bluuebunny/crossref_metadata_2025 huggingface dataset:
Glioblastoma Mesenchymal Transition and Invasion are Dependent on a NF-κB/BRD2 Chromatin Complex
Authors: Vadla Raghavendra , Miki Shunichiro , Taylor Brett , Kawauchi Daisuke , Jones Brandon M , Nathwani Nidhi , Pham Philip , Tsang Jonathan , Nathanson David A. , Furnari Frank B.
Published: July 2023

Glioblastoma (GBM) represents the most aggressive subtype of glioma, noted for its profound invasiveness and molecular heterogeneity. The mesenchymal (MES) transcriptomic subtype is frequently associated with therapy resistance, rapid recurrence, and increased tumor-associated macrophages. Notably, activation of the NF-κB pathway and alterations in thePTENgene are both associated with this malignant transition. Although PTEN aberrations have been shown to be associated with enhanced NF-κB signaling, the relationships between PTEN, NF-κB and MES transition are poorly understood in GBM. Here, we show that PTEN regulates the chromatin binding of bromodomain and extraterminal (BET) family proteins, BRD2 and BRD4, mediated by p65/RelA localization to the chromatin. By utilizing patient-derived glioblastoma stem cells and CRISPR gene editing of theRELAgene, we demonstrate a crucial role for RelA lysine 310 acetylation in recruiting BET proteins to chromatin for MES gene expression and GBM cell invasion uponPTENloss. Remarkably, we found that BRD2 is dependent on chromatin associated acetylated RelA for its recruitment to MES gene promoters and their expression. Furthermore, loss of BRD2 results in the loss of MES signature, accompanied by an enrichment of proneural signature and enhanced therapy responsiveness. Finally, we demonstrate that disrupting the NF-κB/BRD2 interaction with a brain penetrant BET-BD2 inhibitor reduces mesenchymal gene expression, GBM invasion, and therapy resistance in GBM models. This study uncovers the role of hitherto unexplored PTEN-NF-κB-BRD2 pathway in promoting MES transition and suggests inhibiting this complex with BET-BD2 specific inhibitors as a therapeutic approach to target the MES phenotype in GBM.
Longitudinal hair cortisol in bipolar disorder and a mechanism based on HPA dynamics
Authors: Milo Tomer , Maimon Lior , Cohen Ben , Haran Dafna , Segman Dror , Danon Tamar , Bren Anat , Mayo Avi , Cohen Rappaport Gadi , McInnis Melvin , Alon Uri
Published: July 2023

Bipolar disorder (BD) is a dynamic disease in which mania, depression and mixed states vary on a timescale of months to years. BD patients characteristically exhibit elevated levels of the hormone cortisol. Persistently elevated cortisol can also cause mood episodes in a substantial fraction of the general population. Although BD is a dynamic disease that is related to cortisol, longitudinal cortisol dynamics in BD have rarely been studied. Here we use hair to measure past cortisol where each cm of hair corresponds to a month of growth. Cortisol was measured in 12 cm hair samples from people with BD (n=26) and controls (n=59), corresponding to one year of cortisol data. We found that hair cortisol exhibited a frequency spectrum with enhanced year-scale fluctuations whose amplitude was about 4-fold higher on average in BD compared to controls. Cortisol in the proximal 2 cm hair segment correlated with mood scales that report on mood in the past two months. In line with the notion that cortisol correlates with mood, we find that the mean frequency spectrum of depression (n=266) and mania (n=273) scores from a large longitudinal study of BD is similar to the hair cortisol spectrum from the present cohort. Taken together, these results suggest a mechanism for BD as the intersection of two neuropsychological traits: cortisol-induced mood episodes (CIM) and high emotional reactivity (ER). High ER causes fluctuations in which cortisol is elevated for months, as shown by a mathematical model of the hypothalamic-pituitary-adrenal (HPA) axis that regulates cortisol. In individuals with CIM, the magnitude of these persistent cortisol fluctuations can be high enough to trigger mood episodes. Thus, this study combines longitudinal cortisol measurements and mathematical modeling to provide a potential mechanistic link between the timescales of cortisol and moods in BD.
Identification and distribution of novel candidate T6SS effectors encoded inSalmonellaPathogenicity island 6
Authors: Blondel Carlos J. , Amaya Fernando A. , Bustamante Paloma , Santiviago Carlos A. , Pezoa David
Published: July 2023

