bluuebunny/crossref_metadata_2025 huggingface dataset:
Chromosome Structure II: Stem-loops and circle-loops
Authors: Ke Wenfan , Fujioka Miki , Schedl Paul , Jaynes James B
Published: January 2024

The chromosomes in multicellular eukaryotes are organized into a series of topologically independent loops called TADs. In flies, TADs are formed by physical interactions between neighboring boundaries. Fly boundaries exhibit distinct partner preferences, and pairing interactions between boundaries are typically orientation-dependent. Pairing can be head-to-tail or head-to-head. The former generates a stem-loop TAD, while the latter gives a circle-loop TAD. The TAD that encompasses the Drosophila even skipped (eve) gene is formed by the head-to-tail pairing of the nhomie and homie boundaries. To explore the relationship between loop topology and the physical and regulatory landscape, we flanked the nhomie boundary region with two attP sites. The attP sites were then used to generate four boundary replacements: lambda DNA, nhomie forward (WT orientation), nhomie reverse (opposite of WT orientation), and homie forward (same orientation as WT homie). The nhomie forward replacement restores the WT physical and regulatory landscape: In MicroC experiments, the eve TAD is a volcano triangle topped by a plume, and the eve gene and its regulatory elements are sequestered from interactions with neighbors. The lambda DNA replacement lacks boundary function: the endpoint of the new eve TAD on the nhomie side is ill-defined, and eve stripe enhancers activate a nearby gene, eIF3j. While nhomie reverse and homie forward restore the eve TAD, the topology is a circle-loop, and this changes the local physical and regulatory landscape. In MicroC experiments, the eve TAD interacts with its neighbors, and the plume at the top of the eve triangle peak is instead flanked by a pair of clouds of contacts with the next-door TADs. Consistent with the loss of isolation afforded by the stem-loop topology, the eve enhancers weakly activate genes in the neighboring TADs. Conversely, eve function is partially disrupted.
Shaping the physical world to our ends: The left PF technical-cognition area
Authors: Osiurak François , Federico Giovanni , Fournel Arnaud , Gaujoux Vivien , Lamberton Franck , Ibarrola Danielle , Rossetti Yves , Lesourd Mathieu
Published: January 2024

Our propensity to materiality, which consists in using, making, creating, and passing on technologies, has enabled us to shape the physical world according to our ends. To explain this proclivity, scientists have calibrated their lens to either low-level skills such as motor cognition or high-level skills such as language or social cognition. Yet, little has been said about the intermediate-level cognitive processes that are directly involved in mastering this materiality, that is, technical cognition. We aim to focus on this intermediate level for providing new insights into the neurocognitive bases of human materiality. Here we show that a technical-reasoning process might be specifically at work in physical problem-solving situations. We found via two distinct neuroimaging studies that the area PF (parietal F) within the left parietal lobe is central for this reasoning process in both tool-use and non-tool-use physical problem-solving and can work along with social-cognitive skills to resolve day-to-day interactions that combine social and physical constraints. Our results demonstrate the existence of a specific cognitive module in the human brain dedicated to materiality, which might be the supporting pillar allowing the accumulation of technical knowledge over generations. Intensifying research on technical cognition could nurture a comprehensive framework that has been missing in fields interested in how early and modern humans have been interacting with the physical world through technology, and how this interaction has shaped our history and culture.
Longitudinal transcriptional changes reveal genes from the natural killer cell-mediated cytotoxicity pathway as critical players underlying COVID-19 progression
Authors: Medina Matias A. , Fuentes-Villalobos Francisco , Quevedo Claudio , Aguilera Felipe , Riquelme Raul , Rioseco Maria Luisa , Barria Sebastian , Pinos Yazmin , Calvo Mario , Burbulis Ian , Kossack Camila , Alvarez Raymond A ,   COVID 19 South Chile Group, Garrido Jose Luis , Barria Maria Ines
Published: January 2024

Patients present a wide range of clinical severities in response SARS-CoV-2 infection, but the underlying molecular and cellular reasons why clinical outcomes vary so greatly within the population remains unknown. Here, we report that negative clinical outcomes in severely ill patients were associated with divergent RNA transcriptome profiles in peripheral immune cells compared with mild cases during the first weeks after disease onset. Protein-protein interaction analysis indicated that early-responding cytotoxic NK cells were associated with an effective clearance of the virus and a less severe outcome. This innate immune response was associated with the activation of select cytokine-cytokine receptor pathways and robust Th1/Th2 cell differentiation profiles. In contrast, severely ill patients exhibited a dysregulation between innate and adaptive responses affiliated with divergent Th1/Th2 profiles and negative outcomes. This knowledge forms the basis of clinical triage that may be used to preemptively detect high-risk patients before life-threatening outcomes ensue.

Highlights

- Mild COVID-19 patients displayed an early transcriptional commitment with NK cell function, whereas severe patients do so with neutrophil function.

- The identified co-expressed genes give insights into a coordinated transcriptional program of NK cell cytotoxic activity being associated with mild patients.

- Key checkpoints of NK cell cytotoxicity that were enriched in mild patients include:KLRD1,CD247, andIFNG.

