title: SARS-CoV-2 mRNA vaccine is re-adenylated in vivo, enhancing antigen production and immune response

city: Warsaw

first: Andrzej

last: Dziembowski

address: 4 Ks. Trojdena Street

country: Poland

institute: International Institute Of Molecular And Cell Biology In Warsaw

summary: Though mRNA vaccines against COVID-19 have revolutionized vaccinology and have been administered in billions of doses, we know incredibly little about how mRNA vaccines are metabolized in vivo. Here we implemented enhanced nanopore Direct RNA sequencing (eDRS), to enable the analysis of single Moderna’s mRNA-1273 molecules, giving in vivo information about the sequence and poly(A) tails. We show that mRNA-1273, with all uridines replaced by N1-methylpseudouridine (mΨ), is terminated by a long poly(A) tail (~100 nucleotides) followed by an mΨCmΨAG sequence. In model cell lines, mRNA-1273 is swiftly degraded in a process initiated by the removal of mΨCmΨAG, followed by CCR4-NOT-mediated deadenylation. In contrast, intramuscularly inoculated mRNA-1273 undergoes more complex modifications. Notably, mRNA-1273 molecules are re-adenylated after mΨCmΨAG removal. Detailed analysis of immune cells involved in antigen production revealed that in macrophages, after mΨCmΨAG removal, vaccine mRNA is very efficiently re-adenylated, and poly(A) tails can reach up to 200A. In contrast, in dendritic cells, vaccine mRNA undergoes slow deadenylation-dependent decay. We further demonstrate that enhancement of mRNA stability in macrophages is mediated by TENT5 poly(A) polymerases, whose expression is induced by the vaccine itself. Lack of TENT5-mediated re-adenylation results in lower antigen production and severely compromises specific immunoglobulin production following vaccination. Together, our findings provide an unexpected principle for the high efficacy of mRNA vaccines and open new possibilities for their improvement. They also emphasize that, in addition to targeting a protein of interest, the design of mRNA therapeutics should be customized to its cellular destination.

sample_organism:
Mus musculus
synthetic construct

platform_organism:
Mus musculus
synthetic construct

overall_design: mRNA profiles from mRNA-1273 injection sites, 2, 8, and 24h after after mice immunization, generated by deep sequencing, in duplicate or triplicate, using Illumina NovaSeq 6000

first: Pawel

last: Krawczyk

first: Olga

last: Gewartowska

first: Michał

last: Mazur

first: Wiktoria

last: Orzeł

first: Seweryn

last: Mroczek


title: IL-23 tunes inflammatory functions of human MAIT cells

status: Public on Jan 09 2025

city: Paris

first: Sebastien

last: Mella

address: 28 rue du Dr Roux

department: Immunology

country: France

institute: Institut Pasteur

type: Expression profiling by high throughput sequencing

summary: IL-23 signaling plays a key role in the pathogenesis of chronic inflammatory and infectious diseases, yet the cellular targets and signaling pathways affected by this cytokine remain poorly understood. We show that IL-23 receptors are expressed on the large majority of human MAIT, but not of conventional T cells, suggesting that these innate-like T cells are critical mediators of IL-23 functions.  In this study, we investigated the effects of IL-23 on human MAIT cell transcriptional and chromatin accessibility profiles, using RNA- and CITE-seq and ATAC-seq, respectively. Protein and transcriptional profiling at the population and single cell level demonstrates that stimulation with IL-23 or the structurally related cytokine IL-12 drives distinct functional profiles, revealing a high level of plasticity of MAIT cells. IL-23, in particular, affects key molecules and pathways related to autoimmunity and cytotoxic functions. Integrated analysis of transcriptomic and chromatin accessibility shows that AP-1 transcription factors constitute a key regulatory node of the IL-23 pathway in MAIT cells.

overall_design: PBMC were isolated from healthy donors. MAIT cells were activated for 6 days in PBMC cultures using MR1 tetramers loaded with 5-OP-RU (MR1/5-OP-RU), in the absence or presence of IL-23, or IL-12. After 6 days, MAIT cells were sorted and we performed RNA-seq, ATAC-seq, and CITE-seq. To identify the effects of IL-23 or IL-12 on MAIT cells, we compared cells activated in the presence of IL-23 or IL-12 with those stimulated with MR1/5-OP-RU only.

