bluuebunny/crossref_metadata_2025 huggingface dataset:
An Aetiological and Observational Cohort study of Interstitial Lung Abnormalities (ILAs) Including Progressive Fibrosing Phenotype Interstitial Lung Diseases in a Large General Medical Check-up Population (Kumamoto ILA study in Japan) (KILA-J)
Authors: Ichikado Kazuya , Ichiyasu Hidenori , Iyonaga Kazuhiro , Kawamura Kodai , Higashi Noritaka , Johkoh Takeshi , Fujimoto Kiminori , Morinaga Jun , Yoshida Minoru , Mitsuzaki Katsuhiko , Suga Moritaka , Tanabe Naoya , Handa Tomohiro , Hirai Toyohiro , Sakagami Takuro
Published: January 2023

Introduction

Interstitial lung abnormalities (ILAs) are subtle or mild parenchymal abnormalities observed in more than 5 % of the lungs on CT scans in patients in whom interstitial lung disease was not previously clinically suspected and is considered. ILA is considered to be partly undeveloped stages of idiopathic pulmonary fibrosis (IPF) or progressive pulmonary fibrosis (PPF). This study aims to clarify the frequency of subsequent IPF or PPF diagnosis, the natural course from the preclinical status of the diseases, and the course after commencing treatment.

Methods and analysis

This is an ongoing, prospective, multicentre observational cohort study of patients with ILA referred from general health screening facilities with more than 70000 annual attendances. Up to 500 participants will be enrolled annually over 3 years, with 5-year assessments every six months. Treatment intervention including anti-fibrotic agents will be introduced in disease progression cases. The primary outcome is the frequency of subsequent IPF or PPF diagnoses. Additionally, secondary and further endpoints are associated with the efficacy of early therapeutic interventions in cases involving disease progression.

Ethics and dissemination

This study protocol and informed consent documents have been approved by the Institutional Review Boards of Kumamoto University Hospital (approval number: 2368), Saiseikai Kumamoto Hospital (approval number: 809), and each participating institution. Additionally, written informed consent will be obtained from all participants. Patient recruitment commenced on 20 June 2022. The results will be disseminated through peer-reviewed publications and international conferences.

Trial registration number

UMIN000045149

Strengths and limitations of this study

This is the first prospective, multicentre, observational study to clarify the following points:

the aetiological data of patients with ILA from a large general health check-up population

the natural course of IPF or PPF from the asymptomatic stage

the effects and outcomes of early therapeutic intervention including anti-fibrotic agents for progressive cases of ILA.

The limitations of the study:

participants missing the regular 6-monthly visits after consenting to participate in the study because of their asymptomatic or the pandemic viral infection.
OH-EpiCap: a semi-quantitative tool for the evaluation of One Health epidemiological surveillance capacities and capabilities
Authors: Tegegne Henok Ayalew , Bogaardt Carlijn , Collineau Lucie , Cazeau Géraldine , Lailler Renaud , Reinhardt Johana , Taylor Emma L. , Prada Joaquin , Hénaux Viviane
Published: January 2023

Although international health agencies encourage the development of One Health (OH) surveillance, many systems remain mostly compartmentalized, with limited collaborations among sectors and disciplines. In the framework of the OH European Joint Programme “MATRIX” project, a generic evaluation tool called OH-EpiCap has been developed to enable individual institutes/governments to characterize, assess and monitor their own OH epidemiological surveillance capacities and capabilities. The tool is organized around three dimensions: organization, operational activities, and impact of the OH surveillance system; each dimension is then divided into four targets, each including four indicators. A semi-quantitative questionnaire enables the scoring of each indicator, with four levels according to the degree of satisfaction in the studied OH surveillance system. The evaluation is conducted by a panel of surveillance representatives (during a half-day workshop or with a back-and-forth process to reach a consensus). An R Shiny-based web application facilitates implementation of the evaluation and visualization of the results, and includes a benchmarking option. The tool was piloted on several foodborne hazards (i.e.Salmonella, Campylobacter, Listeria), emerging threats (e.g. antimicrobial resistance) and other zoonotic hazards (psittacosis) in multiple European countries in 2022. These case studies showed that the OH-EpiCap tool supports the tracing of strengths and weaknesses in epidemiological capacities and the identification of concrete and direct actions to improve collaborative activities at all steps of surveillance. It appears complementary to the existing EU-LabCap tool, designed to assess the capacity and capability of European microbiology laboratories. In addition, it provides opportunity to reinforce trust between surveillance stakeholders from across the system and to build a good foundation for a professional network for further collaboration.
Acceptability of decentralizing childhood tuberculosis diagnosis in low-income countries with high tuberculosis incidence: experiences and perceptions from health care workers in sub-Saharan Africa and South-East Asia
Authors: Joshi Basant , De Lima Yara Voss , Massom Douglas Mbang , Kaing Sanary , Banga Marie-France , Kamara Egerton Tamba , Sesay Sheriff , Borand Laurence , Taguebue Jean-Voisin , Moh Raoul , Khosa Celso , Breton Guillaume , Mwanga-Amumpaire Juliet , Bonnet Maryline , Wobudeya Eric , Marcy Olivier , Orne-Gliemann Joanna
Published: January 2023

Background

Decentralizing childhood tuberculosis services, including diagnosis, is now recommended by the WHO and could contribute to increasing tuberculosis detection in high burden countries. However, implementing microbiological tests and clinical evaluation could be challenging for health care workers (HCWs) in Primary Health Centers (PHCs) and even District Hospitals (DHs). We sought to assess the acceptability of decentralizing a comprehensive childhood tuberculosis diagnosis package from HCWs’ perspective.