- The early innate immune response related to NK cells connects with the Th1/Th2 adaptive immune responses, supporting their relevance in COVID-19 progression.
Development of robust antiviral assays using relevant apical-out human airway organoids
Authors: Lee Ji-Hoon , LeCher Julia C. , Parigoris Eric , Shinagawa Noriyuki , Sentosa Jason , Manfredi Candela , Goh Shu Ling , De Ramyani , Tao Sijia , Zandi Keivan , Amblard Franck , Sorscher Eric J. , Spence Jason R. , Tirouvanziam Rabindra , Schinazi Raymond F. , Takayama Shuichi
Published: January 2024

While breakthroughs with organoids have emerged as next-generationin vitrotools, standardization for drug discovery remains a challenge. This work introduces human airway organoids with reversed biopolarity (AORBs), cultured and analyzed in a high-throughput, single-organoid-per-well format, enabling milestones towards standardization. AORBs exhibit a spatio-temporally stable apical-out morphology, facilitating high-yield direct intact-organoid virus infection. Single-cell RNA sequencing and immunohistochemistry confirm the physiologically relevant recapitulation of differentiated human airway epithelia. The cellular tropism of five severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) strains along with host response differences between Delta, Washington, and Omicron variants, as observed in transcriptomic profiles, also suggest clinical relevance. Dose-response analysis of three well-studied SARS-CoV-2 antiviral compounds (remdesivir, bemnifosbuvir, and nirmatrelvir) demonstrates that AORBs efficiently predict human efficacy, comparable to gold-standard air-liquid interface cultures, but with higher throughput (∼10-fold) and fewer cells (∼100-fold). This combination of throughput and relevance allows AORBs to robustly detect false negative results in efficacy, preventing irretrievable loss of promising lead compounds. While this work leverages the SARS-CoV-2 study as a proof-of-concept application, the standardization capacity of AORB holds broader implications in line with regulatory efforts to push alternatives to animal studies.
De Novo Atomic Protein Structure Modeling for Cryo-EM Density Maps Using 3D Transformer and Hidden Markov Model
Authors: Giri Nabin , Cheng Jianlin
Published: January 2024

Accurately building three-dimensional (3D) atomic structures from 3D cryo-electron microscopy (cryo-EM) density maps is a crucial step in the cryo-EM-based determination of the structures of protein complexes. Despite improvements in the resolution of 3D cryo-EM density maps, the de novo conversion of density maps into 3D atomic structures for protein complexes that do not have accurate homologous or predicted structures to be used as templates remains a significant challenge. Here, we introduce Cryo2Struct, a fully automated ab initio cryo-EM structure modeling method that utilizes a 3D transformer to identify atoms and amino acid types in cryo-EM density maps first, and then employs a novel Hidden Markov Model (HMM) to connect predicted atoms to build backbone structures of proteins. Tested on a standard test dataset of 128 cryo-EM density maps with varying resolutions (2.1 - 5.6 Å) and different numbers of residues (730 - 8,416), Cryo2Struct built substantially more accurate and complete protein structural models than the widely used ab initio method - Phenix in terms of multiple evaluation metrics. Moreover, on a new test dataset of 500 recently released density maps with varying resolutions (1.9 - 4.0 Å) and different numbers of residues (234 - 8,828), it built more accurate models than on the standard dataset. And its performance is rather robust against the change of the resolution of density maps and the size of protein structures.
Molecular detection using hybridization capture and next-generation sequencing reveals cross-species transmission of feline coronavirus type-1 between a domestic cat and a captive wild felid
Authors: Olarte-Castillo Ximena A. , Goodman Laura B. , Whittaker Gary R
Published: January 2024

Feline coronavirus (FCoV) infection normally causes mild or subclinical signs and is common in domestic cats. However, in some cats, FCoV infection can also lead to the development of feline infectious peritonitis (FIP)—a typically lethal disease. FCoV has two serotypes or genotypes, FCoV-1 and FCoV-2, both of which can cause FIP. The main difference between the genotypes is the viral spike (S) protein that determines tropism and pathogenicity, crucial mechanisms in the development of FIP. Subclinical infection and FIP have both been reported in wild felids, including in threatened species. Due to the high genetic variability of the S gene and the technical challenges to sequencing it, detection and characterization of FCoV in wild felids have mainly centered on other more conserved genes. Therefore, the genotype causing FIP in most wild felids remains unknown. Here, we report a retrospective molecular epidemiological investigation of FCoV in a zoological institution in the United States. In 2008, a domestic cat (Felis catus) and a Pallas’ cat (Otocolobus manul) sharing the same room succumbed to FIP. Using hybridization capture and next-generation sequencing, we detected and sequenced the complete 3’-end of the viral genome (∼8.2 Kb), including the S gene, from both felids. Our data show for the first time that FCoV-1 can be transmitted between domestic and wild felids and extends the known host range of FCoV-1. Our findings highlight the importance of identifying the genotype causing FIP, to develop effective control measures.
A Neural Network for High-Precise and Well-Interpretable Electrocardiogram Classification
Authors: Liu Xiaoqiang , He Liang , Yan Jiadong , Huang Yisen , Wang Yubin , Lin Chanchan , Huang Yingxuan , Liu Xiaobo
Published: January 2024

Manual heart disease diagnosis with the electrocardiogram (ECG) is intractable due to the intertwined signal features and lengthy diagnosis procedure, especially for the 24-hour dynamic ECG signals. Consequently, even experienced cardiologists may face difficulty in producing all accurate ECG reports. In recent years, neural network-based automatic ECG diagnosis methods have exhibited promising performance, suggesting a potential alternative to the labor-intensive examination conducted by cardiologists. However, many existing approaches failed to adequately consider the temporal and channel dimensions when assembling features and ignored interpretability. And clinical theory underscores the necessity of prolonged signal observations for diagnosing certain ECG conditions such as tachycardia. Moreover, specific heart diseases manifest primarily through distinct ECG leads represented as channels. In response to these challenges, this paper introduces a novel neural network architecture for ECG classification (diagnosis). The proposed model incorporates Lead Fusing blocks, transformer-XL encoder-based Encoder modules, and hierarchical temporal attentions. Importantly, this classifier operates directly on raw ECG time-series signals rather than cardiac cycles. Signal integration begins with the Lead Fusing blocks, followed by the Encoder modules and hierarchical temporal attentions, enabling the extraction of long-dependent features. Furthermore, we argue that existing convolution-based methods compromise interpretability, while our proposed neural network offers improved clarity in this regard. Experimental evaluation on a comprehensive public dataset confirms the superiority of our classifier over state-of-the-art methods. Moreover, visualizations reveal the enhanced interpretability provided by our approach.

