bluuebunny/crossref_metadata_2025 huggingface dataset:
Newer Surveillance Data Extends our Understanding of the Niche ofRickettsia montanensis(Rickettsiales: Rickettsiaceae) Infection of the American Dog Tick (Acari: Ixodidae) in the United States
Authors: Lippi Catherine A. , Gaff Holly D. , Nadolny Robyn M. , Ryan Sadie J.
Published: January 2023

Background

Understanding the geographic distribution ofRickettsia montanensisinfections inDermacentor variabilisis important for tick-borne disease management in the United States, as both a tick-borne agent of interest and a potential confounder in surveillance of other rickettsial diseases. Two previous studies modeled niche suitability forD. variabiliswith and withoutR. montanensis, from 2002-2012, indicating that theD. variabilisniche overestimates the infected niche. This study updates these, adding data since 2012.

Methods

Newer surveillance and testing data were used to update Species Distribution Models (SDMs) ofD. variabilis, andR. montanensisinfectedD. variabilis, in the United States. Using random forest (RF) models, found to perform best in previous work, we updated the SDMs and compared them with prior results. Warren’s I niche overlap metric was used to compare between predicted suitability for all ticks and ‘pathogen positive niche’ models across datasets.

Results

Warren’s I indicated <2% change in predicted niche, and there was no change in order of importance of environmental predictors, forD. variabilisorR. montanensispositive niche. The updatedD. variabilisniche model overpredicted suitability compared to the updatedR. montanensispositive niche in key peripheral parts of the range, but slightly underpredicted through the northern and midwestern parts of the range. This reinforces previous findings of a more constrained pathogen-positive niche than predicted byD. variabilisrecords alone.

Conclusions

The consistency of predicted niche suitability forD. variabilisin the United States, with the addition of nearly a decade of new data, corroborates this is a species with generalist habitat requirements. Yet a slight shift in updated niche distribution, even of low suitability, included more southern areas, pointing to a need for continued and extended monitoring and surveillance. This further underscores the importance of revisiting vector and vector-borne disease distribution maps.
SCS: cell segmentation for high-resolution spatial transcriptomics
Authors: Chen Hao , Li Dongshunyi , Bar-Joseph Ziv
Published: January 2023

Spatial transcriptomics promises to greatly improve our understanding of tissue organization and cell-cell interactions. While most current platforms for spatial transcriptomics only offer multi-cellular resolution, with 10-15 cells per spot, recent technologies provide a much denser spot placement leading to sub-cellular resolution. A key challenge for these newer methods is cell segmentation and the assignment of spots to cells. Traditional image-based segmentation methods are limited and do not make full use of the information profiled by spatial transcrip-tomics. Here we present SCS, which combines imaging data with sequencing data to improve cell segmentation accuracy. SCS assigns spots to cells by adaptively learning the position of each spot relative to the center of its cell using a transformer neural network. SCS was tested on two new sub-cellular spatial transcriptomics technologies and outperformed traditional image-based segmentation methods. SCS achieved better accuracy, identified more cells, and provided more realistic cell size estimation. Sub-cellular analysis of RNAs using SCS spots assignments provides information on RNA localization and further supports the segmentation results.
Fast, accurate ranking of engineered proteins by receptor binding propensity using structure modeling
Authors: Ding Xiaozhe , Chen Xinhong , Sullivan Erin E. , Shay Timothy F. , Gradinaru Viviana
Published: January 2023

Deep learning-based methods for protein structure prediction have achieved unprecedented accuracy. However, the power of these tools to guide the engineering of protein-based therapeutics remains limited due to a gap between the ability to predict the structures of candidate proteins and the ability to assess which of those proteins are most likely to bind to a target receptor. Here we bridge this gap by introducing Automated Pairwise Peptide-Receptor AnalysIs for Screening Engineered proteins (APPRAISE), a method for predicting the receptor binding propensity of engineered proteins. After generating models of engineered proteins competing for binding to a target using an established structure-prediction tool such as AlphaFold-Multimer or ESM-Fold, APPRAISE performs a rapid (under 1 CPU second per model) scoring analysis that takes into account biophysical and geometrical constraints. As a proof-of-concept, we demonstrate that APPRAISE can accurately classify receptor-dependent vs. receptor-independent adeno-associated viral vectors and diverse classes of engineered proteins such as miniproteins targeting the SARS-CoV-2 spike, nanobodies targeting a G-protein-coupled receptor, and peptides that specifically bind to transferrin receptor or PD-L1. APPRAISE is accessible through a web-based notebook interface using Google Colaboratory (https://tiny.cc/APPRAISE). With its accuracy, interpretability, and generalizability, APPRAISE promises to expand the utility of protein structure prediction and accelerate protein engineering for biomedical applications.
Immature leaves are the dominant volatile sensing organs of maize
Authors: Wang Lei , Jäggi Simon , Walthert Mario , Waterman Jamie M. , Cofer Tristan M. , Erb Matthias
Published: January 2023