Design/Methods

We conducted implementation research nested within the TB-Speed Decentralization study. HCWs from two health districts of Cambodia, Cameroon, Côte d’Ivoire, Mozambique, Sierra Leone, and Uganda implemented systematic screening, nasopharyngeal aspirates (NPA) and stool sample collection with molecular testing, clinical evaluation and chest X-ray (CXR) interpretation. We investigated their experiences and perceptions in delivering the diagnostic package components in 2020-21 using individual semi-structured interviews. We conducted thematic analysis, supported by the Theoretical Framework of Acceptability.

Results

HCWs (n=130, 55% female, median age 36 years, 53% nurses, 72% PHC-based) perceived that systematic screening, although increasing workload, was beneficial as it improved childhood tuberculosis awareness. Most HCWs shared satisfaction and confidence in performing NPA, despite procedure duration, need to involve parents/colleagues and discomfort for children. HCWs shared positive attitudes towards stool sample-collection but were frustrated by delayed collection associated with cultural practices, transport and distance challenges. Molecular testing, conducted by nurses or laboratory technicians, was perceived as providing quality results, contributing to diagnosis. Clinical evaluation and diagnosis raised self-efficacy issues and need for continuous training and clinical mentoring. HCWs valued CXR, however complained that technical and logistical problems limited access to digital reports. Referral from PHC to DH was experienced as burdensome.

Conclusion

HCWs at DH and PHC-levels perceived and experienced decentralized childhood tuberculosis diagnosis as acceptable. Implementation however could be hampered by feasibility issues, and calls for innovative referral mechanisms for patients, samples and CXR.
Symptomatic malaria enhances protection from reinfection with homologousPlasmodium falciparumparasites
Authors: Markwalter Christine F. , Petersen Jens E. V. , Zeno Erica E. , Sumner Kelsey M. , Freedman Elizabeth , Mangeni Judith N. , Abel Lucy , Obala Andrew A. , Prudhomme-O’Meara Wendy , Taylor Steve M.
Published: January 2023

A signature remains elusive of naturally-acquired immunity againstPlasmodium falciparum. We identifiedP. falciparumin a 14-month cohort of 239 people in Kenya, genotyped at immunogenic parasite targets expressed in the pre-erythrocytic (circumsporozoite protein, CSP) and blood (apical membrane antigen 1, AMA-1) stages, and classified into epitope type based on variants in the DV10, Th2R, and Th3R epitopes in CSP and the c1L region of AMA-1. Compared to asymptomatic index infections, symptomatic malaria was associated with a reduced reinfection by parasites bearing homologous CSP-Th2R (adjusted hazard ratio [aHR]:0.63; 95% CI:0.45–0.89; p=0.008) CSP-Th3R (aHR:0.71; 95% CI:0.52–0.97; p=0.033), and AMA-1 c1L (aHR:0.63; 95% CI:0.43–0.94; p=0.022) epitope types. The association of symptomatic malaria with reduced risk of homologous reinfection was strongest for rare epitope types. Symptomatic malaria more effectively promotes functional immune responses. The phenotype represents a legible molecular epidemiologic signature of naturally-acquired immunity by which to identify new antigen targets.
Spatial molecular imaging of the human type 2 diabetic islet
Authors: Kolar Grant R. , Ross David , Killingbeck Emily E. , Samson Willis K. , Yosten Gina L. C.
Published: January 2023

The islets of Langerhans are complex micro-organs comprised of multiple cell types essential for the maintenance of glucose homeostasis. The endocrine cell types of the islets engage in intimate, intercellular communication that is necessary for normal secretory activity. Disruption of this intercellular communication, which is at least partially dependent on the spatial organization of individual islets, leads to secretory dysfunction and exacerbation of the symptomology of disease states such as type 2 diabetes. However, the molecular determinants of the mechanisms underlying disrupted intercellular communication remain incompletely understood. Herein we describe the utilization of CosMx™ Spatial Molecular Imaging (SMI) to interrogate transcriptomic changes associated with the transition from the obese, prediabetic state to overt type 2 diabetes. Using SMI, we verified previously reported findings regarding islet composition in the obese and type 2 diabetic states, including loss of beta cells and expansion of alpha cell mass. In addition, we identified changes in the islet neighborhood that have implications for the function of islet endocrine cells. In particular, we identified a subset of alpha cells oriented in the periphery of the islets that appear to exhibit a transcriptomic profile suggestive of de-differentiation toward a beta cell-like transcriptome-type. To our knowledge, this is the first study utilizing spatial molecular imaging to investigate single-cell transcriptomes of individual islets. Further exploration of the intersection of islet architecture and gene expression using spatial technologies is expected to yield novel insights into the mechanisms underlying the development and progression of metabolic diseases like type 2 diabetes.
Machine Learning Reveals Lipidome Remodeling Dynamics in a Mouse Model of Ovarian Cancer
Authors: Bifarin Olatomiwa O. , Sah Samyukta , Gaul David A. , Moore Samuel G. , Chen Ruihong , Palaniappan Murugesan , Kim Jaeyeon , Matzuk Martin M. , Fernández Facundo M.
Published: January 2023

Ovarian cancer (OC) is one of the deadliest cancers affecting the female reproductive system. It may present little or no symptoms at the early stages, and typically unspecific symptoms at later stages. High-grade serous ovarian cancer (HGSC) is the subtype responsible for most ovarian cancer deaths. However, very little is known about the metabolic course of this disease, particularly in its early stages. In this longitudinal study, we examined the temporal course of serum lipidome changes using a robust HGSC mouse model and machine learning data analysis. Early progression of HGSC was marked by increased levels of phosphatidylcholines and phosphatidylethanolamines. In contrast, later stages featured more diverse lipids alterations, including fatty acids and their derivatives, triglycerides, ceramides, hexosylceramides, sphingomyelins, lysophosphatidylcholines, and phosphatidylinositols. These alterations underscored unique perturbations in cell membrane stability, proliferation, and survival during cancer development and progression, offering potential targets for early detection and prognosis of human ovarian cancer.