Highlights

Our model extracts long-dependent features of ECG signals based on the Transformer-XL encoder.

The proposed network offers the improved interpretability.

Our classifier achieves superior performance over other state-of-the-art methods.
Enhancing radiotherapy response via intratumoral injection of the TLR9 agonist CpG to stimulate CD8 T cells in an autochthonous mouse model of sarcoma
Authors: Su Chang , Kent Collin L. , Pierpoint Matthew , Floyd Warren , Luo Lixia , Wiliams Nerissa T. , Ma Yan , Peng Brian , Lazarides Alexander L. , Subramanian Ajay , Himes Jonathan E. , Perez Vincent M. , Hernansaiz-Ballesteros Rosa D. , Roche Kimberly E. , Modliszewski Jennifer L. , Selitsky Sara R. , Shinohara Mari , Wisdom Amy J. , Moding Everett J. , Mowery Yvonne M. , Kirsch David G.
Published: January 2024

Radiation therapy is frequently used to treat cancers including soft tissue sarcomas. Prior studies established that the toll-like receptor 9 (TLR9) agonist cytosine-phosphate-guanine oligodeoxynucleotide (CpG) enhances the response to radiation therapy (RT) in transplanted tumors, but the mechanism(s) remain unclear. Here, we used CRISPR/Cas9 and the chemical carcinogen 3-methylcholanthrene (MCA) to generate autochthonous soft tissue sarcomas with high tumor mutation burden. Treatment with a single fraction of 20 Gy RT and two doses of CpG significantly enhanced tumor response, which was abrogated by genetic or immunodepletion of CD8+ T cells. To characterize the immune response to RT + CpG, we performed bulk RNA-seq, single-cell RNA-seq, and mass cytometry. Sarcomas treated with 20 Gy and CpG demonstrated increased CD8 T cells expressing markers associated with activation and proliferation, such as Granzyme B, Ki-67, and interferon-γ. CpG + RT also upregulated antigen presentation pathways on myeloid cells. Furthermore, in sarcomas treated with CpG + RT, TCR clonality analysis suggests an increase in clonal T-cell dominance. Collectively, these findings demonstrate that RT + CpG significantly delays tumor growth in a CD8 T cell-dependent manner. These results provide a strong rationale for clinical trials evaluating CpG or other TLR9 agonists with RT in patients with soft tissue sarcoma.
Full-length RNA transcript sequencing traces brain isoform diversity in house mouse natural populations
Authors: Zhang Wenyu , Guenther Anja , Gao Yuanxiao , Ullrich Kristian , Huettel Bruno , Ahmad Aftab , Duan Lei , Wei Kaizong , Tautz Diethard
Published: January 2024

The ability to generate multiple RNA transcript isoforms from the same gene is a general phenomenon in eukaryotes. However, the complexity and diversity of alternative isoforms in natural populations remain largely unexplored. Using a newly developed full-length transcripts enrichment protocol with 5’ CAP selection, we sequenced full-length RNA transcripts of 48 individuals from outbred populations and subspecies ofMus musculus, and from the closely related sister speciesMus spretusandMus spicilegusas outgroups. The dataset represents the most extensive full-length high-quality isoform catalog at the population level to date. In total, we reliably identified 117,728 distinct isoforms, of which only 51% were previously annotated. We show that the population-specific distribution pattern of isoforms is phylogenetically informative and reflects the segregating SNP diversity between the populations. We find that ancient housekeeping genes are a major source of the overall isoform diversity, and that the generation of alternative first exons plays a major role in generating new isoforms. Given that our data allow us to distinguish between population-specific isoforms and isoforms that are conserved across multiple populations, it is possible to refine the annotation of the reference mouse genome to a set of about 40,000 isoforms that should be most relevant for comparative functional analysis across species.
Epigenetic Modulation to perturb theSYNGAP1Intellectual Disability (ID) that ameliorates synaptic and behavioural deficits
Authors: Singh Akash Kumar , Joshi Ila , Reddy Neeharika M. N. , Purushotham Sushmitha S. , Eswaramoorthy M. , Vasudevan Madavan , Banerjee Sourav , Clement J. P , Kundu Tapas K
Published: January 2024