first: Laetitia

last: Camard

first: Tharshana

last: Stephen

first: Hanane

last: Yahia-Cherbal

first: Vincent

last: Guillemot

first: Sébastien

first: Victoire

last: Baillet

first: Hélène

last: Lopez-Maestre

first: Daniele

last: Capocefalo

first: Laura

last: Cantini

first: Claire

last: Leloup

first: Julie

last: Marsande

first: Juan

last: Sienes Bailo

first: Ambre

last: Dangien

first: Natalia

last: Pietrosemoli

first: Milena

last: Hasan

first: Huimeng

last: Wang

first: Sidonia

middle: B

last: Eckle

first: Anne

middle: M

last: Fourie

first: Carrie

last: Greving

first: Barbara

middle: J

last: Shaikh

first: Raphaelle

last: Parker

first: Daniel

last: Cua

first: Elisabetta

last: Bianchi

first: Lars

last: Rogge


title: Integrated Analysis of Chromatin and Transcriptomic Profiling of Striatum After Cerebral Hypoperfusion in Mice (RNA-Seq)

status: Public on Jan 08 2025

city: Shanghai

first: Lei

last: Zhu

address: No. 12, Wulumuqi Road (M)

department: Department of Vascular Surgery

country: China

institute: Huashan Hospital, Fudan University

summary: Vascular cognitive impairment (VCI) is a major cause of dementia, characterized by cognitive dysfunction resulting from cerebrovascular disease. Chronic cerebral hypoperfusion (CCH) is a significant contributor to VCI, impacting regions such as the white matter and the hippocampus. Despite the known effects on these areas, the response of the striatum to CCH has been less explored. This study aims to elucidate the molecular and cellular mechanisms underlying striatal dysfunction following CCH. Using a mouse model of bilateral common carotid artery stenosis (BCAS), we observed microglial activation in the striatum, suggesting an inflammatory response. We employed RNA-seq and ATAC-seq to integrate transcriptomic and epigenomic data, identifying changes in gene expression and chromatin accessibility associated with CCH. Our analysis revealed upregulation of immune-related pathways and downregulation of neuronal activity pathways, highlighting distinct regulatory mechanisms in response to hypoperfusion. The results provide insights into the pathogenesis of ischemic brain injury and suggest potential therapeutic targets for VCI.

overall_design: In this study, the samples included were striatum tissues from mice subjected to BCAS-induced hypoperfusion and sham-operated control mice. The experimental conditions involved inducing chronic cerebral hypoperfusion to mimic conditions of vascular cognitive impairment. Key variables under investigation included changes in gene expression and chromatin accessibility in the striatum, analyzed through RNA-seq and ATAC-seq. This approach aimed to uncover molecular signatures and epigenetic modifications associated with striatal dysfunction and neuroinflammation following CCH, focusing on the role of microglial activation and immune response pathways.

first: Shijia

last: Le


title: Newly Found Rat CD103− Dendritic Cells Represent a Highly Immunogenic Subpopulation of Type-2 Conventional Dendritic Cells, Corresponding to Known Dendritic Cell Subsets in Mice and Humans

city: Shimotuga

first: Yasushi

last: Sawanobori

state: Tochigi

address: Kitakobayashi 880, Mibu

country: Japan

institute: Dokkyo medical university

summary: Dendritic cells (DCs), the primary antigen-presenting cells, have traditionally been identified by CD103 molecules in rats, whereas mouse and human DCs are identified by CD11c molecules. However, this history does not preclude the existence of CD103- DCs in rats. To explore this possibility, we examined MHCII+ cells in rat spleen and thymus, identifying a novel population of CD103-MHCII+CD45R-CD172a+ cells. These cells are negative for CD103 and B cell marker CD45R, but positive for the type-2 conventional DC (cDC2) marker CD172a. Transcriptomic analyses revealed that they represent a subpopulation of cDC2.

overall_design: cDC1, cDC2, newly found CD103- dendritic cells (newAPC), and HIS36+ macrophages were sorted from thymic and spleens of male Lewis strain rats. mRNA was extracted and analyzed with Clariom™ S Assay, rat.

first: Tadayuki

last: Ogawa

first: Hisashi

last: Ueta

first: Yusuke

last: Kitazawa

first: Nobuko

last: Tokuda


title: Acclimation of the Escherichia coli transcriptome after 10 hours of incubation in the presence of gallium nitrate at sublethal levels.