Plants perceive herbivory induced volatiles and respond to them by upregulating their defenses. So far, the organs responsible for volatile perception remain poorly described. Here, we show that responsiveness to the herbivory induced green leaf volatile (Z)-3-hexenyl acetate (HAC) in terms of volatile emission, transcriptional regulation and defense hormone activation is largely constrained to younger maize leaves. Older leaves are much less sensitive to HAC. In a given leaf, responsiveness to HAC is high at immature developmental stages and drops off rapidly during maturation. Responsiveness to the non-volatile elicitor ZmPep3 shows an opposite pattern, demonstrating that hyposmia is not driven by defective canonical defense signaling. Neither stomatal conductance nor leaf cuticle composition explain the unresponsiveness of older leaves to HAC, suggesting perception mechanisms upstream of canonical defense signaling as driving factors. Finally, we show that hyposmia in older leaves is not restricted to HAC, and extends to the full blend of herbivory induced volatiles. In conclusion, our work identifies immature maize leaves as dominant stress volatile sensing organs. The tight spatiotemporal control of volatile perception may facilitate within-plant defense signaling to protect young leaves, and may allow plants with complex architectures to explore the dynamic odor landscapes at the outer periphery of their shoots.
Galectin-9 has non-apoptotic cytotoxic activity towards Acute Myeloid Leukemia independent of cytarabine resistance
Authors: Choukrani Ghizlane , Visser Nienke , Avtenyuk Natasha Ustyanovska , Olthuis Mirjam , Marsman Glenn , Ammatuna Emanuele , Lourens Harm Jan , Niki Toshiro , Huls Gerwin , Bremer Edwin , Wiersma Valerie R.
Published: January 2023

Acute myeloid leukemia (AML) is a malignancy still associated with poor survival rates, among others due to frequent occurrence of therapy-resistant relapse after standard-of-care treatment with cytarabine (AraC). AraC triggers apoptotic cell death, a type of cell death to which AML cells often become resistant. Therefore, therapeutic options that trigger an alternate type of cell death are of particular interest. We previously identified that the glycan-binding protein Galectin-9 (Gal-9) has tumor-selective and non-apoptotic cytotoxicity towards various types of cancer, which depended on autophagy inhibition. Thus, Gal-9 could be of therapeutic interest for (AraC-resistant) AML. In the current study, treatment with Gal-9 was cytotoxic for AML cells, including for CD34+patient-derived AML stem cells, but not for healthy cord blood-derived CD34+stem cells. This Gal-9-mediated cytotoxicity did not rely on apoptosis but negatively associated with autophagic flux. Importantly, both AraC-sensitive and -resistant AML cell lines as well as AML patient samples were sensitive to single agent treatment with Gal-9. Additionally, Gal-9 potentiated the cytotoxic effect of DNA demethylase inhibitor Azacytidine (Aza), a drug that is clinically used for patients that are not eligible for intensive AraC treatment. Thus, Gal-9 is a potential therapeutic agent for the treatment of AML, including AraC resistant AML, by inducing caspase-independent cell death.
High Throughput Sequencing technologies complemented by grower’s perception highlight the impact of tomato virome in diversified vegetable farms
Authors: Temple Coline , Blouin Arnaud G. , Tindale Sophie , Steyer Stephan , Marechal Kevin , Massart Sebastien
Published: January 2023

The number of small-scale diversified vegetable growers in industrialized countries has risen sharply over the last ten years. The risks associated with plant viruses in these systems have been barely studied in Europe, yet dramatic virus emergence events, such as tomato brown fruit rugose virus, sometimes occur. We developed a methodology that aimed to understand better the implications related to viruses for tomato production in Belgian’s vegetable farms by comparing growers’ perception of the presence of viral symptoms (visual inspection) with non targeting detection of nearly all viruses present in the plants by high throughput sequencing technologies (HTS). Virus presence and impact were interpreted considering the farm’s typology and cultural practices, the grower’s professional profiles, and visual inspection of plant-viral-like symptoms. Overall, The data indicated that most growers have limited understanding of tomato viruses and are not concerned about them. Field observations were correlated to this perception as the prevalence of symptomatic plants was usually lower than 1%. However, important and potentially emergent viruses, mainly transmitted by insects, were detected in several farms. Noteworthy, the presence of these viruses was correlated with the number of plant species grown per site (diversity) but not with a higher awareness of the growers regarding plant viral diseases or a higher number of symptomatic plants. In addition, both HTS and perception analysis underlined the rising incidence and importance of an emergent virus: Physostegia chlorotic mottle virus. Overall, the original methodology developed here, combining social science with HTS technologies, could be applied to other crops in other systems to identify emergent risks associated with plant viruses and can highlight the communication needed toward growers to mitigate epidemics.
Grapevine scion gene expression is driven by rootstock and environment interaction
Authors: Harris Zachary N , Pratt Julia E , Kovacs Laszlo G , Klein Laura L , Kwasniewski Misha T. , Londo Jason P , Wu Angela , Miller Allison J
Published: January 2023

BACKGROUND

Grafting is a horticultural practice used widely across woody perennial crop species to fuse together the root and shoot system of two distinct genotypes, the rootstock and the scion, combining beneficial traits from both. In grapevine, grafting is used in nearly 80% of all commercial vines to optimize fruit quality, regulate vine vigor, and enhance biotic and abiotic stress-tolerance. Rootstocks have been shown to modulate elemental composition, metabolomic profiles, and the shape of leaves in the scion, among other traits. However, it is currently unclear how rootstock genotypes influence shoot system gene expression as previous work has reported complex and often contradictory findings.

RESULTS

In the present study, we examine the influence of grafting on scion gene expression in leaves and reproductive tissues of grapevines growing under field conditions for three years. We show that the influence from the rootstock genotype is highly tissue and time dependent, manifesting only in leaves, primarily during a single year of our three-year study. Further, the degree of rootstock influence on scion gene expression is driven by interactions with the local environment.

