1 primarily functions in recruiting telomerase to telomeres. Furthermore, we demonstrated that short telomere disease-associated telomerase mutations compromise RPA's ability to stimulate telomerase, establishing a link between impaired RPA-dependent processivity and telomeropathies. Our findings redefine human telomerase regulation by establishing RPA as a critical regulator and provide new insights into the molecular basis of telomere-related diseases.
Antiviral Library Screening Leads to Discovery of Novel and Potent Anticlostridial Compounds
Authors: Stolz Brice Jasarino , Seleem Mohamed N. , Abouelkhair Ahmed A
Published: August 2024

Clostridium difficile(C. difficile) is a major cause of nosocomial infections, often associated with individuals who have gut dysbiosis from previous antibiotic therapies.C. difficileinfections have a high recurrence rate and impose significant financial and mortality burdens on the healthcare system. Novel anti-C. difficiledrugs are urgently needed to treat and reduce the severity and recurrence of infection. In this study, we screened a library of 618 antiviral drugs to identify a potential candidate for repurposing as novel anti-C. difficiletreatments. Following our preliminary screening, we identified 9 novel compounds that inhibitC. difficileat a concentration of 16 µM or lower. Among these, 4 antiviral compounds demonstrated the most potent anti-C. difficileactivity against a panel of 15C. difficileisolates, with minimum inhibitory concentrations (MICs) comparable to the drug of choice, vancomycin. These include rottlerin (MIC50 = 0.25 µg/mL), α-mangostin (MIC50 = 1 µg/mL), dryocrassin ABBA (MIC50 = 1 µg/mL), and obefazimod (MIC50 = 4 µg/mL).  All exhibited minimal to no activity against representative members of the gut microbiota. Interestingly, α-mangostin, a natural xanthone derived from the mangosteen fruit, exhibited strong bactericidal action, clearing a high inoculum ofC. difficilein less than an hour. All other drugs exhibited bacteriostatic activity. Given the characteristics of these compounds, they show great promise as novel treatments forC. difficileinfections.
A rapid growth rate underpins the dominance ofHanseniaspora uvarumin spontaneous grape juice fermentations
Authors: Onetto Cristobal A. , McCarthy Jane , Schmidt Simon A.
Published: August 2024

Hanseniaspora uvarumis consistently observed as the dominant non-Saccharomycesspecies in spontaneous grape juice fermentations. However, the physiological mechanisms and physicochemical variables influencing the prevalence ofH. uvarumover other non-Saccharomycesspecies remain unclear. We tested the physicochemical parameters contributing toH. uvarumdominance by inoculating a chemically diverse set of grape juices with a mock community whose composition was defined following a comprehensive microbial survey of spontaneous fermentations. Our findings indicated that the chemical composition of grape juice had minimal impact on the microbial dynamics of fermentation, withH. uvarumemerging as the dominant non-Saccharomycesspecies in nearly all conditions tested. Grape juice composition primarily influenced the total yeast abundance of the mock community. Flow cytometry analysis confirmed thatH. uvarumhas a faster growth rate thanSaccharomyces cerevisiaeand several otherHanseniaspora spp.. Moreover, its growth was not affected by the presence ofS. cerevisiae, explaining its rapid dominance in spontaneous fermentations. The rapid growth ofH. uvarumnegatively impacted the growth ofS. cerevisiae, with significant implications for fermentation performance and sugar consumption. The results of this study suggest that the fast growth rate ofH. uvarumenables it to quickly dominate the grape juice environment during the early stages of fermentation. This physiological advantage indicates that the initial abundance ofH. uvarummay be critical to the outcome of spontaneous fermentations, as evidenced by its direct impact on the growth ofS. cerevisiaeand fermentation performance.
Neuromuscular dysfunction in patient-derived FUSR244RR-ALS iPSC model via axonal downregulation of neuromuscular junction proteins
Authors: von Kuegelgen Nicolai , Ludwik Katarzyna , Mendonsa Samantha , Roemer Christine , Becher Erik , Breimann Laura , Strauch Mara , Mari Tommaso , Mongellaz Sandrine , Grexa Nina , Oliveras-Martinez Anna , Woehler Andrew , Selbach Matthias , la Bella Vincenzo , Ulitsky Igor , Chekulaeva Marina
Published: August 2024

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative condition characterized by the progressive degeneration of motor neurons, ultimately resulting in death due to respiratory failure. A common feature among ALS cases is the early loss of axons, pointing to defects in axonal transport and translation as initial disease indicators. Here, we established a FUS-ALS hiPSC-derived model that recapitulates the motor neuron survival and muscle contractility defects characteristic of ALS patients. Analysis of the protein and mRNA expression profiles in axonal and somatodendritic compartments of ALS-afflicted and isogenic control motor neurons revealed a selective downregulation of proteins essential for the neuromuscular junction function in FUS-ALS axons. Furthermore, analysis of FUS CLIP and RIP data showed that FUS binds mRNAs encoding these proteins. This work shed light on the pathogenic mechanisms of ALS and emphasized the importance of axonal gene expression analysis in elucidating the mechanisms of neurodegenerative disorders.
Clu1/Clu form mitochondria-associated granules upon metabolic transitions and regulate mitochondrial protein translation via ribosome interactions
Authors: Miller-Fleming Leonor , Au Wing Hei , Raik Laura , Guiomar Pedro , Schmitz Jasper , Cho Ha Yoon , Czako Aron , Whitworth Alexander J
Published: August 2024

Mitochondria perform essential metabolic functions and respond rapidly to changes in metabolic and stress conditions. As the majority of mitochondrial proteins are nuclear-encoded, intricate post-transcriptional regulation is crucial to enable mitochondria to adapt to changing cellular demands. The eukaryotic Clustered mitochondria protein family has emerged as an important regulator of mitochondrial function during metabolic shifts. Here, we show that theDrosophila melanogasterandSaccharomyces cerevisiaeClu/Clu1 proteins form dynamic, membraneless granules adjacent to mitochondria in response to metabolic changes. Yeast Clu1 regulates the translation of a subset of nuclear-encoded mitochondrial protein by interacting with their mRNAs while these are engaged in translation. We further show that Clu1 regulates translation by interacting with polysomes, independently of whether it is in a diffuse or granular state. Our results demonstrate remarkable functional conservation with other members of the Clustered mitochondria protein family and suggests that Clu/Clu1 granules isolate and concentrate ribosomes engaged in translating their mRNA targets, thus, integrating metabolic signals with the regulation of mitochondrial protein synthesis.