city: Calgary

first: Daniel Armando

last: Salazar-Alemán

state: Alberta

address: 2500 University Drive NW

department: Department of Biological Sciences

country: Canada

institute: University of Calgary

summary:
Gallium nitrate has been explored as an FDA-approved alternative to treat antibiotic-resistant bacterial strains. Questions remain as it relates to the full-systems response elicited to counteract its toxic effects.
We explored the transcriptomic response profile of Escherichia coli K12 BW25113 when challenged to grow planktonically for 10 hours in the presence of a sublethal concentration of gallium nitrate (1.25 mM, 319.6 µg/mL).
Our results yielded significant changes in gene expresson levels: 581 genes were up-regulated, while 791 were down-regulated in our gallium nitrate treatment. Several biological processes became dysregulated to cope with the presence of this metal stressor, including iron homeostasis, sulfate metabolism, oxidative and nitrosative stress response, cysteine biosynthesis, anaerobic respiration, toxin-antitoxin interactions, and DNA repair.
Altogether, this work provides a valuable snapshot of how our indicator strain adapts to this metal-induced stress. This is a significant step in understanding how bacteria can adjust their physiology to coexist with sublethal concentrations of metal-based antimicrobials.

overall_design: Two treatment groups were set up: a non-metal control with M9-glucose minimal media, and a metal challenge using the same culture media but spiked with a sublethal concentration of gallium nitrate (1.25 mM, 319.6 µg/mL). Both treatments were inoculated with Escherichia coli K12 BW25113, with three biological trials each. RNA extraction took place after an incubation time of 10 hours at 37 ºC 150 rpm. Differential gene expression was identified by contrasting both treatments, using p-adj value < 0.05 and absolute fold-change (FC) of at least 2 (|log2 FC| > 1) as significance thresholds.

middle: A

first: Raymond

last: Turner


title: Eighteen-year survival after GD2-directed Chimeric Antigen Receptor-Modified Immune Effector Cell Treatment for Neuroblastoma

city: Houston

first: Helen

last: Heslop

address: 1102 Bates Ave

country: USA

institute: Baylor College of Medicine

summary: We report long-term outcomes up to 18 years of a clinical trial treating children with neuroblastoma with EBV-specific T lymphocytes and CD3-activated T cells – each expressing chimeric antigen receptors (CAR) targeting GD2 but without an embedded costimulatory sequence (1st generation CARs). These CARs incorporated barcoded sequences to track each infused population. Of 11 patients with active disease at infusion, three patients achieved a complete response that was sustained in 2, one for 8 years until lost to follow up and one for 18+ years. Of eight patients with no evidence of disease at time of CAR-T administration, five are disease-free at their last follow-up between 10-15 years post-infusion. Intermittent low levels of transgene were detected during the follow up period with significantly greater persistence in those who were long-term survivors. In conclusion, despite using first-generation vectors that are nowadays no longer employed because of the lack of costimulatory domains, patients with relapsed/refractory neuroblastoma achieved long-term disease control after receiving GD2 CAR-T cell therapy including one patient now in remission of relapsed disease for >18 years.

overall_design: Single cell mRNA were extracted from three long-term survivors (#1144, 1290, and 1632)' both chimeric antigen receptor-activated T cells (CAR-ATCs) and EBV virus specific T cells (CAR-VSTs) products. Joint scRNA- and scTCR libraries were prepared using the 10x Genomics platform.

first: Che

middle: H

last: Li

first: Sandhya

last: Sharma

first: Andras

last: Heczey

first: Mae

last: Woods

first: David

last: Steffin

first: Chrystal

last: Louis

first: Bambi

last: Grilley

first: Sachin

last: Thakkar

first: Mengfen

last: Wu

first: Tao

last: Wang

first: Cliona

last: Rooney

first: Malcolm

last: Brenner


title: Integrated Analysis of Chromatin and Transcriptomic Profiling of Striatum After Cerebral Hypoperfusion in Mice (ATAC-Seq)

first: Lei

last: Zhu

type: Genome binding/occupancy profiling by high throughput sequencing

first: Shijia

last: Le


title: Ectopic expression of DNMT3L in human trophoblast stem cells restores features of the placental methylome.