CONCLUSIONS

Our results demonstrate that the role of rootstock genotype in modulating scion gene expression is not a consistent, unchanging effect, but rather an effect that varies over time in relation to local environmental conditions.
Automated quantitative trait locus analysis (AutoQTL)
Authors: Freda Philip J. , Ghosh Attri , Zhang Elizabeth , Luo Tianhao , Chitre Apurva , Polesskaya Oksana , St. Pierre Celine L. , Gao Jianjun , Martin Connor D. , Chen Hao , Garcia-Martinez Angel G. , Wang Tengfei , Han Wenyan , Ishiwari Keita , Meyer Paul , Lamparelli Alexander , King Christopher P. , Palmer Abraham A. , Li Ruowang , Moore Jason H.
Published: January 2023

Background

Quantitative Trait Locus (QTL) analysis and Genome-Wide Association Studies (GWAS) have the power to identify variants that capture significant levels of phenotypic variance in complex traits. However, effort and time are required to select the best methods and optimize parameters and pre-processing steps. Although machine learning approaches have been shown to greatly assist in optimization and data processing, applying them to QTL analysis and GWAS is challenging due to the complexity of large, heterogenous datasets. Here, we describe proof-of-concept for an automated machine learning approach, AutoQTL, with the ability to automate many complex decisions related to analysis of complex traits and generate diverse solutions to describe relationships that exist in genetic data.

Results

Using a dataset of 18 putative QTL from a large-scale GWAS of body mass index in the laboratory rat,Rattus norvegicus, AutoQTL captures the phenotypic variance explained under a standard additive model while also providing evidence of non-additive effects including deviations from additivity and 2-way epistatic interactions from simulated data via multiple optimal solutions. Additionally, feature importance metrics provide different insights into the inheritance models and predictive power of multiple GWAS-derived putative QTL.

Conclusions

This proof-of-concept illustrates that automated machine learning techniques can be applied to genetic data and has the potential to detect both additive and non-additive effects via various optimal solutions and feature importance metrics. In the future, we aim to expand AutoQTL to accommodate omics-level datasets with intelligent feature selection strategies.
Fitting agent-based models to tumor images using representation learning
Authors: Cess Colin G. , Finley Stacey D.
Published: January 2023

Agent-based models (ABMs) have enabled great advances in the study of tumor development and therapeutic response, allowing researchers to explore the spatiotemporal evolution of the tumor and its microenvironment. However, these models face serious drawbacks in the realm of parameterization – ABM parameters are typically set individually based on various data and literature sources, rather than through a rigorous parameter estimation approach. While ABMs can be fit to time-course data, that type of data loses the spatial information that is a defining feature of ABMs. Tumor images provide spatial information; however, such images only represent individual timepoints, limiting their utility in calibrating the tumor dynamics predicted by ABMs. Furthermore, it is exceedingly difficult to compare tumor images to ABM simulations beyond a qualitative visual comparison. Without a quantitative method of comparing the similarity of tumor images to ABM simulations, a rigorous parameter fitting is not possible. Here, we present a novel approach that applies neural networks to represent both tumor images and ABM simulations as low dimensional points, with the distance between points acting as a quantitative measure of difference between the two. This enables a quantitative comparison of tumor images and ABM simulations, where the distance between simulated and experimental images can be minimized using standard parameter-fitting algorithms. Here, we describe this method and present two examples to demonstrate the application of the approach to estimate parameters for two distinct ABMs. Overall, we provide a novel method to robustly estimate ABM parameters.
Conserved Roles for the Dynein Intermediate Chain and Ndel1 in Assembly and Activation of Dynein
Authors: Okada Kyoko , Iyer Bharat R. , Lammers Lindsay G. , Gutierrez Pedro , Li Wenzhe , Markus Steven M. , McKenney Richard J.
Published: January 2023

Cytoplasmic dynein, the primary retrograde microtubule transport motor within cells, must be activated for processive motility through the regulated assembly of a dynein-dynactin-adapter (DDA) complex. The interaction between dynein and dynactin was initially ascribed to the N-terminus of the dynein intermediate chain (IC) and a coiled-coil of the dynactin subunit p150Glued. However, cryo-EM structures of DDA complexes have not resolve these regions of the IC and p150Glued, raising questions about the importance of this interaction. The IC N-terminus (ICN) also interacts with the dynein regulators Nde1/Ndel1, which compete with p150Gluedfor binding to ICN. Using a combination of approaches, we reveal that the ICN plays critical, evolutionarily conserved roles in DDA assembly by interacting with dynactin and Ndel1, the latter of which recruits the DDA assembly factor LIS1 to the dynein complex. In contrast to prior models, we find that LIS1 cannot simultaneously bind to Ndel1 and dynein, indicating that LIS1 must be handed off from Ndel1 to dynein in temporally discrete steps. Whereas exogenous Ndel1 or p150Glueddisrupts DDA complex assemblyin vitro, neither perturbs preassembled DDA complexes, indicating that the IC is stably bound to p150Gluedwithin activated DDA complexes. Our study reveals previously unknown regulatory steps in the dynein activation pathway, and provides a more complete model for how the activities of LIS1/Ndel1 and dynactin/cargo-adapters are integrated to regulate dynein motor activity.
Pervasive GxE interactions shape adaptive trajectories and the exploration of the phenotypic space in artificial selection experiments
Authors: Desbiez-Piat Arnaud , Ressayre Adrienne , Marchadier Elodie , Noly Alicia , Remoué Carine , Vitte Clémentine , Belcram Harry , Bourgais Aurélie , Galic Nathalie , Le Guilloux Martine , Tenaillon Maud I. , Dillmann Christine
Published: January 2023