Teaser

Time-resolved lipidome remodeling in an ovarian cancer model is studied through lipidomics and machine learning.
Simultaneous proteome localization and turnover analysis reveals spatiotemporal features of protein homeostasis disruptions
Authors: Currie Jordan , Manda Vyshnavi , Robinson Sean K. , Lai Celine , Agnihotri Vertica , Hidalgo Veronica , Ludwig R. W. , Zhang Kai , Pavelka Jay , Wang Zhao V. , Rhee June-Wha , Lam Maggie P. Y. , Lau Edward
Published: January 2023

The functions of proteins depend on their spatial and temporal distributions, which are not directly measured by static protein abundance. Under endoplasmic reticulum (ER) stress, the unfolded protein response (UPR) pathway remediates proteostasis in part by altering the turnover kinetics and spatial distribution of proteins. A global view of these spatiotemporal changes has yet to emerge and it is unknown how they affect different cellular compartments and pathways. Here we describe a mass spectrometry-based proteomics strategy and data analysis pipeline, termed Simultaneous Proteome Localization and Turnover (SPLAT), to measure concurrently the changes in protein turnover and subcellular distribution in the same experiment. Investigating two common UPR models of thapsigargin and tunicamycin challenge in human AC16 cells, we find that the changes in protein turnover kinetics during UPR varies across subcellular localizations, with overall slowdown but an acceleration in endoplasmic reticulum and Golgi proteins involved in stress response. In parallel, the spatial proteomics component of the experiment revealed an externalization of amino acid transporters and ion channels under UPR, as well as the migration of RNA-binding proteins toward an endosome co-sedimenting compartment. The SPLAT experimental design classifies heavy and light SILAC labeled proteins separately, allowing the observation of differential localization of new and old protein pools and capturing a partition of newly synthesized EGFR and ITGAV to the ER under stress that suggests protein trafficking disruptions. Finally, application of SPLAT toward human induced pluripotent stem cell derived cardiomyocytes (iPSC-CM) exposed to the cancer drug carfilzomib, identified a selective disruption of proteostasis in sarcomeric proteins as a potential mechanism of carfilzomib-mediated cardiotoxicity. Taken together, this study provides a global view into the spatiotemporal dynamics of human cardiac cells and demonstrates a method for inferring the coordinations between spatial and temporal proteome regulations in stress and drug response.
Dimensional Reduction for Single Molecule Imaging of DNA and Nucleosome Condensation by Polyamines, HP1α and Ki-67
Authors: Benning Nils A. , Kæstel-Hansen Jacob , Rashid Fahad , Park Sangwoo , Urteaga Raquel Merino , Liao Ting-Wei , Hao Jingzhou , Berger James M. , Hatzakis Nikos S. , Ha Taekjip
Published: January 2023

Macromolecules organize themselves into discrete membrane-less compartments. Mounting evidence has suggested that nucleosomes as well as DNA itself can undergo clustering or condensation to regulate genomic activity. Current in vitro condensation studies provide insight into the physical properties of condensates, such as surface tension and diffusion. However, such studies lack the resolution needed for complex kinetic studies of multicomponent condensation. Here, we use a supported lipid bilayer platform in tandem with total internal reflection microscopy to observe the 2-dimensional movement of DNA and nucleosomes at the single-molecule resolution. This dimensional reduction from 3-dimensional studies allows us to observe the initial condensation events and dissolution of these early condensates in the presence of physiological condensing agents. Using polyamines, we observed that the initial condensation happens on a timescale of minutes while dissolution occurs within seconds upon charge inversion. Polyamine valency, DNA length and GC content affect threshold polyamine concentration for condensation. Protein-based nucleosome condensing agents, HP1α and Ki-67, have much lower threshold concentration for condensation than charge-based condensing agents, with Ki-67 being the most effective as low as 100 pM for nucleosome condensation. In addition, we did not observe condensate dissolution even at the highest concentrations of HP1α and Ki-67 tested. We also introduce a two-color imaging scheme where nucleosomes of high density labeled in one color is used to demarcate condensate boundaries and identical nucleosomes of another color at low density can be tracked relative to the boundaries after Ki-67 mediated condensation. Our platform should enable the ultimate resolution of single molecules in condensation dynamics studies of chromatin components under defined physicochemical conditions.