city: Cambridge

first: Sarah

last: Inglesfield

address: Babraham Research Campus

department: Bioinformatics

country: United Kingdom

institute: The Babraham Institute

type: Methylation profiling by high throughput sequencing

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Deep learning imputes DNA methylation states in single cells and enhances the detection of epigenetic alterations in schizophrenia

city: BALTIMORE

first: Jiyun

last: Zhou

state: MD

address: 855 N Wolfe St Suite 300

institute: The Lieber Institute for Brain Development

summary: DNA methylation (DNAm) is a crucial epigenetic mark and plays a key role in gene regulation, mammalian development, and various human diseases. Single-cell technologies enable profiling of DNAm states at cytosines within the DNA sequence of individual cells, but they often suffer from limited coverage of CpG sites. In this study, we introduce scMeFormer, a transformer-based deep learning model designed to impute DNAm states for each CpG site in single cells. Through comprehensive evaluations, we demonstrate the superior performance of scMeFormer compared to alternative models across five single-nucleus DNAm datasets from the human brain and mouse generated by distinct technologies. Remarkably, scMeFormer exhibits high-fidelity imputation, even when dealing with significantly reduced coverage, as low as 10% of the original CpG sites. We applied scMeFormer to a single-nucleus DNAm dataset generated from the prefrontal cortex of four schizophrenia patients and four neurotypical controls. This enabled the identification of thousands of differentially methylated regions associated with schizophrenia that would have remained undetectable without imputation and added granularity to our understanding of epigenetic alterations in schizophrenia within specific cell types. Our study highlights the power of deep learning in imputing DNAm states in single cells, advancing our understanding of the epigenetic bases of schizophrenia within specific brain cell types.

overall_design:
Single-nucleus profiling of the human prefrontal cortex in four schizophrenia patients and four neurotypical controls

***************************************************************
Raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access.
***************************************************************

first: Chongyuan

last: Luo

first: Hanqing

last: Liu

first: Matthew

middle: G

last: Heffel

first: Richard

middle: E

last: Straub

first: Joel

last: Kleinman

first: Thomas

last: Hyde

first: Joseph

middle: R

last: Ecker

first: Daniel

last: Weinberger

first: Shizhong

last: Han


title: Immediate early splicing controls de novo protein synthesis after T cell activation

city: Berlin

first: Marco

last: Preussner

address: Takustrasse 6

country: Germany

institute: FU berlin

summary: Immediate early genes have been known and studied for decades. Their fast and transient induction, which depends on phosphorylation cascades and is independent of de novo protein synthesis, orchestrates the cellular response to various stimuli. In this study we show that the same phosphorylation cascades also target the splicing machinery to induce an analogous splicing switch that we call immediate early splicing (IES). We characterize hnRNPC2-controlled IES, which depends on the MEK-ERK pathway and the T cell specific PKCθ. This splicing switch mainly targets components of the translation machinery and contributes to reduced de novo protein synthesis early upon T cell activation, to coordinate extensive transcriptional and posttranscriptional changes in gene expression. Together, our data set a paradigm for fast and transient alternative splicing, the immediate cellular response to activation, and provide evidence for its functional relevance during T cell activation.

overall_design: Jurkat T cells were stimulated with PMA for 0, 30 and 120 minutes. Nascent RNA was analyzed by RNA-seq. Additionally, Hek or HeLa cells treated with a morpholino to change the hnRNPC1/C2 ratio and polyA+ RNA was investigated by RNA-seq.

first: Florian

last: Heyd


title: EVG1 regulates vascular development and regeneration in response to stress

city: Uppsala

first: Ai

last: Zhang

state: Uppsala

address: Biocenter, Almas allé 5

department: Plant Biology

country: Sweden

institute: SLU

summary: Vascular tissues possess remarkable regenerative abilities, quickly healing post damage from wounding, grafting, or drought. The regenerative ability is also relevant for many plant pathogens that invade host tissues and induce the formation of vasculature to derive nutrients, thereby suggesting a possible mechanistic overlap in vascular regeneration between stresses. With comparative transcriptomics we identified a gene that was highly upregulated in Agrobacterium infection, nematode infection, and during graft formation. Loss of function mutant of EVG1 enhanced ectopic xylem formation in Vascular cell Induction culture System Using Arabidopsis Leaves (VISUAL) and enhanced graft formation and was termed ENHANCER OF VISUAL AND GRAFTING 1 (EVG1). EVG1 promoted cambium development and inhibited vascular regeneration during grafting. Transcriptomic analyses found that stress responsive, cell wall related, and ABA response related genes were differentially expressed in evg1. Mutants of EVG1 also phenocopied mutants of RLP44, a receptor that interacts with the receptors of both the brassinosteroid (BR) and phytosulfokine (PSK) signaling pathway to incorporate cell wall damages cues into hormone signaling and help maintain procambium cell identity. We identified that EVG1 is highly stress responsive while RLP44 is not despite showing similar phenotypes in developmental assays. We propose that EVG1 and RLP44 act in the same pathway. EVG1 might act as a stress signal and regulates development while indirectly also influencing the RLP44 associated pathways.