Quantitative genetics models have shown that long-term selection responses depend on initial variance and mutational influx. Understanding limits of selection requires quantifying the role of mutational variance. However, correlative responses to selection on non-focal traits can perturb the selection response on the focal trait; and generations are often confounded with selection environments so that genotype by environment (GxE) interactions are ignored. The Saclay Divergent Selection Experiments (DSE) on maize flowering time were used to track the fate of individual mutations combining genotyping data and phenotyping data from yearly measurements (DSEYM) and common garden experiments (DSECG) with four objectives (1) to quantify the relative contribution of standing and mutational variance to the selection response, (2) to estimate genotypic mutation effects, (3) to study the impact of GxE interactions in the selection response, (4) to analyze how trait correlations modulate the exploration of the phenotypic space. We validated experimentally the expected enrichment of fixed beneficial mutations with an average effect of +0.278 and +0.299 days to flowering, depending on the genetic background. Fixation of unfavorable mutations reached up to 25% of incoming mutations, a genetic load possibly due to antagonistic pleiotropy, whereby mutations fixed in the selection environment (DSEYM) turned to be unfavorable in the evaluation environment (DSECG). Global patterns of trait correlations were conserved across genetic backgrounds but exhibited temporal patterns. Traits weakly or uncorrelated with flowering time triggered stochastic exploration of the phenotypic space, owing to microenvironment-specific fixation of standing variants and pleiotropic mutational input.
SUMO protease FUG1, histone reader AL3 and the PRC1 Complex are integral to repeat-expansion induced epigenetic silencing inArabidopsis thaliana
Authors: Sureshkumar Sridevi , Bandaranayake Champa , Lu Junqing , Dent Craig I , Atri Chhaya , York Harrison M , Tamizhselvan Prashanth , Shamaya Nawar , Folini Giulia , Bhagat Prakash Kumar , Bergey Benjamin G , Yadav Avilash Singh , Kumar Subhasree , Grummisch Oliver , Saini Prince , Yadav Ram K , Arumugam Senthil , Rosonina Emanuel , Sadanandom Ari , Liu Hongtao , Balasubramanian Sureshkumar
Published: January 2023

Epigenetic gene silencing induced by expanded repeats can cause diverse phenotypes ranging from severe growth defects in plants to genetic diseases such as Friedreich’s ataxia in humans1. The molecular mechanisms underlying repeat expansion-induced epigenetic silencing remain largely unknown2,3. Using a plant model, we have previously shown that expanded repeats can induce smallRNAs which in turn can lead to epigenetic silencing through the RNA-dependent DNA methylation pathway4,5. Here, using a genetic suppressor screen, we confirm a key role for the RdDM pathway and identify novel components required for epigenetic silencing caused by expanded repeats. We show that FOURTH ULP LIKE GENE CLASS 1 (FUG1) – a SUMO protease, ALFIN-LIKE 3 – a histone reader and LIKE HETEROCHROMATIN 1 (LHP1) - a component of the PRC1 complex are required for repeat expansion-induced epigenetic silencing. Loss of any of these components suppress repeat expansion-associated phenotypes. SUMO protease FUG1 physically interacts with AL3 and perturbing its potential SUMOylation site disrupts its nuclear localisation. AL3 physically interacts with LHP1 of the PRC1 complex and the FUG1-AL3-LHP1 module is essential to confer repeat expansion-associated epigenetic silencing. Our findings highlight the importance post-translational modifiers and histone readers in epigenetic silencing caused by repeat expansions.
Cost-effective DNA methylation profiling by FML-seq
Authors: Foley Joseph W. , Zhu Shirley X. , West Robert B.
Published: January 2023

Current methods for profiling DNA methylation require costly reagents, sequencing, or labor time. We introduce FML-seq, a sequencing library protocol that greatly reduces all these costs. Relative to other techniques tested on the same human cell lines, FML-seq produces similar measurements of absolute and differential cytosine methylation at a fraction of the price. FML-seq enables inexpensive, high-throughput experimental designs for large-scale epigenetics research projects.
Tobramycin adaptation alters the antibiotic susceptibility ofPseudomonas aeruginosaquorum sensing-null mutants
Authors: Abisado-Duque Rhea G. , Townsend Kade A. , Mckee Brielle M. , Woods Kathryn , Koirala Pratik , Holder Alexandra J. , Craddock Vaughn D. , Cabeen Matthew , Chandler Josephine R.
Published: January 2023

The opportunistic bacteriumPseudomonas aeruginosauses the LasR-I quorum sensing system to increase resistance to the aminoglycoside antibiotic tobramycin. Paradoxically,lasR-null mutants are commonly isolated from chronic human infections treated with tobramycin, suggesting there may be a mechanism allowing thelasR-null mutants to persist under tobramycin selection. We hypothesized that the effects of inactivatinglasRon tobramycin resistance might be dependent on the presence or absence of other gene mutations in that strain, a phenomenon known as epistasis. To test this hypothesis, we inactivatedlasRin several highly tobramycin-resistant isolates from long-term evolution experiments. We show that the effects of ΔlasRon tobramycin resistance are strain dependent, which is due to a single mutation in thefusA1gene encoding the translation elongation factor EF-G1A (G61A nucleotide substitution). ThefusA1G61A mutation confers a strong selective advantage to ΔlasRmutants under tobramycin treatment. The effects offusA1G61A on ΔlasR-dependent tobramycin resistance are dependent on the MexXY efflux pump and the MexXY regulator ArmZ. ThefusA1mutation also modulates ΔlasRmutant resistance to two other antibiotics, ciprofloxacin and ceftazidime. Our results provide a possible explanation for the emergence oflasR-null mutants in clinical isolates and illustrate the importance of epistatic gene interactions in the evolution of quorum sensing.
E4F1 and ZNF148 are transcriptional activators of the A57C and wildtypeTERTpromoter
Authors: Chua Boon Haow , Ferry Laure , Domrane Cecilia , Anuar Nurkaiyisah Zaal , Wittek Anna , Jha Sudhakar , Butter Falk , Tenen Daniel G. , Defossez Pierre-Antoine , Kappei Dennis
Published: January 2023