The type VI secretion system (T6SS) is a contact-dependent contractile multiprotein apparatus widely distributed in Gram-negative bacteria. These systems can deliver different effector proteins into target bacterial and/or eukaryotic cells, contributing to the environmental fitness and virulence of many bacterial pathogens.Salmonellaharbors five different T6SSs encoded in different genomic islands. The T6SS encoded inSalmonellaPathogenicity Island 6 (SPI-6) contributes toSalmonellacompetition with the host microbiota and its interaction with infected host cells. Despite its relevance, information regarding the total number of effector proteins encoded within SPI-6 and its distribution among differentSalmonella entericaserotypes is limited. In this work, we performed bioinformatic and comparative genomics analyses of the SPI-6 T6SS gene cluster to expand our knowledge regarding the T6SS effector repertoire and the global distribution of these effectors inSalmonella. The analysis of a curated dataset of 60Salmonella entericagenomes from the Secret6 database revealed the presence of 23 novel putative T6SS effector/immunity protein (E/I) modules. These effectors were concentrated in the variable regions 1 to 3 (VR1-3) of the SPI-6 T6SS gene cluster. VR1-2 were enriched in candidate effectors with predicted peptidoglycan hydrolase activity, while VR3 was enriched in candidate effectors of the Rhs family with C-terminal extensions with predicted DNase, RNase, deaminase, or ADP-ribosyltransferase activity. A global analysis of known and candidate effector proteins inSalmonella entericagenomes from the NCBI database revealed that T6SS effector proteins are differentially distributed amongSalmonellaserotypes. While some effectors are present in over 200 serotypes, others are found in less than a dozen. A hierarchical clustering analysis identifiedSalmonellaserotypes with distinct profiles of T6SS effectors and candidate effectors, highlighting the diversity of T6SS effector repertoires inSalmonella enterica. The existence of different repertoires of effector proteins suggests that different effector protein combinations may have a differential impact on the environmental fitness and pathogenic potential of these strains.
Emergence of putative energy parasites withinClostridiarevealed by genome analysis of a novel endosymbiotic clade
Authors: Takahashi Kazuki , Kuwahara Hirokazu , Horikawa Yutaro , Izawa Kazuki , Kato Daiki , Inagaki Tatsuya , Yuki Masahiro , Ohkuma Moriya , Hongoh Yuichi
Published: July 2023

TheClostridiais a dominant bacterial class in the guts of various animals and are considered to nutritionally contribute to the animal host. Here, we discovered clostridial endosymbionts of cellulolytic protists in termite guts, which have never been reported with evidence. We obtained (near-)complete genome sequences of three endosymbioticClostridia, each associated with a different parabasalid protist species with various infection rates:Trichonympha agilis,Pseudotrichonympha grassii, andDevescovinasp. All these protists are previously known to harbor permanently-associated, mutualisticEndomicrobiaorBacteroidalesthat supplement nitrogenous compounds. The genomes of the endosymbioticClostridiawere small in size (1.0–1.3 Mbp) and exhibited signatures of an obligately-intracellular parasite, such as an extremely limited capability to synthesize amino acids, cofactors, and nucleotides and a disrupted glycolytic pathway with no known net ATP-generating system. Instead, the genomes encoded ATP/ADP translocase and, interestingly, regulatory proteins that are unique to eukaryotes in general and are possibly used to interfere with host cellular processes. These three genomes formed a clade with metagenome-assembled genomes (MAGs) derived from the guts of other animals, including human and ruminants, and the MAGs shared the characteristics of parasites. Gene flux analysis suggested that the acquisition of the ATP/ADP translocase gene in a common ancestor was probably key to the emergence of this parasitic clade. Taken together, we provide novel insights into the multilayered symbiotic system in the termite gut by adding the presence of parasitism and present an example of the emergence of putative energy parasites from a dominant gut bacterial clade.
Robust Feature Selection strategy detects a panel of microRNAs as putative diagnostic biomarkers in Breast Cancer
Authors: Costa Maria Claudia , Noviello Teresa Maria Rosaria , Ceccarelli Michele , Cerulo Luigi
Published: July 2023

MicroRNAs represent a comprehensive class of short, single-stranded, non-coding RNA transcripts able to interfere with the translation of their targets or degrade them. Since these molecules are dysregulated in several malignancies, they represent reliable biomarkers in various contexts. In this study, the application of several Feature Selection methods uncovers a panel of 20 microRNAs, of whichhsa-mir-337, hsa-mir-378c, and hsa-mir-483are still poorly investigated in the context of Breast Cancer. This signature is capable of discriminating between healthy and tumoral samples, showing better classification performance when compared with differentially expressed microRNAs. Furthermore, a network-based centrality analysis on the gene targets of these transcripts highlightedCDC25, TPX2, KIF18B, CDCA3, TGFBR2, CAV1, TNS1, andLHFPL6as key dysregulated genes. This study providesin silicohypotheses as new insights for futurein vivoorin vitrostudies uncovering the role of these putative diagnostic biomarkers in Breast Cancer.
Diseased human pancreas and liver microphysiological system for preclinical diabetes research
Authors: Rigal Sophie , Casas Belén , Kanebratt Kajsa P. , Huldt Charlotte Wennberg , Magnusson Lisa U. , Müllers Erik , Karlsson Fredrik , Clausen Maryam , Hansson Sara F. , Löfmark Rasmus Jansson , Ämmälä Carina , Marx Uwe , Gennemark Peter , Cedersund Gunnar , Andersson Tommy B. , Vilén Liisa K.
Published: July 2023

Current research on metabolic disorders such as type 2 diabetes relies on animal models because multi-organ diseases cannot be well studied with the standardin vitroassays. Here, we connect models of key metabolism organs, pancreas and liver, on a microfluidic chip to enable diabetes research in a human-based preclinical system. Aided by mechanistic mathematical modelling, we developed a two-organ microphysiological system (MPS) that replicates clinically-relevant phenotypes of diabetic dysregulation both in the liver and pancreas compartments. Exposure to hyperglycemia and high cortisone created a diseased pancreas-liver MPS which displayed beta-cell dysfunction, steatosis, elevated ketone-body secretion, increased glycogen storage, and upregulated gluconeogenic machinery. In turn, normoglycemia and physiological cortisone concentration maintained glucose tolerance and stable liver and beta-cell functions. This method was evaluated for repeatability in two laboratories and was effective in multiple pancreatic islet donors. The model also provides a platform to identify new therapeutic targets as demonstrated with a liver-secreted IL-1R2 protein that induced islet proliferation.
Intratumoral immune triads are required for adoptive T cell therapy-mediated elimination of solid tumors
Authors: Espinosa-Carrasco Gabriel , Scrivo Aurora , Zumbo Paul , Dave Asim , Betel Doron , Hellmann Matthew , Burt Bryan M , Lee Hyun-Sung , Schietinger Andrea
Published: July 2023