overall_design: The  transcriptomic dataset consists of 12 samples, including Col-0, evg1-1, EVG1-OE,rlp44-3. Each genotype contains intact seedlings, each genotype is represented by three biological replicates.

first: Shamik

last: Mazumdar

first: Constance

last: Musseau

first: Muhammad

middle: S

last: Anjam

first: Peter

last: Marhavy

first: Charles

middle: W

last: Melnyk


title: Expression data from RD cells

city: Solna

first: Monika

last: Ehnman

address: Karolinska University Hospital, Akademiska stråket 1

department: Oncology-Pathology

institute: Karolinska Institutet

summary:
Mesenchymal stromal cells belong to a diverse collection of cells in different states that are poorly characterized in soft-tissue sarcomas.
We used microarrays to explore differentially expressed genes in pediatric rhabdomyosarcoma cells (RD) after exposure to non-malignant mesenchymal stromal cells (4 replicates/condition).

overall_design: Conditioned medium (cell culture supernatant from mouse muscle stromal cells in adherent culture) was applied on RD cells. Gene expression changes compared to control medium were analyzed 16 hours later.


title: Ectopic expression of DNMT3L in human trophoblast stem cells restores features of the placental methylome [PBAT BS-Seq]

first: Sarah

last: Inglesfield

summary: The placental DNA methylation landscape is unique, with widespread partially methylated domains (PMDs) and hundreds of placental-specific imprinted domains. Furthermore, the placental ‘methylome’ has been the focus of extensive study for links with pregnancy complications. Human trophoblast stem cells (hTSCs) offer exciting potential for functional epigenetic studies; however, whether the hTSC epigenome recapitulates primary cytotrophoblast, remains poorly described. In this study, we demonstrate that hTSCs exhibit an atypical methylome, with DNA methylation present over transcribed gene bodies but a complete loss of placental PMDs. Using single-cell RNA-seq from human embryogenesis, hTSCs display a notable absence of DNMT3L expression. Ectopic expression of DNMT3L in hTSCs restored placental PMDs. DNMT3L-expressing hTSCs showed comparable stemness but failed to syncytialise in organoid culture, associated with hypermethylation of STB transcription factor motifs. These findings reveal that DNMT3L is essential in establishing the human placental methylome and that DNMT3L downregulation is necessary for successful trophoblast differentiation.

overall_design: To understand determinants of the DNA methylation landscape in hTSCs, we cultured blastocyst- (BTS11) and 1st trimester cytotrophoblast- (CT29) derived hTSCs in 2D conditions (Okae et al., 2018), 2D in organoid media, 3D as organoids (Sheridan et al., 2021), and as organoids cultured in hypoxic conditions. We hypothesized that the absence of DNMT3L from cultured hTSCs results in almost exclusive recruitment of DNMT3B to gene bodies leading to a loss of inter-genic partially methylated domains. To test this, we conducted overexpression (OE) experiments in BTS11 and CT29 hTSCs through lentiviral transduction of DNMT3L, in comparison to a control plasmid. Using post-bisulphite adaptor tagging (PBAT) bisulphite-sequencing, we assayed genome-wide DNA methylation patterns in each cell line and condition and compared to published datasets.

first: Courtney

last: Hanna

first: Georgia

last: Lea

first: Vicente

last: Perez-Garcia


title: Mitochondria-enriched hematopoietic stem cells exhibit elevated self-renewal capabilities, thriving within the context of aged bone marrow [bulk RNA-seq]

city: Singapore

first: Takayoshi

last: Matsumura

address: 14 Medical Drive #12-01

country: Singapore

institute: National University of Singapore

summary: The aging of hematopoietic stem cells (HSCs) significantly alters their characteristics. Mitochondria, essential for cellular metabolism, play a crucial role, and their dysfunction is a hallmark of aging-induced changes. The impact of mitochondrial mass on aged HSCs remains incompletely understood. Here, we demonstrate that HSCs with high mitochondrial mass during aging are not merely cells that have accumulated damaged mitochondria and become exhausted. Instead, these HSCs retain a high regenerative capacity and remain in the aging bone marrow. Furthermore, we identified GPR183 as a novel marker characterizing aged HSCs through single-cell analysis. HSCs marked by GPR183 were also enriched in aged HSCs with high mitochondrial mass, possessing a high capacity of self-renewal. These insights deepen our understanding of HSC aging and provide new perspectives on the assessment of aged HSCs, underscoring the importance of mitochondrial dynamics in the aging.