Point mutations within theTERTpromoter are the most recurrent somatic non-coding mutations identified across different cancer types, including glioblastoma, melanoma, hepatocellular carcinoma, and bladder cancer. They are most abundant at C146T and C124T and rarer at A57C, with the latter originally described as a familial case but subsequently shown also to occur somatically. All three mutations createde novoETS (E-twenty-six specific) binding sites and result in the reactivation of theTERTgene, allowing cancer cells to achieve replicative immortality. Here, we employed a systematic proteomics screen to identify transcription factors preferentially binding to the C146T, C124T and A57C mutations. While we confirmed binding of multiple ETS factors to the mutant C146T and C124T sequences, we identified E4F1 as an A57C-specific binder and ZNF148 as aTERTWT binder that is excluded from theTERTpromoter by the C124T allele. Both proteins are activating transcription factors that bind specifically to the A57C and wildtype (at position 124)TERTpromoter sequence in corresponding cell lines and upregulateTERTtranscription and telomerase activity. Our work describes new regulators of TERT gene expression with possible roles in cancer.
Genome-wide association studies reveal novel loci associated with pyrethroid and organophosphate resistance inAnopheles gambiaes.l.
Authors: Lucas Eric R. , Nagi Sanjay C. , Egyir-Yawson Alexander , Essandoh John , Dadzie Sam , Chabi Joseph , Djogbénou Luc S. , Medjigbodo Adandé A. , Edi Constant V. , Ketoh Guillaume K. , Koudou Benjamin G. , Van’t Hof Arjen E. , Rippon Emily J. , Pipini Dimitra , Harding Nicholas J. , Dyer Naomi A. , Cerdeira Louise T. , Clarkson Chris S. , Kwiatkowski Dominic P. , Miles Alistair , Donnelly Martin J. , Weetman David
Published: January 2023

Resistance to insecticides inAnophelesmosquitoes threatens the effectiveness of the most widespread tools currently used to control malaria. The genetic underpinnings of resistance are still only partially understood, with much of the variance in resistance phenotype left unexplained. We performed a multi-country large scale genome-wide association study of resistance to two insecticides widely used in malaria control: deltamethrin and pirimiphos-methyl. Using a bioassay methodology designed to maximise the phenotypic difference between resistant and susceptible samples, we sequenced 969 phenotyped femaleAn. gambiaeandAn. coluzziifrom ten locations across four countries in West Africa (Benin, Côte d’Ivoire, Ghana and Togo), identifying single nucleotide polymorphisms (SNPs) and copy number variants (CNVs) segregating in the populations. The patterns of resistance association were highly multiallelic and variable between populations, with different genomic regions contributing to resistance, as well as different mutations within a given region. While the strongest and most consistent association with deltamethrin resistance came from the region aroundCyp6aa1, this resistance was based on a combination of several independent CNVs inAn. coluzzii, and on a non-CNV bearing haplotype inAn. gambiae. Further signals involved a range of cytochrome P450, mitochondrial, and immunity genes. Similarly, for pirimiphos-methyl, while the strongest signal came from the region ofAce1, more widespread signals included cytochrome P450s, glutathione S-transferases, and a subunit of thenAChRtarget site of neonicotinoid insecticides. The regions aroundCyp9k1and theTepfamily of immune genes were associated with resistance to both insecticide classes, suggesting possible cross-resistance mechanisms. These locally-varying, multigenic and multiallelic patterns highlight the challenges involved in genomic monitoring and surveillance of resistance, and form the basis for improvement of methods used to detect and predict resistance. Based on simulations of resistance variants, we recommend that yet larger scale studies, exceeding 500 phenotyped samples per population, are required to better identify associated genomic regions.
Exploring the Interdependence of TKS4 and CD2AP: Implications for EMT Process and Early Detection in Colon Cancer
Authors: Kurilla Anita , László Loretta , Takács Tamás , Tilajka Álmos , Lukács Laura , Novák Julianna , Pancsa Rita , Buday László , Vas Virag
Published: January 2023

Background

Colon cancer is a leading cause of death worldwide. Although several biomarkers have been developed, more sensitive and specific methods are needed for its early detection. TKS4 and CD2AP scaffold proteins have been already linked to dynamic actin assembly-related processes, such as colon cancer cell migration, although their co-instructive role during tumour-formation remains unknown. Therefore, this study was designed to validate TKS4-CD2AP interaction and study the interdependent effect of TKS4/CD2AP on oncogenic events.

Methods

CD2AP was identified as a novel TKS4 interacting partner in six cell lines via co-immunoprecipitation-mass spectrometry. The interaction was validated via western blot, immunocytochemistry, DuoLink assay and peptide microarray. Gene silencing and overexpressing experiments were performed to uncover the cooperative effects of TKS4 and CD2AP in cell movement and in epithelial-mesenchymal transition (EMT). The expression levels of TKS4 and CD2AP mRNAs were quantified using a human colon cancer array, and the results were subjected to bioinformatic database analysis.