Sporadic heterozygous mutations inSYNGAP1affects social and emotional behaviour that are often observed in intellectual disability (ID) and autism spectrum disorder (ASD). Although neurophysiological deficits have been extensively studied, the epigenetic landscape ofSYNGAP1mutation-mediated intellectual disability is unexplored. Here, we have surprisingly found that the p300/CBP specific acetylation marks of histones are significantly repressed in the adolescent hippocampus ofSyngap1+/-mouse. To establish the causal relationship ofSyngap1+/-phenotype and the altered histone acetylation signature we have treated 2-4 months oldSyngap1+/-mouse with glucose-derived carbon nanosphere (CSP) conjugated potent small molecule activator (TTK21) of p300/CBP lysine acetyltransferase (CSP-TTK21). The enhancement of the p300/CBP specific acetylation marks of histones by CSP-TTK21 restored deficits in spine density, synaptic function, and social preferences ofSyngap1+/-mouse that is very closely comparable to wild type littermates. The hippocampal RNA-Seq analysis of the treated mice revealed that the expression of many critical genes related to the ID/ASD reversed due to the treatment of the specific small molecule activator. This study could be the first demonstration of the reversal of autistic behaviour and neural wiring upon the modulation of altered epigenetic modification (s).
A Bag-Of-Motif Model Captures Cell States at Distal Regulatory Sequences
Authors: Cornejo-Páramo Paola , Zhang Xuan , Louis Lithin , Yang Yi-Hua , Li Zelun , Humphreys David , Wong Emily S.
Published: January 2024

Deciphering the intricate regulatory code governing cell-type-specific gene expression is a fundamental goal in genetics. Current methods struggle to capture the complex interplay between gene distal regulatory sequences and cell context. We developed a computational approach, BOM (Bag-of-Motifs), which represents cis-regulatory sequences by the type and number of TF binding motifs it contains, irrespective of motif order, orientation, and spacing. This simple yet powerful representation allows BOM to efficiently capture the complexity of cell-type-specific information encoded within these sequences. We apply BOM to mouse, human, and zebrafish distal regulatory regions, demonstrating remarkable accuracy. Notably, the method outperforms more complex deep learning models at the same task using fewer parameters. BOM can also uncover cross-species sequence similarities unrecognized by genome alignments. We experimentally validate ourin silicopredictions using enhancer reporter assay, showing that motifs with the most significant explanatory power are sequence determinants of cell-type specific enhancer activity. BOM offers a novel systematic framework for studying cell-type or condition-specific cis-regulatory sequences. Using BOM, we demonstrate the existence of a highly predictive sequence code at distal regulatory regions in mammals driven by TF binding motifs.
Prototype mRNA vaccines imprint broadly neutralizing human serum antibodies after Omicron variant-matched boosting
Authors: Liang Chieh-Yu , Raju Saravanan , Liu Zhuoming , Li Yuhao , Arunkumar Guha A. , Case James Brett , Zost Seth J. , Acreman Cory M. , Carvalho dos Anjos Deborah Carolina , McLellan Jason S. , Crowe James E. , Whelan Sean P.J. , Elbashir Sayda M. , Edwards Darin K. , Diamond Michael S.
Published: January 2024

Immune imprinting is a phenomenon in which an individual’s prior antigenic experiences influence responses to subsequent infection or vaccination. Here, using antibody depletion and multiplexed spike-binding assays, we characterized the type-specificity and cross-reactivity of serum antibody responses after mRNA vaccination in mice and human clinical trial participants. In mice, a single priming dose of a preclinical version of mRNA-1273 vaccine encoding Wuhan-1 spike minimally imprinted serum responses elicited by Omicron boosters, enabling a robust generation of type-specific antibodies. However, substantial imprinting was observed in mice receiving an Omicron booster after two priming doses of mRNA-1273, an effect that was mitigated by a second booster dose of Omicron mRNA vaccine. In humans who received two BA.5 or XBB.1.5 Omicron-matched boosters after two or more doses of the prototype mRNA-1273 vaccine, spike-binding and neutralizing serum antibodies cross-reacted with circulating Omicron variants as well as more distantly related sarbecoviruses. Because the serum neutralizing response against Omicron strains and other sarbecoviruses was completely abrogated after pre-clearing with the Wuhan-1 spike protein, antibodies induced by XBB.1.5 boosting in humans focus on conserved epitopes shaped and shared by the antecedent mRNA-1273 primary series. Our depletion analysis also identified cross-reactive neutralizing antibodies that recognize distinct epitopes in the receptor binding domain (RBD) and S2 proteins with differential inhibitory effects on members of the sarbecovirus subgenus. Thus, although the serum antibody response to Omicron-based boosters in humans is dominantly imprinted by prior immunizations with prototype mRNA-1273 vaccines, this outcome can be beneficial as it drives expansion of multiple classes of cross-neutralizing antibodies that inhibit infection of emerging SARS-CoV-2 variants and extend activity to distantly related sarbecoviruses.
Targeting transcription factors through an IMiD independent zinc finger domain
Authors: Liu Bee Hui , Liu Miao , Radhakrishnan Sridhar , Jaladanki Chaitanya Kumar , Gao Chong , Tang Jing Ping , Kumari Kalpana , Go Mei Lin , Vu Kim Anh L. , Seo Hyuk-Soo , Song Kijun , Tian Xi , Feng Li , Tan Justin L. , Bassal Mahmoud A. , Arthanari Haribabu , Qi Jun , Dhe-Paganon Sirano , Fan Hao , Tenen Daniel G. , Chai Li
Published: January 2024

Immunomodulatory imide drugs (IMiDs) degrade specific C2H2 zinc finger degrons in transcription factors, making them effective against certain cancers. SALL4, a cancer driver, contains seven C2H2 zinc fingers in four clusters, including an IMiD degron in zinc finger cluster two (ZFC2). Surprisingly, IMiDs do not inhibit growth of SALL4 expressing cancer cells. To overcome this limit, we focused on a non-IMiD degron, SALL4 zinc finger cluster four (ZFC4). By combining AlphaFold and the ZFC4-DNA crystal structure, we identified a potential ZFC4 drug pocket. Utilizing anin silicodocking algorithm and cell viability assays, we screened chemical libraries and discovered SH6, which selectively targets SALL4-expressing cancer cells. Mechanistic studies revealed that SH6 degrades SALL4 protein through the CUL4A/CRBN pathway, while deletion of ZFC4 abolished this activity. Moreover, SH6 led to significant 62% tumor growth inhibition of SALL4+ xenografts in vivo and demonstrated good bioavailability in pharmacokinetic studies. In summary, these studies represent a new approach for IMiD independent drug discovery targeting C2H2 transcription factors in cancer.