overall_design: Bulk RNA-seq of LT-HSCs, ST-HSCs, and MPPs of 4 young (12 weeks old) and 4 aged (107 weeks old) male mice

first: Toshio

last: Suda


title: Mitochondria-enriched hematopoietic stem cells exhibit elevated self-renewal capabilities, thriving within the context of aged bone marrow [single cell multiome]

overall_design: Single cell mutiome analysis (single cell RNA-seq and single cell ATAC-seq)  of young and aged mitoDendra2 Low and High HSCs


title: Single nuclear gene expression profile of postnatal day 1 mouse aortic valve structures

city: Milwaukee

first: Joy

last: Lincoln

state: WI

address: 8701 Watertown Plank Road

department: Pediatrics

institute: Medical College of Wisconsin

summary: The heterogenity of aortic valve cells  present at birth which contruibute to during postnatal maturation is unknown. We utilized single nuclear gene expression profiling to understand the transcritptionally diverse cell type subpopulations present at postnatal day which contribute to postnatal valve maturation.

overall_design: Aortic valve structures were issolated from postnatal day 1 mouse pups and flash frozen, valve structures were pooled (n=3 pooled aortic structures replicates, 22-24 aortic roots per replicate). Nuclei were issolated and subjected to single nuclear gene expression profiling utilizing 10X genomics technology.

first: Theresa

last: Bluemn

first: Jenny

last: Drnevich


title: ZNFX1 Functions as a Master Regulator of Epigenetically Induced Pathogen Mimicry and Inflammasome Signaling in Cancer.

status: Public on Jan 07 2025

city: Baltimore

first: Michael

last: Topper

address: 1650 Orleans St.

institute: Johns Hopkins University, Sidney Kimmel Comprehensive Cancer Center

summary: DNA methyltransferase and poly (ADP-ribose) polymerase inhibitors (DNMTis, PARPis) induce a stimulator of interferon genes (STING)-dependent pathogen mimicry response (PMR) in ovarian and other cancers. Here, we showed that combining DNMTis and PARPis upregulates expression of the nucleic-acid sensor NFX1-type zinc finger-containing 1 protein (ZNFX1). ZNFX1 mediated induction of PMR in mitochondria, serving as a gateway for STING-dependent interferon/inflammasome signaling. Loss of ZNFX1 in ovarian cancer cells promoted proliferation and spheroid formation in vitro and tumor growth in vivo. In patient ovarian cancer databases, expression of ZNFX1 was elevated in advanced stage disease, and ZNFX1 expression alone significantly correlated with an increase in overall survival in a phase 3 trial for therapy-resistant ovarian cancer patients receiving bevacizumab in combination with chemotherapy. RNA-sequencing revealed an association between inflammasome signaling through ZNFX1 and abnormal vasculogenesis. Together, this study identified that ZNFX1 as a tumor suppressor that controls PMR signaling through the mitochondria and may serve as a biomarker to facilitate personalized therapy in ovarian cancer patients.

overall_design: RNA-seq profiling of the ovarian cancer cell line TYK-nu comparing WT vs ZNFX1 KO conditions assessed in biological triplicate.

first: Lora

last: Stojanovic

first: Rachel

last: Abbotts

first: Stephen

last: Baylin

first: Kenneth

last: Nephew

first: Feyruz

last: Rassool


title: Single-nuclei RNA sequencing analysis from a WT mouse brain cortex after Intrathecal(IT) injection of AAV9/JeT-hSLC6A1 vector

city: Dallas

first: Weirui

last: Guo

state: Texas

address: 6000 Harry Hines Blvd.

institute: UT Southwestern Medical Center

summary: Single-nuclei RNA sequencing analysis conducted at 6 months old from a WT mouse brain cortex, following Intrathecal(IT) injection of AAV9/JeT-hSLC6A1 vector at PND10.