Results

The molecular biology analysis revealed that one of the SH3-domain of CD2AP interacts with a proline-rich short linear motif in TKS4. Functional studies showed that TKS4 and CD2AP form a scaffold complex that regulates migration- and EMT-related pathways of colon cancer cells. The relative TKS4 and CD2AP expression level measurements pointed out that CD2AP/TKS4 ratio is a sensitive biomarker for the identification of tumorous colon tissue.

Conclusions

This is the first study demonstrating the TKS4-CD2AP protein-protein interaction in vitro and in vivo and their interdependent regulative effect on mesenchymal transition-like process in colon cancer. Furthermore, the results highlight that the relative expression levels of CD2AP and TKS4 might serve as a biomarker for the diagnosis of early-stage colon cancer.
In vitroandin vivoevidences propound therapeutic potential of Lipocalin 2 in cervical carcinoma
Authors: Dwivedi Nehanjali , Mazumder Tahmina , Veeraraghavan Gayathri , Siva Ramanujam , Smitha P K , Ranade Rohit , Das Manjula , Dhar Sujan K
Published: January 2023

Cervical cancer (CC), the second most common in developing countries and the third most common in developed nations, is the fourth most common type of cancer in women overall. The HPV16 high-risk genotype of the virus, which is responsible for about 61% of cervical cancer incidences, was found to have higher LCN2 levels in advanced clinical CC stages. In this study, we assessed the impact of suppressing LCN2 activity after treatment with an anti-LCN2 monoclonal antibody (MAb) in bothin vitroandin vivosettings. Anti-LCN2 antibody was found to reduce proliferation and invasion of HeLa cells, the first immortal cells from a HPV positive aggressive adenocarcinoma of the cervix. LCN2 and its ligand MMP9 was found to be highly expressed in the cells and abrogated on treatment with anti-LCN2. The five receptors of LCN2 - SLC22A17, MC1R, MC2R, MC4R and LRP2 were barely detected with or without treatment. Anti-LCN2 Mab caused tumors to regress and softenin vivo, in a xenograft mouse model. Analysis of histology images of the treated and untreated tumor established the necrotic capability of the therapeutic molecule explaining the regression and softening of the tumor. Differential gene expression analysis between untreated and treated tumor proved that LCN2 inhibition abolished the migratory, invasive, and hypoxic pathways while significantly increasing the necrosis and cell death pathways in tumor after treatment with the monoclonal antibody. LCN2 inhibition was shown molecularly to lead to tumor regression via a negative feedback loop of LCN2 through the TNFα-IL17 axis exponentially increasing the effect of the anti-LCN2 monoclonal antibody. In conclusion, LCN2 appears to be a viable therapeutic target, and the monoclonal antibody used in this study can be further developed for clinical usage in cervical cancer.
scDual-Seq ofToxoplasma gondii-infected mouse bone marrow-derived dendritic cells reveals host cell heterogeneity and differential infection dynamics
Authors: Hildebrandt Franziska , Mohammed Mubasher , Dziedziech Alexis , Bhandage Amol , Divne Anna-Maria , Barrenäs Fredrik , Barragan Antonio , Henriksson Johan , Ankarklev Johan
Published: January 2023

Host-parasite interactions include complex interplays between an invading and a defending organism, each continuously adapting to gain the upper hand. The protozoan parasite,Toxoplasma gondii, can invade every nucleated cell type in a vertebrate host, including immune cells while the host mounts a protective response.

Here, we utilize Dual-scSeq to parse out heterogeneous transcription of bone marrow-derived dendritic cells (BMDCs) infected withT. gondiitype I, RH (LDM) or type II, ME49 (PTG) parasites, over multiple time points post infection.

We find that the two parasite lineages distinctly manipulate two subpopulations of infected BMDCs. Co-expression networks establish host and parasite genes, with implications for modulation of host immunity and host-pathogen interactions. Integration of published data validates immune pathways and suggests novel candidate genes involved in host-pathogen interactions. This study aims to provide a comprehensive resource for future characterization of host-pathogen interplay among other protozoan parasites within their host niches, as well as that of bacterial and viral pathogens.
An intravenous DNA-binding priming agent protects cell-free DNA and improves the sensitivity of liquid biopsies
Authors: Tabrizi Shervin , Martin-Alonso Carmen , Xiong Kan , Blewett Timothy , Sridhar Sainetra , An Zhenyi , Patel Sahil , Rodriguez-Aponte Sergio , Naranjo Christopher A. , Wang Shih-Ting , Shea Douglas , Golub Todd R. , Bhatia Sangeeta N. , Adalsteinsson Viktor , Love J. Christopher
Published: January 2023