Abstract Figure
Detection of tumor-derived extracellular vesicles interactions with immune cells is dependent on EV-labelling methods
Authors: Loconte Luisa , Arguedas Davinia , Chipont Anna , El Rojbin , Guyonnet Lea , Guerin Coralie , Piovesana Ester , Vázquez-Ibar José Luis , Joliot Alain , Théry Clotilde , Martín-Jaular Lorena
Published: January 2023

Cell-cell communication within the complex tumor microenvironment is critical to cancer progression. Tumor-derived extracellular vesicles (TD-EVs) are key players in this process. They can interact with immune cells and modulate their activity, either suppressing or activating the immune system. Understanding the interactions between TD-EVs and immune cells is essential for understanding immune modulation by cancer cells. Fluorescent labelling of TD-EVs is a method of choice to study such interaction. This work aims to determine the impact of EV labelling methods on the detection of EV interaction and capture by the different immune cell types within human Peripheral Blood Mononuclear Cells (PBMCs), analyzed by imaging flow cytometry and multicolor spectral flow cytometry. EVs released by the triple-negative breast carcinoma cell line MDA-MB-231 were labeled either with the lipophilic dye MemGlow-488 (MG-488), with Carboxyfluorescein diacetate, succinimidyl ester (CFDA-SE), or through expression of a MyrPalm-superFolder GFP (sfGFP) that incorporates into EVs during their biogenesis using a genetically engineered cell line. Our results showed that these different labeling strategies, although analyzed with the same techniques, led to diverging results. While MG-488-labelled EVs incorporate in all cell types, CFSE-labelled EVs are restricted to a minor subset of cells and sfGFP-labelled EVs are mainly detected in CD14+ monocytes which are the main uptakers of EVs and other particles, regardless of the labeling method. Moreover, MG-488-labeled liposomes behaved similarly to MG-488 EVs, highlighting the predominant role of the labelling strategy on the visualization and analysis of TD-EVs uptake by immune cell types. Consequently, the use of different EV labeling methods has to be considered as they can provide complementary information on various types of EV-cell interaction and EV fate.
Drug-Mediated Control of Receptor Valency Enhances Immune Cell Potency
Authors: Finn Paul B. , Chavez Michael , Chen Xinyi , Wang Haifeng , Rane Draven A. , Gurjar Jitendra , Qi Lei S.
Published: January 2023

Designer T cells offer a novel paradigm for treating diseases like cancer, yet they are often hindered by target recognition evasion and limited in vivo control. To overcome these challenges, we develop valency-controlled receptors (VCRs), a novel class of synthetic receptors engineered to enable precise modulation of immune cell activity. VCRs use custom-designed valency-control ligands (VCLs) to modulate T cell signaling via spatial molecular clustering. Using multivalent DNA origami as VCL, we first establish that valency is important for tuning the activity of CD3-mediated immune activation. We then generate multivalent formats of clinically relevant drugs as VCL and incorporate VCR into the architecture of chimeric antigen receptors (CARs). Our data demonstrate that VCL-mediated VCRs can significantly amplify CAR activities and improve suboptimal CARs. Finally, through medicinal chemistry, we synthesize programmable, bioavailable VCL drugs that potentiate targeted immune response against low-antigen tumors both in vitro and in vivo. Our findings establish receptor valency as a core mechanism for enhancing CAR functionality and offer a synthetic chemical biology platform for strengthening customizable, potent, and safer cell therapies.
The AFB1 auxin receptor controls the cytoplasmic auxin response pathway inArabidopsis thaliana
Authors: Dubey Shiv Mani , Han Soeun , Stutzman Nathan , Prigge Michael J , Medvecká Eva , Platre Matthieu Pierre , Busch Wolfgang , Fendrych Matyáš , Estelle Mark
Published: January 2023

The phytohormone auxin triggers root growth inhibition within seconds via a non-transcriptional pathway. Among members of the TIR1/AFBs auxin receptor family, AFB1 has a primary role in this rapid response. However, the unique features that confer this specific function have not been identified. Here we show that the N-terminal region of AFB1, including the F-box domain and residues that contribute to auxin binding, are essential and sufficient for its specific role in the rapid response. Substitution of the N-terminal region of AFB1 with that of TIR1 disrupts its distinct cytoplasm-enriched localization and activity in rapid root growth inhibition. Importantly, the N-terminal region of AFB1 is indispensable for auxin-triggered calcium influx which is a prerequisite for rapid root growth inhibition. Furthermore, AFB1 negatively regulates lateral root formation and transcription of auxin-induced genes, suggesting that it plays an inhibitory role in canonical auxin signaling. These results suggest that AFB1 may buffer the transcriptional auxin response while it regulates rapid changes in cell growth that contribute to root gravitropism.
Dynamical control enables the formation of demixed biomolecular condensates
Authors: Lin Andrew Z. , Ruff Kiersten M. , Jalihal Ameya , Dar Furqan , King Matthew R. , Lalmansingh Jared M. , Posey Ammon E. , Seim Ian , Gladfelter Amy S. , Pappu Rohit V.
Published: January 2023

Macromolecular phase separation underlies the regulated formation and dissolution of biomolecular condensates. What is unclear is how condensates of distinct and shared macromolecular compositions form and coexist within cellular milieus. Here, we use theory and computation to establish thermodynamic criteria that must be satisfied to achieve compositionally distinct condensates. We applied these criteria to an archetypal ribonucleoprotein condensate and discovered that demixing into distinct protein-RNA condensates cannot be the result of purely thermodynamic considerations. Instead, demixed, compositionally distinct condensates arise due to asynchronies in timescales that emerge from differences in long-lived protein-RNA and RNA-RNA crosslinks. This type of dynamical control is also found to be active in live cells whereby asynchronous production of molecules is required for realizing demixed protein-RNA condensates. We find that interactions that exert dynamical control provide a versatile and generalizable way to influence the compositions of coexisting condensates in live cells.
How random is the review outcome? A systematic study of the impact of external factors oneLifepeer review
Authors: Liang Weixin , Mahowald Kyle , Raymond Jennifer , Krishna Vamshi , Smith Daniel , Jurafsky Daniel , McFarland Daniel , Zou James
Published: January 2023