Graphical Abstract
Two Distinct Regulatory Systems Control Pulcherrimin Biosynthesis inBacillus subtilis
Authors: Fernandez Nicolas L. , Simmons Lyle A.
Published: January 2024

Regulation of transcription is a fundamental process that allows bacteria to respond to external stimuli with appropriate timing and magnitude of response. In the soil bacteriumBacillus subtilis,transcriptional regulation is at the core of developmental processes needed for cell survival. Gene expression in cells transitioning from exponential phase to stationary phase is under the control of a group of transcription factors called transition state regulators (TSRs). TSRs influence numerous developmental processes including the decision between biofilm formation and motility, genetic competence, and sporulation, but the extent to which TSRs influence bacterial physiology remains to be fully elucidated. Here, we demonstrate two TSRs, ScoC and AbrB, along with the MerR-family transcription factor PchR negatively regulate production of the iron chelator pulcherrimin inB. subtilis.Genetic analysis of the relationship between the three transcription factors indicate that all are necessary to limit pulcherrimin production during exponential phase and influence the rate and total amount of pulcherrimin produced. Similarly, expression of the pulcherrimin biosynthesis geneyvmCwas found to be under control of ScoC, AbrB, and PchR and correlated with the amount of pulcherrimin produced by each background. Lastly, our in vitro data indicate a weak direct role for ScoC in controlling pulcherrimin production along with AbrB and PchR. The layered regulation by two distinct regulatory systems underscores the important, and somewhat enigmatic, role for pulcherrimin inB. subtilisphysiology.

Author Summary

Regulation of gene expression is important for survival in ever changing environments. In the soil bacteriumBacillus subtilis,key developmental processes are controlled by overlapping networks of transcription factors, some of which are termed transition state regulators (TSRs). Despite decades of research, the scope of how TSRs influenceB. subtilisphysiology is still being uncovered. We found that three transcription factors, two of which are TSRs, converge to inhibit production of the iron-chelator pulcherrimin. Only when all three are missing is pulcherrimin production elevated. Finally, we demonstrate that expression of pulcherrimin biosynthesis genes occurs via direct and indirect regulation by the trio of transcription factors. Due to its iron chelating ability, pulcherrimin has been characterized as a modulator of niche development with antioxidant properties. Thus, our findings that TSRs control pulcherrimin, concurrently with other developmental phenotypes, provides new insight into how TSRs impactB. subtilisand its interaction with the environment.
Differences in selective profiles among single or double stranded DNA or RNA viruses. Statistical detection of its double or single stranded condition
Authors: Valenzuela Carlos Y
Published: January 2024

I developed a test to study the probability distribution of bases in dinucleotides separated by 0, 1, 2…K nucleotides sites. This analysis define selective profiles of dinucleotides by their sign and selection coefficients, their distance to neutrality measured by a chi-square value and the order of significance of their distance to randomness among the 16 possible values. In double stranded (ds) DNA or RNA an index (index = I, with 5’-3’ sense) dinucleotide participates in evolutionary processes together with three dinucleotides defined by base complementarity and 5’-3’ sense. The anti-sense dinucleotide in the same strand, the parallel (Par, with 3’-5’ sense) and anti-parallel (a-Par, with 5’-3’ sense) dinucleotides in the complementary strand. Single stranded (ss) RNA or DNA do not have these set of dinucleotides and would not present similarities or differences, among those four dinucleotides. I studied viromes taken from GenBank, with ds and ss RNA and DNA by this dinucleotide analysis. The core matrix has rows with Separations (Sep) of bases and columns with their selective profiles of the Index, Par and a-Par dinucleotides. Profiles distances I-Par and I-a-Par where compared. The ds RNA or DNA showed larger selective profiles differences between I-Par than I-a-Par. Ss RNA or DNA viruses showed non-significant differences between these comparisons. Those ss virus that exist as ds virus in their hosts, as Hindbis and HIV virus behaved similarly to ds viruses. Coli phage PhiX174 an ss DNA virus showed smaller I-Par than I-a-Par selective distances.

IMPORTANCE

I present a new method for the comparison of selective profiles that allows know whether a RNA or DNA behaves preferentially as ds or ss nucleic acid and gives information about their dynamic or evolutionary mechanisms of replication cycles. This changes our vision of viruses constituted in the virion as ds or ss viruses towards a functional evolutionary vision where viruses are ds and ss according to their behavior along with their life cycle. This evolutionary vision may be crucial in epidemiology.
Evaluation of Gremlin-1 as a therapeutic target in metabolic dysfunction-associated steatohepatitis
Authors: Horn Paul , Norlin Jenny , Almholt Kasper , Viuff Birgitte M. , Galsgaard Elisabeth D. , Hald Andreas , Zosel Franziska , Demuth Helle , Poulsen Svend , Norby Peder L. , Rasch Morten G. , Vyberg Mogens , Werge Mikkel Parsberg , Gluud Lise Lotte , Rink Marco R. , Shepherd Emma , Northall Ellie , Lalor Patricia F. , Weston Chris J. , Fog-Tonnesen Morten , Newsome Philip N.
Published: January 2024