overall_design: WT C57BL/6J mouse was IT injected with 5 µL AAV9/ JeT-SLC6A1 vector at postnatal 10 days old. At 6 months post-injection, treated mouse was perfused, the brain cortex tissue was collected and processed for Single-nuclei RNA sequencing analysis (10x Genomics)

first: Matthew

last: Rioux

first: Frances

last: Shaffo

first: Yuhui

last: Hu

first: Ze

last: Yu

first: Chao

last: Xing

first: Steven

last: Gray


title: Impacts of perR on oxygen sensitivity, gene expression, and murine infection in Clostridioides difficile 630∆erm

city: Madison

first: Anna

last: Gregory

address: 1550 Linden Dr., MSB Rm3435

department: Department of Gastroenterology and Department of Medical Microbiology and Immunology

institute: University of Wisconsin-Madison

summary: Clostridioides difficile infection (CDI), characterized by colitis and diarrhea, afflicts approximately half a million people in the United States every year, burdening both individuals and the healthcare system. C. difficile 630Δerm is an erythromycin-sensitive variant of the clinical isolate C. difficile 630 and is commonly used in the C. difficile research community due to its genetic tractability. 630Δerm possesses a point mutation in perR, an autoregulated transcriptional repressor that regulates oxidative stress resistance genes. This point mutation results in a constitutively de-repressed PerR operon in 630Δerm. To address the impacts of perR on phenotypes relevant for oxygen tolerance and relevant to a murine model of CDI, we corrected the point mutant to restore PerR function in 630∆erm (herein, 630∆erm perRWT). We demonstrate that there is no difference in growth between 630Δerm and a 630Δerm perRWT under anaerobic conditions or when exposed to concentrations of O2 that mimic those found near the surface of the colonic epithelium. However, 630∆erm perRWT is more sensitive to ambient oxygen than 630∆erm, which coincides with alterations in expression of a variety of perR-dependent and perR-independent genes. Finally, we show that 630∆erm and 630∆erm perRWT do not differ in their ability to infect and cause disease in a well-established murine model of CDI. Together, these data support the hypothesis that the perR mutation in 630∆erm arose as a result of exposure to ambient oxygen and that the perR mutation in 630∆erm is unlikely to impact CDI-relevant phenotypes in laboratory studies

overall_design: C. difficile 630Δerm and 630Δerm perRWT overnight cultures were back-diluted 1:100 into 35 mL of pre-reduced mRCM in Erlenmeyer flasks and incubated anaerobically at 37°C until cultures reached mid-log phase (OD600=0.3-0.4). At mid-log phase, 5 mL aliquots of the cultures were diluted 1:1 in chilled 1:1 ethanol:acetone and stored at -20° to preserve RNA for 0 minute timepoints. The remaining cultures were aerobically shaken (220 rpms) at 37°C for 60 minutes. After 60 minutes, 5 mL aliquots were diluted 1:1 in chilled 1:1 ethanol:acetone and stored at -20° to preserve RNA for the 60 minute timepoints. RNA was extracted by centrifuging samples at 3,000 xg for 5 minutes at 4°C. Pellets were washed with 5 mL cold, nuclease free PBS, and centrifuged at 3,000 xg for 5 minutes at 4°C. The supernatant was removed, and remaining pellets were resuspended in 1 mL TRIzol and processed using a TRIzol Plus RNA Purification Kit (Thermo) with on-column DNase treatment. Purified RNA integrity was confirmed via 2100 Agilent BioAnalyzer and frozen at -80°C. RNA-seq was performed by Microbial Genome Sequencing Center (MiGS) on high-quality rRNA-depleted RNA extracts (12 M paired end reads per sample).

middle: L

first: Hailey

last: Bussan

first: Madeline

last: Topf

first: Andrew

last: Hryckowian


title: Mitochondrial fatty acid oxidation regulates monocytic type I interferon signaling via histone acetylation.

city: Bethesda

first: Michael

last: Sack

address: 10 Center Drive

institute: NHLBI

summary: Although lipid-derived acetyl-CoA is a major carbon source for histone acetylation, the contribution of fatty acid -oxidation (FAO) to this process remains poorly characterized. To investigate this, we generated mitochondrial acetyl-CoA acetyltransferase 1 (ACAT1, distal FAO enzyme) knockout macrophages. 13C-carbon tracing confirmed reduced FA-derived carbon incorporation into histone H3 and RNA-seq identified diminished interferon stimulated gene expression in the absence of ACAT1. Chromatin accessibility at Stat1 locus was diminished in ACAT1-/- cells. CHIP analysis demonstrated reduced acetyl-H3 binding to Stat1 promoter/enhancer regions and increasing histone acetylation rescued Stat1 expression. IFNβ release was blunted in ACAT1-/- and recovered by ACAT1 reconstitution. Furthermore, ACAT1-dependent histone acetylation required an intact acetylcarnitine shuttle. Finally, obese subjects’ monocytes exhibited increased ACAT1 and histone acetylation levels. Thus, our study identifies a novel link between FAO-mediated epigenetic control of type 1 interferon signaling and uncovers a potential mechanistic link between obesity and type I inferon signaling.