The advance of science rests on a robust peer review process. However whether or not a paper is accepted can depend on random external factors--e.g. the timing of the submission, the matching of editors and reviewers--that are beyond the quality of the work. This article systematically investigates the impact of these random factors independent of the paper’s quality on peer review outcomes in a major biomedical journal,eLife. We analyzed all of the submissions toeLifebetween 2016 to 2018, with 23,190 total submissions. We examined the effects of random factors at each decision point in the review process, from the gatekeeping senior editors who may desk-reject papers to review editors and reviewers who recommend the final outcome. Our results suggest that the peer-review process ineLifeis robust overall and that random external factors have relatively little quantifiable bias.
In vitroALPK1 kinase assay reveals new insights into ADP-heptose sensing pathway and kinase activity of disease-associated ALPK1 mutants
Authors: García-Weber Diego , Dangeard Anne-Sophie , Teixeira Veronica , Hauke Martina , Carreaux Alexis , Josenhans Christine , Arrieumerlou Cécile
Published: January 2023

Alpha-protein kinase 1 (ALPK1) is a pathogen recognition receptor that detects ADP-heptose (ADPH), a lipopolysaccharide biosynthesis intermediate, recently described as a pathogen-associated molecular pattern in Gram-negative bacteria. ADPH binding to ALPK1 activates its kinase domain and triggers TIFA phosphorylation on threonine 9. This leads to the assembly of large TIFA oligomers called TIFAsomes, activation of NF-κB and pro-inflammatory gene expression. Furthermore, mutations inALPK1are associated with inflammatory syndromes and cancers. While this kinase is of growing medical interest, only few tools are available to characterize its activity. Here, we describe a versatile non-radioactive ALPK1in vitrokinase assay based on protein thiophosphorylation. We confirm that ALPK1 phosphorylates TIFA T9 and show that T2, T12 and T19 are also weakly phosphorylated by ALPK1. Interestingly, we find that ALPK1 itself is phosphorylated in response to ADPH recognition duringShigella flexneriandHelicobacter pyloriinfection and that disease-associated ALPK1 mutants exhibit altered kinase activity. In particular, T237M and V1092A mutations associated with ROSAH syndrome and spiradenoma/spiradenocarcinoma respectively, exhibit enhanced ADPH-induced kinase activity and constitutive assembly of TIFAsomes. Altogether, this study provides new insights into the ADPH sensing pathway and a new tool for measuring the activity of ALPK1.
FAXC Proteins of Vertebrates and Invertebrates: Relationship to Metaxin Proteins
Authors: Adolph Kenneth W.
Published: January 2023

FAXC proteins of vertebrates and invertebrates are shown to possess structural features similar to the metaxins, and can therefore be categorized as metaxin-like proteins. A variety of vertebrate and invertebrate species were found to encode FAXC (Failed Axon Connections) proteins. These are predicted proteins. Among the vertebrate species are human, mouse, zebrafish, andXenopus. For invertebrates, FAXC proteins were most common in phyla that include Mollusca, Arthropoda, Cnidaria, and Placozoa. The presence of characteristic GST_N_Metaxin and GST_C_Metaxin protein domains was an important criterion in identifying the FAXCs as metaxin-like. In addition, the Tom37 domain was frequently found, especially in vertebrates. Also important was the possession of segments of α-helical secondary structure in a pattern of eight helices like that of metaxin 1 and other metaxins. Both the domain structures and α-helical structures are characteristic metaxin-like features. But alignment of protein sequences of both vertebrate and invertebrate FAXCs with metaxin proteins demonstrated that FAXCs and metaxins have low percentages of identical amino acids, about 20%. Therefore, FAXCs and metaxins are related, but are not the same protein. As revealed by phylogenetic analysis, vertebrate FAXCs and vertebrate metaxins 1, 2, and 3 constitute separate but related groups of proteins. Distinct phylogenetic groups were also observed for invertebrate FAXCs and invertebrate metaxins 1 and 2. The FAXC genes have neighboring genes that differ from the neighboring genes of metaxins. For vertebrates, the genes adjacent to FAXC genes are highly conserved. For invertebrates, the genes that neighbor FAXC genes are not the same as for vertebrates, but are conserved among different invertebrates. Invertebrates can have multiple FAXC genes, up to at least 12 or 13 genes. In some cases, the multiple FAXC genes exist as a cluster of genes in close proximity.
Single-cell atlases of two lophotrochozoan larvae highlight their complex evolutionary histories
Authors: Piovani Laura , Leite Daniel J. , Yañez Guerra Luis Alfonso , Simpson Fraser , Musser Jacob M. , Salvador-Martínez Irepan , Marlétaz Ferdinand , Jékely Gáspár , Telford Maximilian J.
Published: January 2023

Pelagic larval stages are widespread across animals, yet it is unclear if larvae were present in the last common ancestor of animals or whether they evolved multiple times due to common selective pressures. Many marine larvae are at least superficially similar, they are small, swim through beating of ciliated bands and sense the environment with an apical organ structure. To understand these similarities, we have generated single cell atlases for marine larvae from two animal phyla and have compared their cell types. We found clear similarities among ciliary band cells and neurons of the apical organ in the two larvae pointing to possible homology of these structures suggesting a single origin of larvae within the clade analysed here (Lophotrochozoa). We also find several clade specific innovations in each larva, including distinct myocytes and shell gland cells in the oyster larva. Oyster shell gland cells express many novel genes which have made previous gene age estimates for trochophore larvae too young.
Endophytic fungi related to the ash dieback causal agent encode signatures of pathogenicity on European ash
Authors: Rafiqi Maryam , Kosawang Chatchai , Peers Jessica A. , Jelonek Lukas , Yvanne Hélène , McMullan Mark , Nielsen Lene R.
Published: January 2023