Gremlin-1 has been implicated in liver fibrosis in metabolic dysfunction-associated steatohepatitis (MASH) via inhibition of bone-morphogenetic protein (BMP) signalling and has thereby been identified as a potential therapeutic target. Using ratin vivoand humanin vitroandex vivomodel systems of MASH fibrosis, we show that neutralisation of Gremlin-1 activity with monoclonal therapeutic antibodies does not reduce liver inflammation or liver fibrosis. Still, Gremlin-1 was upregulated in human and rat MASH fibrosis, but expression was restricted to a small subpopulation of COL3A1/THY1+myofibroblasts. Lentiviral overexpression of Gremlin-1 in LX-2 cells and primary hepatic stellate cells led to changes in BMP-related gene expression, which did not translate to increased fibrogenesis. Furthermore, we show that Gremlin-1 binds to heparin with high affinity, which prevents Gremlin-1 from entering systemic circulation, prohibiting Gremlin-1-mediated organ crosstalk. Overall, our findings suggest a redundant role for Gremlin-1 in the pathogenesis of liver fibrosis, which is unamenable to therapeutic targeting.
Ileal mucus viscoelastic properties differ in Crohn’s disease
Authors: Kramer Catharina , Rulff Hanna , Ziegler Jörn Felix , Alzain Nadra , Addante Annalisa , Kuppe Aditi , Timm Sara , Schrade Petra , Bischoff Philip , Glauben Rainer , Dürr Julia , Ochs Matthias , Mall Marcus A. , Gradzielski Michael , Siegmund Britta
Published: January 2024

Crohn’s disease (CD) is an inflammatory bowel disease (IBD) that can affect any part of the gastrointestinal tract, frequently involving the terminal ileum. While colonic mucus alterations in CD patients have been described, terminal ileal mucus and its mechanobiological properties have been neglected. Our study is the first of its kind to decipher the viscoelastic and network properties of ileal mucus.

With that aim, oscillatory rheological shear measurements based on an airway mucus protocol that was thoroughly validated for ileal mucus were performed. Our pilot study analyzed terminal ileum mucus from controls (n=11) and CD patients (n=11). Mucus network structure was visualized by scanning electron microscopy (SEM).

Interestingly, a statistically significant increase in viscoelasticity as well as a decrease in mesh size was observed in ileal mucus from CD patients compared to controls. Furthermore, rheological data were analyzed in relation to study participants clinical characteristics, such as inflammatory status, revealing noteworthy trends.

In conclusion, this study provides the first data on the viscoelastic properties and structure of human ileal mucus in the healthy state and in CD, demonstrating significant alterations between groups and highlighting the need for further research on mucus and its effect on the underlying epithelial barrier.
Differential activation of rhodopsin triggers distinct endocytic trafficking and recycling in vivo via differential phosphorylation
Authors: Ferng Darwin , Sun Wesley , Shieh Bih-Hwa
Published: January 2024

Activated GPCRs are phosphorylated and internalized mostly via clathrin-mediated endocytosis (CME), which are then sorted for recycling or degradation. We investigated how differential activation of the same GPCR affects its endocytic traffickingin vivousing rhodopsin as a model in flies expressing mCherry-tagged rhodopsin 1 (Rh1-mC) or GFP-tagged arrestin 1 (Arr1-GFP). Upon blue light stimulation, activated Rh1 recruited Arr1-GFP to the rhabdomere, which became co-internalized and accumulated in cytoplasmic vesicles of photoreceptors. This internalization was eliminated inshits1mutants affecting dynamin. Moreover, it was blocked by eitherrdgAorrdgBmutations affecting the PIP2biosynthesis. Together, the blue light-initiated internalization of Rh1 and Arr1 belongs to CME. Green light stimulation also triggered the internalization and accumulation of activated Rh1-mC in the cytoplasm but with faster kinetics. Importantly, Arr1-GFP was also recruited to the rhabdomere but not co-internalized with Rh1-mC. This endocytosis was not affected inshits1norrdgAmutants, indicating it is not CME. We explored the fate of internalized Rh1-mC following CME and observed it remained in cytoplasmic vesicles following 30 min of dark adaptation. In contrast, in the non-CME Rh1-mC appeared readily recycled back to the rhabdomere within five min of dark treatment. This faster recycling may be regulated by rhodopsin phosphatase, RdgC. Together, we demonstrate two distinct endocytic and recycling mechanisms of Rh1 via two light stimulations. It appears that each stimulation triggers a distinct conformation leading to different phosphorylation patterns of Rh1 capable of recruiting Arr1 to rhabdomeres. However, a more stable interaction leads to the co-internalization of Arr1 that orchestrates CME. A stronger Arr1 association appears to impede the recycling of the phosphorylated Rh1 by preventing the recruitment of RdgC. We conclude that conformations of activated rhodopsin determine the downstream outputs upon phosphorylation that confers differential protein-protein interactions.
Mitochondrial reverse electron transport in myeloid cells perpetuates neuroinflammation
Authors: Peruzzotti-Jametti L. , Willis C.M. , Hamel R. , Krzak G. , Reisz J.A. , Prag H.A. , Wu V. , Xiang Y. , van den Bosch A.M.R. , Nicaise A.M. , Roth L. , Bates G.R. , Huang H. , Vincent A.E. , Frezza C. , Viscomi C. , Marioni J.C. , D’Alessandro A. , Takats Z. , Murphy M.P. , Pluchino S.
Published: January 2024