Tumor-reactive CD8 T cells found in cancer patients are frequently dysfunctional, unable to halt tumor growth. Adoptive T cell transfer (ACT), the administration of large numbers ofin vitro-generated cytolytic tumor-reactive CD8 T cells, is an important cancer immune therapy being pursued. However, a limitation of ACT is that transferred CD8 T cells often rapidly lose effector function, and despite exciting results in certain malignancies, few ACT clinical trials have shown responses in solid tumors. Here, we developed preclinical cancer mouse models to investigate if and how tumor-specific CD4 T cells can be enlisted to overcome CD8 T cell dysfunction in the setting of ACT.In situconfocal microscopy of color-coded cancer cells, tumor-specific CD8 and CD4 T cells, and antigen presenting cells (APC), combined with functional studies, revealed that the spatial positioning and interactions of CD8 and CD4 T cells, but not their numbers, dictates ACT efficacy and anti-tumor responses. We uncover a new role of antigen-specific CD4 T cells in addition to the known requirement for CD4 T cells during priming/activation of naïve CD8 T cells. CD4 T cells must co-engage with CD8 T cells and APC cross-presenting CD8-and CD4-tumor antigens during the effector phase, forming a three-cell-cluster (triad), to license CD8 T cell cytotoxicity and mediate cancer cell elimination. Triad formation transcriptionally and epigenetically reprogram CD8 T cells, prevent T cell dysfunction/exhaustion, and ultimately lead to the elimination of large established tumors and confer long-term protection from recurrence. When intratumoral triad formation was disrupted, adoptively transferred CD8 T cells could not be reprogrammed, and tumors progressed despite equal numbers of tumor-infiltrating CD8 and CD4 T cells. Strikingly, the formation of CD4 T cell::CD8 T cell::APC triads in tumors of patients with lung cancers treated with immune checkpoint blockade was associated with clinical responses, but not CD4::APC dyads or overall numbers of CD8 or CD4 T cells, demonstrating the importance of triads in non-ACT settings in humans. Our work uncovers intratumoral triads as a key requirement for anti-tumor immunity and a new role for CD4 T cells in CD8 T cell cytotoxicity and cancer cell eradication.
Investigation of swine caecal microbiomes in the northern region of Thailand
Authors: Eiamsam-ang Thanaporn , Tadee Pakpoom , Buddhasiri Songphon , Chuammitri Phongsakorn , Kittiwan Nattinee , Pascoe Ben , Patchanee Prapas
Published: July 2023

Introduction

The northern region of Thailand serves as a crucial area for swine production, contributing to the global food supply. Previous studies have highlighted the presence of foodborne pathogens originating from swine farms in this region, posing a threat to both human and animal health.

Gap statement

Multiple swine pathogens have been studied at a species level, but the distribution and co-occurrence of pathogens in agricultural swine has not been well established.

Aim

Our study employed the intestinal scraping technique to directly examine the microorganisms interacting with the swine host.

Methodology

We used shotgun metagenomic sequencing to analyse the caecal microbiomes of swine from five commercial farms in northern Thailand.

Results

Swine caecal microbiomes contained commensal bacteria such asBifidobacterium,Lactobacillus, andFaecalibacterium, which are associated with healthy physiology and feed utilisation. We also identified multiple pathogenic and opportunistic bacteria present in all samples, includingEscherichia coli,Clostridium botulinum,Staphylococcus aureus, and theCorynebacteriumgenus. From a One Health perspective, these species are important foodborne and opportunistic pathogens in both humans and agricultural animals. Antimicrobial resistance genes were also detected in all samples, specifically conferring resistance to tetracycline and aminoglycosides which have historically been used extensively in swine farming.

Conclusion

The findings further support the need for improved sanitation standards in swine farms, and additional monitoring of agricultural animals and farm workers to reduce contamination and improved produce safety for human consumption.
Structural characterization of the Sel1-like repeat protein LceB fromLegionella pneumophila
Authors: Penner Tiffany V. , Lorente Cobo Neil , Patel Deepak T. , Patel Dhruvin H. , Savchenko Alexei , Brassinga Ann Karen C. , Prehna Gerd
Published: July 2023

Legionellaare freshwater Gram-negative bacteria that in their normal environment infect protozoa. However, this adaptation also allowsLegionellato infect human alveolar macrophages and cause pneumonia. Central toLegionellapathogenesis are more than 330 secreted effectors, of which there are 9 core effectors that are conserved in all pathogenic species. Despite their importance, the biochemical function of several core effectors remains unclear. To address this, we have taken a structural approach to characterize the core effector of unknown function LceB, or Lpg1356, fromLegionella pneumophila.Here we solve an X-ray crystal structure of LceB using an AlphaFold model for molecular replacement. The experimental structure shows that LceB adopts a Sel1-like repeat fold as predicted. However, the crystal structure captured multiple conformations of LceB all of which differed from the AlphaFold model. Comparison of the predicted model and the experimental models suggests that LceB is highly flexible in solution. Additionally, molecular analysis of LceB using its close structural homologues reveals sequence and structural motifs of known biochemical function. Specifically, LceB harbors a repeated KAAEQG motif that both stabilizes the Sel1-like repeat fold and is known to participate in protein-protein interactions with eukaryotic host proteins. We also observe that LceB forms several higher-order oligomers in solution. Overall, our results have revealed that LceB has conformational flexibility, self-associates, and contains a molecular surface for binding a target host-cell protein. Additionally, our data provides structural insights into the Sel1-like repeat family of proteins that remain poorly studied.
Large-scale deorphanization ofNematostella vectensisneuropeptide GPCRs supports the independent expansion of bilaterian and cnidarian peptidergic systems
Authors: Thiel Daniel , Yañez-Guerra Luis Alfonso , Kieswetter Amanda , Cole Alison G. , Temmerman Liesbet , Technau Ulrich , Jékely Gáspár
Published: July 2023