Tree diseases constitute a significant threat to biodiversity worldwide. Pathogen discovery in natural habitats is of vital importance to understanding current and future threats and prioritising efforts towards developing disease management strategies. Ash dieback is a fungal disease of major conservational concern that is infecting common ash trees, Fraxinus excelsior, in Europe. The disease is caused by a non-native fungal pathogen, Hymenoscyphus fraxineus. Other dieback causing-species have not previously been identified in the genus Hymenoscyphus. Here, we discover the pathogenicity potential of two newly identified related species of Asian origin, H. koreanus and H. occultus, and one Europe-native related species, H. albidus. We sequence the genomes of all three Hymenoscyphus species and compare them to that of H. fraxineus. Phylogenetic analysis of core eukaryotic genes identified H. albidus and H. koreanus as sister species, whilst H. occultus diverged prior these and H. fraxineus. All four Hymenoscyphus genomes are of comparable sizes (55-62 Mbp) and GC contents (42–44%) and encode for polymorphic secretomes. Surprisingly, 1,133 predicted secreted proteins are shared between the ash dieback pathogen H. fraxineus and the three related Hymenoscyphus endophytes. Amongst shared secreted proteins are cell death-inducing effector candidates, such as necrosis, and ethylene-inducing peptide 1-like proteins, NLPs, that are upregulated during in planta growth of all Hymenoscyphus species. Indeed, pathogenicity tests showed that all four related Hymenoscyphus species develop pathogenic growth on European ash stems, with native H. albidus being the least virulent. Our results identify the threat Hymenoscypohus species pose to the survival of European ash trees, and highlight the importance of promoting pathogen surveillance in environmental landscapes. Identifying new pathogens and including them in the screening for durable immunity of common ash trees is key to the long-term survival of ash.
Disruption of the standard kinetochore in holocentricCuscutaspecies
Authors: Pavel Neumann , Oliveira Ludmila , Jang Tae-Soo , Novák Petr , Koblížková Andrea , Schubert Veit , Houben Andreas , Macas Jiří
Published: January 2023

Segregation of chromosomes depends on the centromere. Most species are monocentric, with the centromere restricted to a single region per chromosome. In some organisms, monocentric organization changed to holocentric, in which the centromere activity is distributed over the entire chromosome length. However, the causes and consequences of this transition are poorly understood. Here, we show that the transition in the genusCuscutawas associated with dramatic changes in the kinetochore, a protein complex that mediates the attachment of chromosomes to microtubules. We found that in holocentricCuscutaspecies the KNL2 genes were lost; the CENP-C, KNL1, and ZWINT1 genes were truncated; the centromeric localization of CENH3, CENP-C, KNL1, MIS12, and NDC80 proteins was disrupted; and the spindle assembly checkpoint (SAC) was degenerated. Our results demonstrate that holocentricCuscutaspecies lost the ability to form a standard kinetochore and do not employ SAC to control the attachment of microtubules to chromosomes.
MitoQuicLy: a high-throughput method for quantifying cell-free DNA from human plasma, serum, and saliva
Authors: Michelson Jeremy , Rausser Shannon , Peng Amanda , Yu Temmie , Sturm Gabriel , Trumpff Caroline , Kaufman Brett A. , Rai Alex J. , Picard Martin
Published: January 2023

Circulating cell-free mitochondrial DNA (cf-mtDNA) is an emerging biomarker of psychobiological stress and disease which predicts mortality and is associated with various disease states. To evaluate the contribution of cf-mtDNA to health and disease states, standardized high-throughput procedures are needed to quantify cf-mtDNA in relevant biofluids. Here, we describe MitoQuicLy:Mitochondrial DNAQuantification incell-free samples byLysis. We demonstrate high agreement between MitoQuicLy and the commonly used column-based method, although MitoQuicLy is faster, cheaper, and requires a smaller input sample volume. Using 10 µL of input volume with MitoQuicLy, we quantify cf-mtDNA levels from three commonly used plasma tube types, two serum tube types, and saliva. We detect, as expected, significant inter-individual differences in cf-mtDNA across different biofluids. However, cf-mtDNA levels between concurrently collected plasma, serum, and saliva from the same individual differ on average by up to two orders of magnitude and are poorly correlated with one another, pointing to different cf-mtDNA biology or regulation between commonly used biofluids in clinical and research settings. Moreover, in a small sample of healthy women and men (n=34), we show that blood and saliva cf-mtDNAs correlate with clinical biomarkers differently depending on the sample used. The biological divergences revealed between biofluids, together with the lysis-based, cost-effective, and scalable MitoQuicLy protocol for biofluid cf-mtDNA quantification, provide a foundation to examine the biological origin and significance of cf-mtDNA to human health.
Loss of NR5A1 in Sertoli cells after sex determination changes their cellular identity and induces their death by anoikis
Authors: Souali-Crespo Sirine , Condrea Diana , Vernet Nadège , Féret Betty , Klopfenstein Muriel , Grandgirard Erwan , Alunni Violaine , Cerciat Marie , Jung Matthieu , Mayere Chloé , Nef Serge , Mark Manuel , Chalmel Frédéric , Ghyselinck Norbert B.
Published: January 2023