Sustained smouldering, or low grade, activation of myeloid cells is a common hallmark of several chronic neurological diseases, including multiple sclerosis (MS)1. Distinct metabolic and mitochondrial features guide the activation and the diverse functional states of myeloid cells2. However, how these metabolic features act to perpetuate neuroinflammation is currently unknown. Using a multiomics approach, we identified a new molecular signature that perpetuates the activation of myeloid cells through mitochondrial complex II (CII) and I (CI) activity driving reverse electron transport (RET) and the production of reactive oxygen species (ROS). Blocking RET in pro-inflammatory myeloid cells protected the central nervous system (CNS) against neurotoxic damage and improved functional outcomes in animal disease modelsin vivo. Our data show that RET in myeloid cells is a potential new therapeutic target to foster neuroprotection in smouldering inflammatory CNS disorders3.
GM-CSF-mediated epithelial-immune cell crosstalk orchestrates pulmonary immunity toAspergillus fumigatus
Authors: Mills Kathleen A. M. , Westermann Frederike , Espinosa Vanessa , Rosiek Eric , Desai Jigar V. , Aufiero Mariano A. , Guo Yahui , Mitchell Kennedy A. , Tuzlak Selma , De Feo Donatella , Lionakis Michail S. , Rivera Amariliz , Becher Burkhard , Hohl Tobias M.
Published: January 2024

Aspergillus fumigatuscauses life-threatening mold pneumonia in immune compromised patients, particularly in those with quantitative or qualitative defects in neutrophils. While innate immune cell crosstalk licenses neutrophil antifungal activity in the lung, the role of epithelial cells in this process is unknown. Here, we find that that surfactant protein C (SPC)-expressing lung epithelial cells integrate infection-induced IL-1 and type III interferon signaling to produce granulocyte-macrophage colony-stimulating factor (GM-CSF) preferentially at local sites of fungal infection and neutrophil influx. Usingin vivomodels that distinguish the role of GM-CSF during acute infection from its homeostatic function in alveolar macrophage survival and surfactant catabolism, we demonstrate that epithelial-derived GM-CSF increases the accumulation and fungicidal activity of GM-CSF-responsive neutrophils, with the latter being essential for host survival. Our findings establish SPC+epithelial cells as a central player in regulating the quality and strength of neutrophil-dependent immunity against inhaled mold pathogens.

HIGHLIGHTS

GM-CSF is essential for host defense againstA. fumigatusin the lung

IL-1 and IFN-λ promote GM-CSF production by lung epithelial cells in parallel

Epithelial cell-derived GM-CSF increases neutrophil accumulation and fungal killing capacity

Epithelial cells preferentially upregulate GM-CSF in local sites of inflammation

GRAPHICAL ABSTRACT
A Murine Model of Post-acute Neurological Sequelae Following SARS-CoV-2 Variant Infection
Authors: Singh Ankita , Adam Awadalkareem , Aditi  , Peng Bi-Hung , Yu Xiaoying , Zou Jing , Kulkarni Vikram V , Kan Peter , Jiang Wei , Shi Pei-Yong , Samir Parimal , Cisneros Irma , Wang Tian
Published: January 2024

Viral variant is one known risk factor associated with post-acute sequelae of COVID-19 (PASC), yet the pathogenesis is largely unknown. Here, we studied SARS-CoV-2 Delta variant-induced PASC in K18-hACE2 mice. The virus replicated productively, induced robust inflammatory responses in lung and brain tissues, and caused weight loss and mortality during the acute infection. Longitudinal behavior studies in surviving mice up to 4 months post-acute infection revealed persistent abnormalities in neuropsychiatric state and motor behaviors, while reflex and sensory functions recovered over time. Surviving mice showed no detectable viral RNA in the brain and minimal neuroinflammation post-acute infection. Transcriptome analysis revealed persistent activation of immune pathways, including humoral responses, complement, and phagocytosis, and reduced levels of genes associated with ataxia telangiectasia, impaired cognitive function and memory recall, and neuronal dysfunction and degeneration. Furthermore, surviving mice maintained potent T helper 1 prone cellular immune responses and high neutralizing antibodies against Delta and Omicron variants in the periphery for months post-acute infection. Overall, infection in K18-hACE2 mice recapitulates the persistent clinical symptoms reported in long COVID patients and may be useful for future assessment of the efficacy of vaccines and therapeutics against SARS-CoV-2 variants.
Somatic cancer driver mutations are enriched and associated with inflammatory states in Alzheimer’s disease microglia
Authors: Huang August Yue , Zhou Zinan , Talukdar Maya , Miller Michael B. , Chhouk Brian , Enyenihi Liz , Rosen Ila , Stronge Edward , Zhao Boxun , Kim Dachan , Choi Jaejoon , Khoshkhoo Sattar , Kim Junho , Ganz Javier , Travaglini Kyle , Gabitto Mariano , Hodge Rebecca , Kaplan Eitan , Lein Ed , De Jager Philip L. , Bennett David A. , Lee Eunjung Alice , Walsh Christopher A.
Published: January 2024