Neuropeptides are ancient signaling molecules in animals but only few peptide receptors are known outside bilaterians. Cnidarians possess a large number of G protein-coupled receptors (GPCRs) – the most common receptors of bilaterian neuropeptides – but most of these remain orphan with no known ligands. We searched for neuropeptides in the sea anemoneNematostella vectensisand created a library of 64 peptides derived from 33 precursors. In a large-scale pharmacological screen with these peptides and 161N. vectensisGPCRs, we identified 31 receptors specifically activated by one of 14 peptides. Mapping GPCR and neuropeptide expression to single-cell sequencing data revealed how cnidarian tissues are extensively wired by multilayer peptidergic networks. Phylogenetic analysis identified no direct orthology to bilaterian peptidergic systems and supports the independent expansion of neuropeptide signaling in cnidarians from a few ancestral peptide-receptor pairs.
SH2 domain protein E (SHE) and ABL signaling regulate blood vessel size
Authors: Schumacher Jennifer A. , Wright Zoë A. , Florat Diandra Rufin , Anand Surendra K. , Dasyani Manish , Klimkaite Laurita , Bredemeier Nina O. , Gurung Suman , Koller Gretchen M. , Aguera Kalia N. , Chadwick Griffin P. , Johnson Riley D. , Davis George E. , Sumanas Saulius
Published: July 2023

Blood vessels in different vascular beds vary in lumen diameter, which is essential for their function and fluid flow along the vascular network. Molecular mechanisms involved in the formation of a vascular lumen of appropriate size, or tubulogenesis, are still only partially understood.Src homology 2 domain containing E (She)protein was previously identified in a screen for proteins that interact with Abelson (Abl)-kinase. However, its biological role has remained unknown. Here we demonstrate that She and Abl signaling regulate vascular lumen size in zebrafish embryos and human endothelial cell culture. Zebrafishshemutants displayed increased endothelial cell number and enlarged lumen size of the dorsal aorta (DA) and defects in blood flow. Vascular endothelial specific overexpression ofsheresulted in a reduced diameter of the DA lumen, which correlated with the reduced arterial cell number and lower endothelial cell proliferation. Chemical inhibition of Abl signaling in zebrafish embryos caused a similar reduction in the DA diameter and alleviated theshemutant phenotype, suggesting that She acts as a negative regulator of Abl signaling. Enlargement of the DA lumen inshemutants correlated with an increased endothelial expression ofclaudin 5aand5b (cldn5a / cldn5b), which encode proteins enriched in tight junctions. Inhibition ofcldn5aexpression partially rescued the enlarged DA inshemutants, suggesting that She regulates DA lumen size, in part, by promotingcldn5aexpression. SHE knockdown in human endothelial umbilical vein cells resulted in a similar increase in the diameter of vascular tubes, and also increased phosphorylation of a known ABL downstream effector CRKL. These results argue that SHE functions as an evolutionarily conserved inhibitor of ABL signaling and regulates lumen size during vascular tubulogenesis.
Ym1 protein crystals promote type 2 immunity
Authors: Heyndrickx Ines , Deswarte Kim , Verstraete Kenneth , Verschueren Koen H. G. , Smole Ursula , Aegerter Helena , Dansercoer Ann , Hammad Hamida , Savvides Savvas N. , Lambrecht Bart N.
Published: July 2023

Spontaneous protein crystallization is a rare event, yet protein crystals are frequently found in eosinophil-rich inflammation. In humans, Charcot-Leyden crystals (CLCs) are made from galectin-10 (Gal10) protein, an abundant protein in eosinophils. Whereas mice do not encode Gal10 in their genome, they do form pseudo-CLCs, made from the chitinase-like proteins Ym1 and/or Ym2, encoded byChil3andChil4and made by myeloid and epithelial cells respectively. Here, we investigated the biological effects of pseudo-CLCs since their function is currently unknown. We produced recombinant Ym1 crystals which were shown to have identical crystal packing and structure by X-ray crystallography asin vivonative crystals derived from murine lung. When administered to the airways of mice, crystalline but not soluble Ym1 stimulated innate and adaptive immunity and acted as a type 2 immune adjuvant for eosinophilic inflammation via triggering of dendritic cells (DCs). Murine Ym1 protein crystals found at sites of eosinophilic inflammation reinforce type 2 immunity and could serve as a surrogate model for studying the biology of human CLCs.
Interrogation of RNA-protein interaction dynamics in bacterial growth
Authors: Monti Mie , Herman Reyme , Davey Karen , Dawson Charlotte S. , Ule Jernej , Thomas Gavin H. , Willis Anne E. , Lilley Kathryn S. , Villanueva Eneko
Published: July 2023