Blood-based, or “liquid,” biopsies enable minimally invasive diagnostics but have limits on sensitivity due to scarce cell-free DNA (cfDNA). Improvements to sensitivity have primarily relied on enhancing sequencing technologyex vivo. Here, we sought to augment the level of circulating tumor DNA (ctDNA) detected in a blood draw by attenuating the clearance of cfDNAin vivo. We report a first-in-class intravenous DNA-binding priming agent given 2 hours prior to a blood draw to recover more cfDNA. The DNA-binding antibody minimizes nuclease digestion and organ uptake of cfDNA, decreasing its clearance at 1 hour by over 150-fold. To improve plasma persistence and limit potential immune interactions, we abrogated its Fc-effector function. We found that it protects GC-rich sequences and DNase-hypersensitive sites, which are ordinarily underrepresented in cfDNA. In tumor-bearing mice, priming improved tumor DNA recovery by 19-fold and sensitivity for detecting cancer from 6% to 84%. These results suggest a novel method to enhance the sensitivity of existing DNA-based cancer testing using blood biopsies.
Screening megasynthetase mutants at high throughput using droplet microfluidics
Authors: Pourmasoumi Farzaneh , Hengoju Sundar , Beck Katharina , Stephan Philipp , Klopfleisch Lukas , Hoernke Maria , Rosenbaum Miriam A. , Kries Hajo
Published: January 2023

Nonribosomal peptide synthetases (NRPSs) are giant enzymatic assembly lines that deliver many pharmaceutically valuable natural products, including antibiotics. As the search for new antibiotics motivates attempts to redesign nonribosomal metabolic pathways, more robust and rapid sorting and screening platforms are needed. Here, we establish a microfluidic platform that reliably detects production of the model nonribosomal peptide gramicidin S. The detection is based on calcein-filled sensor liposomes yielding increased fluorescence upon permeabilization. From a library of NRPS mutants, the sorting platform enriches the gramicidin S producer 14.5-fold, decreases the number of stop codons 250-fold, and generates enrichment factors correlating with enzyme activity. Screening for NRPS activity with a reliable non-binary sensor will enable more sophisticated structure-activity studies and new engineering applications in the future.
Comprehensive detection of structural variation and transposable element differences between wild type laboratory lineages ofC. elegans
Authors: Bush Zachary D. , Naftaly Alice F. S. , Dinwiddie Devin , Albers Cora , Hillers Kenneth J. , Libuda Diana E.
Published: January 2023

Genomic structural variations (SVs) and transposable elements (TEs) can be significant contributors to genome evolution, altered gene expression, and risk of genetic diseases. Recent advancements in long-read sequencing have greatly improved the quality ofde novogenome assemblies and enhanced the detection of sequence variants at the scale of hundreds or thousands of bases. Comparisons between two diverged wild isolates ofCaenorhabditis elegans, the Bristol and Hawaiian strains, have been widely utilized in the analysis of small genetic variations. Genetic drift, including SVs and rearrangements of repeated sequences such as TEs, can occur over time from long-term maintenance of wild type isolates within the laboratory. To comprehensively detect both large and small structural variations as well as TEs due to genetic drift, we generatedde novogenome assemblies and annotations for each strain from our lab collection using both long- and short-read sequencing and compared our assemblies and annotations with that of other lab wild type strains. Within our lab assemblies, we annotate over 3.1Mb of sequence divergence between the Bristol and Hawaiian isolates: 337,584 SNPs, 94,503 small insertion-deletions (<50bp), and 4,334 structural variations (>50bp). Further, we define the location and movement of specific DNA TEs between N2 Bristol and CB4856 Hawaiian wild type isolates. Specifically, we find the N2 Bristol genome has 20.6% more TEs from theTc1/marinerfamily than the CB4856 Hawaiian genome. Moreover, we identified Zator elements as the most abundant and mobile TE family in the genome. Using specific TE sequences with unique SNPs, we also identify 38 TEs that moved intrachromosomally and 9 TEs that moved interchromosomally between the N2 Bristol and CB4856 Hawaiian genomes. By comparing thede novogenome assembly of our lab collection Bristol isolate to the VC2010 Bristol assembly, we also reveal that lab lineages display over 2 Mb of total variation: 1,162 SNPs, 1,528 indels, and 897 SVs with 95% of the variation due to SVs. Overall, our work demonstrates the unique contribution of SVs and TEs to variation and genetic drift between wild type laboratory strains assumed to be isogenic despite growing evidence of genetic drift and phenotypic variation.

Author Summary

For multiple model organisms, propagation of wild type strains in independent labs can lead to multiple phenotypic differences over time. To assess recombination, map mutations, and understand genomic changes during speciation,Caenorhabditis elegansresearchers primarily use the wild type isolates Bristol and Hawaiian. Here, we map structural variations, transposable elements, and sequence divergence between the Bristol and Hawaiian natural isolates and between genomes of different lab lineages of these same strains.
Recovery of the gut microbiome following enteric infection and persistence of antimicrobial resistance genes in specific microbial hosts
Authors: Hansen Zoe A. , Vasco Karla A. , Rudrik James T. , Scribner Kim T. , Zhang Lixin , Manning Shannon D.
Published: January 2023

Enteric pathogens cause widespread foodborne illness and are increasingly found to harbor antimicrobial resistance. The ecological impact of these pathogens on the human gut microbiome and resistome, however, has yet to be fully elucidated. This study applied shotgun metagenome sequencing to stools from 60 patients (cases) with enteric bacterial infections for comparison to stools collected from the same patients’ post-recovery (follow-ups). Overall, the case samples harbored more antimicrobial resistance genes (ARGs) and had greater resistome diversity than the follow-up samples (p<0.001), while follow-ups had much more diverse microbiomes (p<0.001). Although cases were primarily defined by generaEscherichia, Salmonella, andShigellaalong with ARGs for multi-compound and multidrug resistance, follow-ups had a greater abundance of Bacteroidetes and Firmicutes phyla and genes for tetracycline, macrolides, lincosamides, and streptogramins (MLS), and aminoglycoside resistance. A host-tracking analysis revealed thatEscherichiawas the primary carrier of ARGs in both cases and follow-ups, with a greater abundance occurring during infection. Eleven distinct extended spectrum beta-lactamases (ESBLs) were identified during infection, some of which appear to be lost or transferred to different microbial hosts upon recovery. The increasing incidence of disease caused by foodborne pathogens, coupled with their evolving role in harboring and transferring antimicrobial resistance determinants within communities, justifies further examination of the repercussions of enteric infection on human gut ecology.
Assembly of gut-derived bacterial communities follows “early-bird” resource utilization dynamics
Authors: Aranda-Díaz Andrés , Willis Lisa , Nguyen Taylor H. , Ho Po-Yi , Vila Jean , Thomsen Tani , Chavez Taylor , Yan Rose , Yu Feiqiao Brian , Neff Norma , Sanchez Alvaro , Estrela Sylvie , Huang Kerwyn Casey
Published: January 2023