NR5A1 is an orphan nuclear receptor crucial for gonadal development in mammals. In the mouse testis it is expressed both in Sertoli cells (SC) and Leydig cells (LC). To investigate its role posteriorly to sex determination, we have generated and analysed mice lacking NR5A1 in SC from embryonic day (E) 13.5 onwards (Nr5a1SC−/−mutants). Ablation ofNr5a1impairs the expression of genes characteristic of SC identity (e.g.,Sox9, Amh), makes SC to progressively die from E14.5 by aTrp53-independent mechanism, and induces disorganization of the testis cords, which, together, yields germ cells (GC) to prematurely enter meiosis and die, instead of becoming quiescent. Single-cell RNA-sequencing experiments revealed thatNr5a1-deficient SC acquire a pre-granulosa cell-like identity, and profoundly modify the landscape of the adhesion molecules and extracellular matrix they express. We propose therefore that SC lacking NR5A1 transdifferentiate and die by anoikis. Fetal LC do not display major changes in their transcriptome, indicating that SC are not required beyond E14.5 for their emergence or maintenance. In contrast, adult LC were missing inNr5a1SC−/−postnatal testes. In addition, adult males display Müllerian duct derivatives (i.e., uterus, vagina), as well as a decreased anogenital distance and a shorter penis that can be explained by loss of AMH production and defective HSD17B1- and HSD17B3-mediated synthesis of testosterone in SC during fetal life. Together, our findings indicate thatNr5a1expressed in SC after the period of sex determination safeguards SC identity, which maintains proper seminiferous cord organization and prevents GC to enter meiosis.
Human Brain Endothelial Cell Seeded on Inner Surface of Alginate Hollow Microfibers
Authors: Aykar Saurabh S. , Alimoradi Nima , Petersen Isaac S. , Montazami Reza , Brockman Amanda L. , Hashemi Nicole N.
Published: January 2023

Barrier functionality of the blood-brain barrier (BBB) is provided by the tight junctions formed by a monolayer of the human brain endothelial cells (HBECs) internally around the blood capillaries. To mimic such barrier functionality in vitro, replicating the hollow tubular structure of the BBB along with the HBECs monolayer on its inner surface is crucial. Here, we developed an invasive microfluidic technique to obtain the HBECs monolayer on the inner surface of alginate-based hollow microfibers. The HBECs were seeded on the inner surface of these microfibers using a custom-built microfluidic device. The seeded HBECs were monitored for 9 days after manufacturing and cultured to form a monolayer on the inner surface of the alginate hollow microfibers in the maintenance media. A higher cell seeding density of 217 cells/mm length of the hollow microfiber was obtained using our microfluidic technique. Moreover, high accuracy of around 96 % was obtained in seeding cells on the inner surface of alginate hollow microfibers. The microfluidic method illustrated in this study could be extrapolated to obtain a monolayer of different cell types on the inner surface of alginate hollow microfibers with cell-compatible ECM matrix proteins. Furthermore, it will enable us to mimic a range of microvascular systems in vitro by closely replicating the structural attributes of the native structure.
Urban birds become less fearful following COVID-19 reopenings
Authors: Diamant Eleanor S. , MacGregor-Fors Ian , Blumstein Daniel T. , Yeh Pamela J.
Published: January 2023

Following the COVID-19 pandemic, many people around the world stayed home, drastically altering human activity in cities. This exceptional moment provided researchers the opportunity to test how urban animals respond to human disturbance, in some cases testing fundamental questions on the mechanistic impact of urban behaviors on animal behavior. However, at the end of this “anthropause,” human activity returned to cities. How might each of these strong shifts affect wildlife in the short and long term? We focused on fear response, a trait essential to tolerating urban life. We measured flight initiation distance—at both individual and population-levels—for an urban bird before, during, and after the anthropause to examine if birds experienced longer-term changes after a year of lowered human presence. Dark-eyed juncos did not change fear levels during the anthropause, but they became drastically less fearful afterwards. These surprising and counter-intuitive findings, made possible by following the behavior of individuals over time, has led to a novel understanding that fear response can be driven by plasticity, yet not habituation-like processes. The pandemic-caused changes in human activity have shown that there is great complexity in how humans modify a behavioral trait fundamental to urban tolerance in animals.
High Throughput Viability Assay for Microbiology
Authors: Meyer Christian T. , Lynch Grace K. , Stamo Dana F. , Miller Eugene J. , Chatterjee Anushree , Kralj Joel M.
Published: January 2023

Counting viable cells is a universal practice in microbiology. The colony forming unit (CFU) assay has remained the gold standard to measure viability across disciplines; however, it is time-intensive and resource-consuming. Herein, we describe the Geometric Viability Assay (GVA) that replicates CFU measurements over 6-orders of magnitude while reducing over 10-fold the time and consumables. GVA computes a sample’s viable cell count based on the distribution of embedded colonies growing inside a pipette tip. GVA is compatible with gram-positive and -negative planktonic bacteria, biofilms, and yeast. Laborious CFU experiments such as checkerboard assays, treatment time-courses, and drug screens against slow-growing cells are simplified by GVA. We therefore screened a drug library against exponential and stationary phaseE. colileading to the discovery of the ROS-mediated, bactericidal mechanism of diphenyliodonium. The ease and low cost of GVA evinces it can accelerate existing viability assays and enable measurements at previously impractical scales.
Phylogenomics reveals patterns of ancient hybridization and differential diversification contributing to phylogenetic conflict inPopulusL. andSalixL
Authors: Sanderson Brian J. , Ghambir Diksha , Feng Guanqiao , Hu Nan , Cronk Quentin C. , Percy Diana M. , Freaner Francisco Molina , Johnson Matthew G. , Smart Lawrence B. , Keefover-Ring Ken , Yin Tongming , Ma Tao , DiFazio Stephen P. , Liu Jianquan , Olson Matthew S.
Published: January 2023