Characterising RNA-protein interaction dynamics is fundamental to understand how bacteria respond to their environment. In this study, we have analysed the dynamics of 91% of theEscherichia coliexpressed proteome and the RNA-interaction properties of 271 RNA-binding proteins (RBPs) at different growth phases. We find that 68% of RBPs differentially bind RNA across growth phases and characterise 17 previously unannotated proteins as bacterial RBPs including YfiF, a ncRNA-binding protein. While these new RBPs are mostly present in Proteobacteria, two of them have human orthologs in the form of mitochondrial proteins associated with rare metabolic disorders. Moreover, we reveal novel RBP functions for proteins such as the chaperone HtpG, a new stationary phase tRNA-binding protein. Altogether, we provide the first dynamic RBPome of a bacterium, showcasing how this approach can reveal the function of uncharacterised proteins, and identify critical RNA-protein interactions for cell growth which could inform new antimicrobial therapies.
Multimodal optical mesoscopy reveals the quantity and spatial distribution of gram-positive biofilms inex vivotonsils
Authors: Clapperton Megan , Kunanandam Tash , Florea Catalina D. , Douglas Catriona M. , McConnell Gail
Published: July 2023

Biofilms are known to be present in tonsils, but little is known about their spatial location and size distribution throughout the tonsil. Studies of the location and distribution of biofilms in tonsil specimens have thus far been limited to either high-magnification methods such as electron microscopy, which enables high resolution imaging but only from a tiny tissue volume, or lower magnification techniques such as light microscopy, which allow imaging of larger specimens but with poor spatial resolution. To overcome these limitations, we report the use of multimodal optical mesoscopy to visualize and quantify the number and spatial distribution of gram-positive biofilms in fresh, excised paediatric tonsils. This methodology supports simultaneous imaging of both the tonsil host and biofilms in whole mounts of tissue up to 5 mm × 5 mm × 3 mm with subcellular resolution throughout. A quantitative assessment of thirty-six tonsil specimens revealed no statistically significant difference between biofilm presence on the tonsil surface and the interior of the tonsil. This new quantitative mesoscale imaging approach may prove useful in understanding the role of biofilms in tonsillar diseases and other infections.
Direct recruitment of Mis18 to interphase spindle poles promotes CENP-A chromatin assembly
Authors: London Nitobe , Medina-Pritchard Bethan , Spanos Christos , Rappsilber Juri , Arulanandam Jeyaprakash , Allshire Robin C.
Published: July 2023

CENP-A chromatin specifies mammalian centromere identity, and its chaperone HJURP replenishes CENP-A when recruited by the Mis18 complex (Mis18C) via M18BP/KNL2 to CENP-C at kinetochores during interphase. However, the Mis18C recruitment mechanism remains unresolved in species lacking M18BP1, such as fission yeast. Fission yeast centromeres cluster at G2 spindle pole bodies (SPBs) when CENP-ACnp1is replenished and where Mis18C also localizes. We show that SPBs play an unexpected role in concentrating Mis18C near centromeres through the recruitment of Mis18 by direct binding to the major SPBLInker ofNucleoskeleton andCytoskeleton (LINC) complex component Sad1. Mis18 recruitment by Sad1 is important for CENP-ACnp1chromatin establishment and acts in parallel with a CENP-C-mediated Mis18C recruitment pathway to maintain centromeric CENP-ACnp1, but is independent of Sad1-mediated centromere clustering. SPBs therefore provide a non-chromosomal scaffold for both Mis18C recruitment and centromere clustering during G2. This centromere-independent Mis18-SPB recruitment provides a mechanism that governsde novoCENP-ACnp1chromatin assembly by the proximity of appropriate sequences to SPBs and highlights how nuclear spatial organization influences centromere identity.
Phylogenomic analysis reveals the evolutionary origins of five independent clades of forage grasses within the African genusUrochloa
Authors: Masters Lizo E. , Tomaszewska Paulina , Schwarzacher Trude , Zuntini Alexandre R. , Heslop-Harrison Pat , Vorontsova Maria S.
Published: July 2023

Background and Aims

The grass genusUrochloa(Brachiaria) includes forage crops that are important for beef and dairy industries in tropical and sub-tropical Africa, South America, and Oceania/Australia. Economically important species includeU. brizantha,U. decumbens,U. humidicola,U. mutica,U. arrecta,U. trichopus,U. mosambicensis, andM. maximus, all native to the African continent. Perennial growth habits, large, fast growing palatable leaves, intra- and interspecific morphological variability, apomictic reproductive systems, and frequent polyploidy are widely shared within the genus. The combination of these traits likely favoured the selection for forage domestication and weediness, but trait emergence acrossUrochloacannot be modelled, as a robust phylogenetic assessment of the genus has not been conducted.

Methods

Using a target enrichment sequencing approach (Angiosperms353 baits), we inferred a species level phylogeny forUrochloa sensu lato, encompassing 57 species (∼50% of the genus) and outgroups. We determined the phylogenetic placement of agriculturally important species and identify their closest wild relatives. Further, we mapped key traits associated with forage crop potential to the species tree, exploring trait distribution across the genus.

Key Results

Agricultural species belong to five independent clades, includingU. brizanthaandU. decumbenslying in a previously defined species complex. Crop wild relatives were identified for these clades supporting previous sub-generic groupings inUrochloabased on morphology. Using ancestral trait estimation models, we find that five morphological traits that correlate with forage potential (perennial growth habits, culm height, leaf size, a winged rachis, and large seeds) independently evolved in forage clades.