Alzheimer’s disease (AD) is an age-associated neurodegenerative disorder characterized by progressive neuronal loss and pathological accumulation of the misfolded proteins amyloid-β and tau1,2. Neuroinflammation mediated by microglia and brain-resident macrophages plays a crucial role in AD pathogenesis1–5, though the mechanisms by which age, genes, and other risk factors interact remain largely unknown. Somatic mutations accumulate with age and lead to clonal expansion of many cell types, contributing to cancer and many non-cancer diseases6,7. Here we studied somatic mutation in normal aged and AD brains by three orthogonal methods and in three independent AD cohorts. Analysis of bulk RNA sequencing data from 866 samples from different brain regions revealed significantly higher (∼two-fold) overall burdens of somatic single-nucleotide variants (sSNVs) in AD brains compared to age-matched controls. Molecular-barcoded deep (>1000X) gene panel sequencing of 311 prefrontal cortex samples showed enrichment of sSNVs and somatic insertions and deletions (sIndels) in cancer driver genes in AD brain compared to control, with recurrent, and often multiple, mutations in genes implicated in clonal hematopoiesis (CH)8,9. Pathogenic sSNVs were enriched in CSF1R+ microglia of AD brains, and the high proportion of microglia (up to 40%) carrying some sSNVs in cancer driver genes suggests mutation-driven microglial clonal expansion (MiCE). Analysis of single-nucleus RNA sequencing (snRNAseq) from temporal neocortex of 62 additional AD cases and controls exhibited nominally increased mosaic chromosomal alterations (mCAs) associated with CH10,11. Microglia carrying mCA showed upregulated pro-inflammatory genes, resembling the transcriptomic features of disease-associated microglia (DAM) in AD. Our results suggest that somatic driver mutations in microglia are common with normal aging but further enriched in AD brain, driving MiCE with inflammatory and DAM signatures. Our findings provide the first insights into microglial clonal dynamics in AD and identify potential new approaches to AD diagnosis and therapy.
Mutation of a highly conserved isoleucine residue in loop 2 of several 𝛽-coronavirus macrodomains indicates that enhanced ADP-ribose binding is detrimental to infection
Authors: Kerr Catherine M , Pfannenstiel Jessica J , Alhammad Yousef M , O'Connor Joseph J , Ghimire Roshan , Shrestha Rakshya , Khattabi Reem , Saenjamsai Pradtahna , Parthasarathy Srivatsan , McDonald Peter R , Gao Philip , Johnson David K , More Sunil , Roy Anuradha , Channappanavar Rudragouda , Fehr Anthony R
Published: January 2024

All coronaviruses (CoVs) encode for a conserved macrodomain (Mac1) located in nonstructural protein 3 (nsp3). Mac1 is an ADP-ribosylhydrolase that binds and hydrolyzes mono-ADP-ribose from target proteins. Previous work has shown that Mac1 is important for virus replication and pathogenesis. Within Mac1, there are several regions that are highly conserved across CoVs, including the GIF (glycine-isoleucine-phenylalanine) motif. To determine how the biochemical activities of these residues impact CoV replication, the isoleucine and the phenylalanine residues were mutated to alanine (I-A/F-A) in both recombinant Mac1 proteins and recombinant CoVs, including murine hepatitis virus (MHV), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The F-A mutant proteins had ADP-ribose binding and/or hydrolysis defects that led to attenuated replication and pathogenesis in cell culture and mice. In contrast, the I-A mutations had normal enzyme activity and enhanced ADP-ribose binding. Despite increased ADP-ribose binding, I-A mutant MERS-CoV and SARS-CoV-2 were highly attenuated in both cell culture and mice, indicating that this isoleucine residue acts as a gate that controls ADP-ribose binding for efficient virus replication. These results highlight the function of this highly conserved residue and provide unique insight into how macrodomains control ADP-ribose binding and hydrolysis to promote viral replication.
Loss of αBa-crystallin, but not αA-crystallin, increases age-related cataract in the zebrafish lens
Authors: Posner Mason , Garver Taylor , Kaye Taylor , Brdicka Stuart , Suttle Madison , Patterson Bryce , Farnsworth Dylan R.
Published: January 2024

The vertebrate eye lens is an unusual organ in that most of its cells lack nuclei and the ability to replace aging protein. The small heat shock protein α-crystallins evolved to become key components of this lens, possibly because of their ability to prevent aggregation of aging protein that would otherwise lead to lens opacity. Most vertebrates express two α-crystallins, αA- and αB-crystallin, and mutations in each are linked to human cataract. In a mouse knockout model only the loss of αA-crystallin led to early-stage lens cataract. We have used the zebrafish as a model system to investigate the role of α-crystallins during lens development. Interestingly, while zebrafish express one lens-specific αA-crystallin gene (cryaa), they express two αB-crystallin genes, with one evolving lens specificity (cryaba) and the other retaining the broad expression of its mammalian ortholog (cryabb). In this study we used individual mutant zebrafish lines for all three α-crystallin genes to determine the impact of their loss on age-related cataract. Surprisingly, unlike mouse knockout models, we found that the loss of the αBa-crystallin genecryabaled to an increase in lens opacity compared tocryaanull fish at 24 months of age. Loss of αA-crystallin did not increase the prevalence of cataract. We also used single cell RNA-Seq and RT-qPCR data to show a shift in the lens expression of zebrafish α-crystallins between 5 and 10 days post fertilization (dpf), with 5 and 6 dpf lenses expressingcryaaalmost exclusively, and expression ofcryabaandcryabbbecoming more prominent after 10 dpf. These data show thatcryaais the primary α-crystallin during early lens development, while the protective role forcryababecomes more important during lens aging. This study is the first to quantify cataract prevalence in wild-type zebrafish, showing that lens opacities develop in approximately 25% of fish by 18 months of age. None of the three α-crystallin mutants showed a compensatory increase in the expression of the remaining two crystallins, or in the abundant βB1-crystallin. Overall, these findings indicate an ontogenetic shift in the functional importance of individual α-crystallins during zebrafish lens development. Our finding that the lens-specific zebrafish αBa-crystallin plays the leading role in preventing age-related cataract adds a new twist to our understanding of vertebrate lens evolution.