Despite the economic, ecological, and scientific importance of the generaPopulusL. (poplars, cottonwoods, and aspens) andSalixL. Salicaceae (willows), we know little about the sources of differences in species diversity between the genera and of the phylogenetic conflict that often confounds estimating phylogenetic trees.Salixsubgenera and sections, in particular, have been difficult to classify, with one recent attempt termed a ‘spectacular failure’ due to a speculated radiation of the subgeneraVetrixandChamaetia. Here we use targeted sequence capture to understand the evolutionary history of this portion of the Salicaceae plant family. Our phylogenetic hypothesis was based on 787 gene regions and identified extensive phylogenetic conflict among genes. Our analysis supported some previously described subgeneric relationships and confirmed polyphyly of others. Using an fbranchanalysis we identified several cases of hybridization in deep branches of the phylogeny, which likely contributed to discordance among gene trees. In addition, we identified a rapid increase in diversification rate near the origination of theVetrix-Chamaetiaclade inSalix. This region of the tree coincided with several nodes that lacked strong statistical support, indicating a possible increase in incomplete lineage sorting due to rapid diversification. The extraordinary level of both recent and ancient hybridization in bothPopulusandSalixhave played important roles in the diversification and diversity in these two genera.

Supplementary data files will be provided by request tomatt.olson@ttu.edu
Vitamin D inhibits osteosarcoma by reprogramming nonsense-mediated RNA decay and SNAI2-mediated epithelial-to-mesenchymal transition
Authors: Capobianco Enrico , McGaughey Vanessa , Seraphin Gerbenn , Heckel John , Rieger Sandra , Lisse Thomas S.
Published: January 2023

Osteosarcomas are immune-resistant and metastatic as a result of elevated nonsense-mediated RNA decay (NMD), reactive oxygen species (ROS), and epithelial-to-mesenchymal transition (EMT). Although vitamin D has anti-cancer effects, its effectiveness and mechanism of action against osteosarcomas are poorly understood. In this study, we assessed the impact of vitamin D and its receptor (VDR) on the NMD-ROS-EMT signaling axis inin vitroandin vivoosteosarcoma animal models. Initiation of VDR signaling facilitated the enrichment of EMT pathway genes, after which 1,25(OH)2D, the active vitamin D derivative, inhibited the EMT pathway in osteosarcoma subtypes. The ligand-bound VDR directly downregulated the EMT inducerSNAI2, differentiating highly metastatic from low metastatic subtypes and 1,25(OH)2D sensitivity. Moreover, epigenome-wide motif and putative target gene analysis revealed the VDR’s integration with NMD tumorigenic and immunogenic pathways. In an autoregulatory manner, 1,25(OH)2D inhibited NMD machinery genes and upregulated NMD target genes implicated in anti-oncogenic activity, immunorecognition, and cell-to-cell adhesion. Dicer substrate siRNA knockdown ofSNAI2revealed superoxide dismutase 2 (SOD2)-mediated antioxidative responses and 1,25(OH)2D sensitization via non-canonical SOD2 nuclear-to-mitochondrial translocalization leading to overall ROS suppression. In a mouse xenograft metastasis model, the therapeutically relevant vitamin D derivative calcipotriol inhibited osteosarcoma metastasis and tumor growth shown for the first time. Our results uncover novel osteosarcoma-inhibiting mechanisms for vitamin D and calcipotriol that may be translated to human patients.
Behavioral and Neuroanatomical Investigation of Attention Deficit Hyperactivity Disorder Pathogenesis in Juvenile Spontaneously Hypertensive Rats
Authors: Aydin Aysegul Gungor , Adiguzel Esat
Published: January 2023

Attention-deficit/hyperactivity disorder (ADHD) is one of the most prevalent neuropsychiatric disorders of childhood, characterized by locomotor hyperactivity, impaired sustained attention, impulsivity, and distractibility. Recently, the dysfunction of different synaptic circuits in the prefrontal cortex (PFC) has been shown. Previous studies have attributed the pathophysiological mechanism of ADHD to disturbances in the dopaminergic system. In this study, we tested the hypothesis that spontaneously hypertensive rats (SHR), which are considered a validated animal model of ADHD, have altered dopaminergic innervation and increased locomotor activity. Here, we performed immunohistochemical tyrosine hydroxylase (TH) and dopamine-beta-hydroxylase (DBH) staining. The mesocortical dopaminergic system appears to be normal in juvenile SHR, as suggested by (i) no alteration in the area density of TH-immunoreactive (TH-ir) dopaminergic neurons in the VTA, (ii) no alterations in the volume density of TH-ir fibers in layer I of the PrL subregion of mPFC, (iii) no alteration in the percentage of TH-ir dopaminergic fibers in layer I of the PrL subregion of mPFC as revealed by TH and/or DBH immunoreactivity. Furthermore, the SHR showed increased locomotor activity than WKY in the open field test.

The demonstration of no alteration in mesocortical dopaminergic neurons and fiber in SHR raises some concern about the position of SHR as an animal model of the inattentive subtype of ADHD. However, these results strengthen this strain as an animal model of hyperactive/impulsive subtype ADHD for future studies that may elucidate the underlying mechanism mediating hyperactivity and test various treatment strategies.