Conclusions

Urochloa s.l.is a highly diverse genus that contains numerous species with agricultural potential, including crop wild relatives that are currently underexploited. The African continent is the centre of origin for these clades and their conservation across their native distributions is essential. Genomic and phenotypic diversity in forage clade species and their wild relatives needs to be better assessed to improve sustainability inUrochloacultivar production.
Predicting bacterial interaction outcomes from monoculture growth and supernatant assays
Authors: Schmitz Désirée A. , Wechsler Tobias , Mignot Ingrid , Kümmerli Rolf
Published: July 2023

How to derive principles of community dynamics and stability is a central question in microbial ecology. Bottom-up experiments, in which a small number of bacterial species are mixed, have become popular to address it. However, experimental setups are typically limited because co-culture experiments are labor-intensive and species are difficult to distinguish. Here, we use a 4-species bacterial community to show that information from monoculture growth and inhibitory effects induced by secreted compounds can be combined to predict the competitive rank order in the community. Specifically, integrative monoculture growth parameters allow building a preliminary competitive rank order, which is then adjusted using inhibitory effects from supernatant assays. While our procedure worked for two different media, we observed differences in species rank orders between media. We then parameterized computer simulations with our empirical data to show that higher-order species interactions largely follow the dynamics predicted from pairwise interactions with one important exception. The impact of inhibitory compounds was reduced in higher-order communities because their negative effects were spread across multiple target species. Altogether, we formulated three simple rules of how monoculture growth and supernatant assay data can be combined to establish a competitive species rank order in an experimental 4-species community.
Evolutionary origin, population diversity, and diagnostics for a cryptic hybrid pathogen
Authors: Steenwyk Jacob L. , Knowles Sonja , Bastos Rafael W. , Balamurugan Charu , Rinker David , Mead Matthew E. , Roberts Christopher D. , Raja Huzefa A. , Li Yuanning , Colabardini Ana Cristina , de Castro Patrícia Alves , dos Reis Thaila Fernanda , Canóvas David , Sanchez Rafael Luperini , Lagrou Katrien , Torrado Egídio , Rodrigues Fernando , Oberlies Nicholas H. , Zhou Xiaofan , Goldman Gustavo H. , Rokas Antonis
Published: July 2023

Cryptic fungal pathogens pose significant identification and disease management challenges due to their morphological resemblance to known pathogenic species while harboring genetic and (often) infection-relevant trait differences. The cryptic fungal pathogenAspergillus latus, an allodiploid hybrid originating fromAspergillus spinulosporusand an unknown close relative ofAspergillus quadrilineatuswithin sectionNidulantes, remains poorly understood. The absence of accurate diagnostics forA. latushas led to misidentifications, hindering epidemiological studies and the design of effective treatment plans. We conducted an in-depth investigation of the genomes and phenotypes of 44 globally distributed isolates (41 clinical isolates and three type strains) fromAspergillussectionNidulantes. We found that 21 clinical isolates wereA. latus; notably, standard methods of pathogen identification misidentified allA. latusisolates. The remaining isolates were identified asA. spinulosporus(8),A. quadrilineatus(1), orA. nidulans(11). Phylogenomic analyses shed light on the origin ofA. latus, indicating one or two hybridization events gave rise to the species during the Miocene, approximately 15.4 to 8.8 million years ago. Characterizing theA. latuspangenome uncovered substantial genetic diversity within gene families and biosynthetic gene clusters. Transcriptomic analysis revealed that both parental genomes are actively expressed in nearly equal proportions and respond to environmental stimuli. Further investigation into infection-relevant chemical and physiological traits, including drug resistance profiles, growth under oxidative stress conditions, and secondary metabolite biosynthesis, highlight distinct phenotypic profiles of the hybridA. latuscompared to its parental and closely related species. Leveraging our comprehensive genomic and phenotypic analyses, we propose five genomic and phenotypic markers as diagnostics forA. latusspecies identification. These findings provide valuable insights into the evolutionary origin, genomic outcome, and phenotypic implications of hybridization in a cryptic fungal pathogen, thus enhancing our understanding of the underlying processes contributing to fungal pathogenesis. Furthermore, our study underscores the effectiveness of extensive genomic and phenotypic analyses as a promising approach for developing diagnostics applicable to future investigations of cryptic and emerging pathogens.
Novel phosphatidylinositol flippases contribute to phosphoinositide homeostasis in the plasma membrane
Authors: Muranaka Yumeka , Shigetomi Ryo , Iwasaki Yugo , Hamamoto Asuka , Nakayama Kazuhisa , Takatsu Hiroyuki , Shin Hye-Won
Published: July 2023

Phosphatidylinositol is a precursor of various phosphoinositides, which play crucial roles in intracellular signaling and membrane dynamics and have impact on diverse aspects of cell physiology. Phosphoinositide synthesis and turnover occur in the cytoplasmic leaflet of the organellar and plasma membranes. P4-ATPases (lipid flippases) are responsible for translocating membrane lipids from the exoplasmic (luminal) to the cytoplasmic leaflet, thereby regulating membrane asymmetry. However, the mechanism underlying phosphatidylinositol translocation across cellular membranes remains elusive. Here, we discovered that the phosphatidylcholine flippases ATP8B1, ATP8B2, and ATP10A can also translocate phosphatidylinositol at the plasma membrane. To explore the function of these phosphatidylinositol flippases, we used cells depleted of CDC50A, a protein necessary for P4-ATPase function. Upon activation of the Gq-coupled receptor, depletion of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] was accelerated inCDC50Aknockout cells compared with control cells, suggesting a decrease in PtdIns4,5P2levels within the plasma membrane of the knockout cells. These findings highlight the pivotal role of P4-ATPases in maintaining phosphoinositide homeostasis and suggest a mechanism for enrichment of phosphatidylinositol in the cytoplasmic leaflet of the plasma membrane.
A YTHDF-PABP axis is required for m6A-mediated organogenesis in plants
Authors: Tankmar Mathias Due , Reichel Marlene , Arribas-Hernández Laura , Brodersen Peter
Published: July 2023