Diet can impact host health through changes to the gut microbiota, yet we lack mechanistic understanding linking nutrient availability and microbiota composition. Here, we use thousands of microbial communities culturedin vitrofrom human feces to uncover simple assembly rules and develop a predictive model of community composition upon addition of single nutrients from central carbon metabolism to a complex medium. Community membership was largely determined by the donor feces, whereas relative abundances were determined by the supplemental carbon source. The absolute abundance of most taxa was independent of the supplementing nutrient, due to the ability of fast-growing organisms to quickly exhaust their niche in the complex medium and then exploit and monopolize the supplemental carbon source. Relative abundances of dominant taxa could be predicted from the nutritional preferences and growth dynamics of species in isolation, and exceptions were consistent with strain-level variation in growth capabilities. Our study reveals that community assembly follows simple rules of nutrient utilization dynamics and provides a predictive framework for manipulating gut commensal communities through nutritional perturbations.
Activation of thePlasmodiumegress effector subtilisin-like protease 1 is achieved by plasmepsin X destruction of the propiece
Authors: Mukherjee Sumit , Nasamu Armiyaw S. , Rubiano Kelly , Goldberg Daniel E.
Published: January 2023

Following each round of replication, daughter merozoites of the malaria parasitePlasmodium falciparumescape (egress) from the infected host red blood cell (RBC) by rupturing the parasitophorous vacuole membrane (PVM) and the RBC membrane (RBCM). A proteolytic cascade orchestrated by the parasite’s serine protease, subtilisin-like protease 1 (SUB1) regulates the membrane breakdown. SUB1 activation involves primary auto-processing of the 82 kDa zymogen to a 54 kDa (p54) intermediate that remains bound to its inhibitory propiece (p31) post cleavage. A second processing step converts p54 to the terminal 47 kDa (p47) form of SUB1. Although the aspartic protease plasmepsin X (PM X) has been implicated in the activation of SUB1, the mechanism remains unknown. Here, we show that upon knockdown of PM X the inhibitory p31/p54 complex of SUB1 accumulates in the parasites. Using recombinant PM X and SUB1, we show that PM X can directly cleave both p31 and p54. We have mapped the cleavage sites on recombinant p31. Furthermore, we demonstrate that the conversion of p54 to p47 can be effected by cleavage at either a SUB1 or PM X cleavage site that are adjacent to one another. Importantly once the p31 is removed, p54 is fully functional inside the parasites suggesting that the conversion to p47 is dispensable for SUB1 activity. Relief of propiece inhibition via a heterologous protease is a novel mechanism for subtilisin activation.

Significance Statement

Malaria parasites replicate inside a parasitophorous vacuole within the host red blood cells. Exit of mature progeny from the infected host cells is essential for further dissemination. Parasite exit is a highly regulated, explosive process that involves membrane breakdown. To do this, the parasite utilizes a serine protease, called the subtilisin-like protease 1 or SUB1 that proteolytically activates various effector proteins. SUB1 activity is dependent on an upstream protease, called plasmepsin X (PM X), although the mechanism was unknown. Here we describe the molecular basis for PM X mediated SUB1 activation. PM X proteolytically degrades the inhibitory segment of SUB1, thereby activating it. Involvement of a heterologous protease is a novel mechanism for subtilisin activation.
A nanoparticle priming agent reduces cellular uptake of cell-free DNA and enhances the sensitivity of liquid biopsies
Authors: Martin-Alonso Carmen , Tabrizi Shervin , Xiong Kan , Blewett Timothy , Patel Sahil , An Zhenyi , Sridhar Sainetra , Bekdemir Ahmet , Shea Douglas , Amini Ava P. , Wang Shih-Ting , Kirkpatrick Jesse , Rhoades Justin , Golub Todd R. , Love J. Christopher , Adalsteinsson Viktor A. , Bhatia Sangeeta N.
Published: January 2023

Liquid biopsies are enabling minimally invasive monitoring and molecular profiling of diseases across medicine, but their sensitivity remains limited by the scarcity of cell-free DNA (cfDNA) in blood. Here, we report an intravenous priming agent that is given prior to a blood draw to increase the abundance of cfDNA in circulation. Our priming agent consists of nanoparticles that act on the cells responsible for cfDNA clearance to slow down cfDNA uptake. In tumor-bearing mice, this agent increases the recovery of circulating tumor DNA (ctDNA) by up to 60-fold and improves the sensitivity of a ctDNA diagnostic assay from 0% to 75% at low tumor burden. We envision that this priming approach will significantly improve the performance of liquid biopsies across a wide range of clinical applications in oncology and beyond.