N6-methyladenosine (m6A) in mRNA is key to eukaryotic gene regulation. Many m6A functions involve specialized RNA-binding proteins that recognize m6A via a YT521-B Homology (YTH) domain. YTH domain proteins contain long intrinsically disordered regions (IDRs) that may mediate phase separation and interaction with protein partners, but whose precise biochemical functions remain largely unknown. TheArabidopsis thalianaYTH domain proteins ECT2, ECT3 and ECT4 accelerate organogenesis through stimulation of cell division in organ primordia. Here, we focus on ECT2 to reveal molecular underpinnings of this function of ECT2/3/4. We show that stimulation of leaf formation requires the long N-terminal IDR, and we identify two short IDR-elements required for ECT2-mediated organogenesis. Of these two, a tyrosine-rich 19-amino acid region is necessary for binding to a small subset of proteins that includes the major cytoplasmic poly(A)-binding proteins PAB2, PAB4 and PAB8. Remarkably, overexpression of PAB4 in leaf primordia partially rescues the delayed leaf formation inect2 ect3 ect4mutants, suggesting that the ECT2-PAB2/4/8 interaction on target mRNAs of organogenesis-related genes may overcome limiting PAB concentrations in primordial cells.
TRPS1 modulates chromatin accessibility to regulate estrogen receptor (ER) binding and ER target gene expression in luminal breast cancer cells
Authors: Scott Thomas G. , Sathyan Kizhakke Mattada , Gioeli Daniel , Guertin Michael J.
Published: July 2023

Breast cancer is the most frequently diagnosed cancer in women. The most common subtype is luminal breast cancer, which is typically driven by the estrogen receptorα(ER), a transcription factor (TF) that activates many genes required for proliferation. Multiple effective therapies target this path-way, but individuals often develop resistance. Thus, there is a need to identify additional targets that regulate ER activity and contribute to breast tumor progression. TRPS1 is a repressive GATA-family TF that is overexpressed in breast tumors. Common genetic variants in the TRPS1 locus are associated with breast cancer risk, and luminal breast cancer cell lines are particularly sensitive to TRPS1 knockout. However, we do not know how TRPS1 regulates target genes to mediate these breast cancer patient and cellular outcomes. We introduced an inducible degron tag into the native TRPS1 locus within a luminal breast cancer cell line to identify the direct targets of TRPS1 and determine how TRPS1 mechanistically regulates gene expression. We acutely deplete over eighty percent of TRPS1 from chromatin within 30 minutes of inducing degradation. We find that TRPS1 regulates transcription of hundreds of genes, including those related to estrogen signaling. TRPS1 directly regulates chromatin structure, which causes ER to redistribute in the genome. ER redistribution leads to both repression and activation of dozens of ER target genes. Downstream from these primary effects, TRPS1 depletion represses cell cycle-related gene sets and reduces cell doubling rate. Finally, we show that high TRPS1 activity, calculated using a gene expression signature defined by primary TRPS1-regulated genes, is associated with worse breast cancer patient prognosis. Taken together, these data suggest a model in which TRPS1 modulates the activity of other TFs, both activating and repressing transcription of genes related to cancer cell fitness.
Bestrophin-4 relays Hes4 and interacts with Twist1 to suppress epithelial-to-mesenchymal transition in colorectal cancer cells
Authors: Wang Zijing , Xia Bihan , Qi Shaochong , Zhang Xian , Zhang Xiaoshuang , Li Yan , Wang Huimin , Zhang Miao , Zhao Ziyi , Kerr David , Yang Li , Cai Shijie , Yang Jinlin
Published: July 2023

Abstract

Bestrophin isoform 4 (BEST4) is a newly identified subtype of the calcium-activated chloride channel family. Analysis of colonic epithelial cell diversity by single cell RNA-sequencing has revealed the existence of a cluster ofBEST4+ mature colonocytes in humans. However, if the role ofBEST4is involved in regulating tumour progression remains largely unknown. In this study, we demonstrate thatBEST4overexpression attenuates cell proliferation, colony formation, and mobility in colorectal cancer (CRC)in vitro, and impedes the tumor growth and the liver metastasisin vivo.BEST4is coexpressed with hairy/enhancer of split 4 (Hes4) in the nucleus of cells, andHes4signalsBEST4by interacting with the upstream region of theBEST4promoter.BEST4is epistatic toHes4and downregulates Twist1, thereby inhibiting epithelial-to-mesenchymal transition (EMT) in CRC. Conversely, knockout of BEST4 using CRISPR/Cas9 in CRC cells revitalises tumor growth and induces EMT. Furthermore, the low level of theBEST4mRNA is correlated with advanced and the worse prognosis, suggesting its potential role in regulating CRC progression.
Histone deacetylation and cytosine methylation compartmentalize heterochromatic regions in the genome organization ofNeurospora crassa
Authors: Scadden Ashley W. , Graybill Alayne S. , Hull-Crew Clayton , Lundberg Tiffany J. , Lande Nickolas M. , Klocko Andrew D.
Published: July 2023

Chromosomes must correctly fold in eukaryotic nuclei for proper genome function. Eukaryotic organisms hierarchically organize their genomes, including in the fungusNeurospora crassa, where chromatin fiber loops compact into Topologically Associated Domain (TAD)-like structures formed by heterochromatic region aggregation. However, insufficient data exists on how histone post-translational modifications, including acetylation, affect genome organization. In Neurospora, the HCHC complex (comprised of the proteins HDA-1, CDP-2, HP1, and CHAP) deacetylates heterochromatic nucleosomes, as loss of individual HCHC members increases centromeric acetylation and alters the methylation of cytosines in DNA. Here, we assess if the HCHC complex affects genome organization by performing Hi-C in strains deleted of thecdp-2orchapgenes. CDP-2 loss increases intra– and inter-chromosomal heterochromatic region interactions, while loss of CHAP decreases heterochromatic region compaction. Individual HCHC mutants exhibit different patterns of histone post-translational modifications genome-wide: without CDP-2, heterochromatic H4K16 acetylation is increased, yet smaller heterochromatic regions lose H3K9 trimethylation and gain inter-heterochromatic region interactions; CHAP loss produces minimal acetylation changes but increases heterochromatic H3K9me3 enrichment. Loss of both CDP-2 and the DIM-2 DNA methyltransferase causes extensive genome disorder, as heterochromatic-euchromatic contacts increase despite additional H3K9me3 enrichment. Our results highlight how the increased cytosine methylation in HCHC mutants ensures genome compartmentalization when heterochromatic regions become hyperacetylated without HDAC activity.

Significance Statement

The mechanisms driving chromosome organization in eukaryotic nuclei, including in the filamentous fungusNeurospora crassa, are currently unknown, but histone post-translational modifications may be involved. Histone proteins can be acetylated to form active euchromatin while histone deacetylases (HDACs) remove acetyl marks to form silent heterochromatin; these heterochromatic regions cluster, forming strong interactions, in Neurospora genome organization. Here, we show that mutants of a heterochromatin-specific HDAC, HCHC, increase heterochromatic histone acetylation genome-wide and contact probability between distant heterochromatic loci. HCHC loss also impacts cytosine methylation, and in strains lacking both the HCHC and cytosine methylation, heterochromatic regions interact more with euchromatin. Our results suggest cytosine methylation normally functions to segregate silent and active loci when heterochromatic acetylation increases.
Identifying novel age-modulating compounds and quantifying cellular aging using novel computational framework for evaluating transcriptional age
Authors: Zhang Chao , Saurat Nathalie , Cornacchia Daniela , Chung Sun Young , Sikder Trisha , Minotti Andrew , Studer Lorenz , Betel Doron
Published: July 2023

The differentiation of human pluripotent stem cells (hPSCs) provides access to most cell types and tissues. However, hPSC-derived lineages capture a fetal-stage of development and methods to accelerate progression to an aged identity are limited. Understanding the factors driving cellular age and rejuvenation is also essential for efforts aimed at extending human life and health span. A prerequisite for such studies is the development of methods to score cellular age and simple readouts to assess the relative impact of various age modifying strategies.

Here we established a transcriptional score (RNAge) in young versus old primary fibroblasts, frontal cortex and substantia nigra tissue. We validated the score in independent RNA-seq datasets and demonstrated a strong cell and tissue specificity. In fibroblasts we observed a reset of RNAge during iPSC reprogramming while direct reprogramming of aged fibroblasts to induced neurons (iN) resulted in the maintenance of both a neuronal and a fibroblast aging signature. Increased RNAge in hPSC-derived neurons was confirmed for several age-inducing strategies such as SATB1 loss, progerin expression or chemical induction of senescence (SLO). Using RNAge as a probe set, we next performed an in-silico screen using the LINCS L1000 dataset. We identified and validated several novel age-inducing and rejuvenating compounds, and we observed that RNAage captures age-related changes associated with distinct cellular hallmarks of age. Our study presents a simple tool to score age manipulations and identifies compounds that greatly expand the toolset of age-modifying strategies in hPSC derived lineages.
SGLT2 transcriptomic expression atlas supports a kidney-centric role for empagliflozin’s benefits in heart failure
Authors: Mourad Omar , Vohra Shabana , Nunes Sara S
Published: July 2023

Sodium-glucose cotransporter 2 inhibitors (SGLT2i), such as empagliflozin, have shown remarkable benefits in reducing cardiovascular events and mortality in patients with heart failure (HF) irrespective of diabetic status. Because of the magnitude of the benefits and broad application in both HF with reduced and preserved ejection fraction (EF), there have been concerted efforts to identify a mechanism for the observed benefits. One hypothesis is that SGLT2i act directly on the heart. Given empagliflozin’s high specificity to SGLT2, we reasoned that SGLT2 expression would be a requirement for cells to respond to treatment. Here, we present a comprehensive transcriptomic analysis ofSLC5A2, which encodes SGLT2, at the single cell level in multiple datasets, confirmingSLC5A2expression in a subset of kidney epithelial cells but no meaningful expression in other cell types. This was true irrespective of developmental stage, disease state, sequencing method or depth, and species. Our findings support a kidney-centric role for the cardiovascular improvements reported in patients treated with SGLT2i.