overall_design: To investigate role of fatty acid oxidation on immunity, we generated mitochondrial acetyl-CoA acetyltransferase 1 (ACAT1) knockout macrophages.

middle: N

first: Jing

last: Wu


title: tRNA m1A modification regulates cholesterol biosynthesis to promote antitumor immunity of CD8+ T cells

first: Xiaohan

last: Song

state: Shanghai

address: South ChongQing Road 227

institute: Shanghai Jiao Tong University School of Medicine

summary: Activation of CD8+ T cells necessitates rapid metabolic reprogramming to fulfill the substantial biosynthetic demands of their effector functions. However, the post-transcriptional mechanisms underpinning this process remain obscure. The tRNA N1-methyladenine (m1A) modification, which plays a role in maintaining tRNA stability and modulating protein translation, has an undefined physiological function in CD8+ T cells, particularly in antitumor responses. Here, we demonstrate that the tRNA m1A 'writer' gene Trmt61a enhances the tumor-killing capacity of CD8+ T cells by regulating cholesterol biosynthesis. We find that Trmt61a expression in CD8+ T cells is upregulated upon activation and correlates positively with T cell-mediated cytotoxicity within the tumor microenvironment of colorectal cancer (CRC) patients. Deletion of Trmt61a in CD8+ T cells leads to a compromised tumor-killing function in both in vivo and in vitro assays, which is dependent on the m1A catalytic activity of TRMT61A. Mechanistically, tRNA m1A promotes antitumor immunity in CD8+ T cells by enhancing the translation of ATP citrate lyase (ACLY), a key enzyme for cholesterol biosynthesis. Supplementation with cholesterol can rescue the impaired tumor-killing function and proliferation of TRMT61A-deficient CD8+ T cells. Our findings underscore the significance of tRNA m1A modification as a regulatory checkpoint in cholesterol metabolism in CD8+ T cells, suggesting potential novel strategies for cancer immunotherapy.

overall_design: For RNA-seq, there are 0h,48h for WT samples and 0h,48h for Trmt61A-cKO samples.

first: Shan

last: Miao

first: Hao

last: Li

first: Yongbo

last: Liu

first: Gaoyang

last: Wang

first: Chen

last: Kan

first: Youqiong

last: Ye

first: Ru-Juan

first: Hua-Bing


title: An in vitro high-content screening unveils miR-429 as a protective molecule in photoreceptor degeneration models

city: Pozzuoli

first: Santiago

last: Negueruela

address: Via Campi Flegrei, 34

department: Genetic Medicie

country: Italy

institute: Telethon Institute of Genetics and Medicine (TIGEM)

summary: Transcriptome profile of optic cups from RhoP23H/+ mice following AAV-mediated subretinal delivery of pre-miR-429 or GFP (control vector) to study the molecular changes underlying the neuroprotective effect of miR-429 overexpression in an Inherited Retinal Disease (IRD) model.

overall_design: Total RNA was extracted at postnatal day (PN) 30 from optic cups of eyes subretinally injected at PN8 with the AAV8-miR-429 vector; RNA extracted from optic cups of controlateral eyes injected with AAV8-GFP control vector.

first: Georgios

last: Petrogiannakis

first: Irene

last: Guadagnino

first: Martina

last: Di Guida

first: Mariateresa

last: Pizzo

first: Annalaura

last: Torella

last: Zanobio

first: Marianthi

last: Karali

first: Diego

last: Medina

first: Sabrina

last: Carrella

first: Sandro

last: Banfi


title: A SnRK1-JMJ15-CRF6 coregulatory module gages mitochondrial functional status to mitigate growth-defense trade-off in Arabidopsis

city: Kaifeng

first: Xiaohong

last: Zhu

address: Jinming Street

institute: Henan University

summary: Mitochondrion is a signaling organelle sensing environmental changes and integrating cellular responses via retrograde signalling. However, it is still largely unknown how mitochondria help plants cope with adversity while it powers plant growth. In this work, we identify two new targets of plant SNF1-Related Kinase1 (SnRK1), namely histone H3K4me3 demethylase JMJ15 and transcription factor CRF6. We show that JMJ15 and CRF6 are required for reprograming transcription in response to Antimycin A (AA)-induced mitochondrial stress. A functional module of JMJ15 and CRF6 with SnRK1 that measures mitochondrial status is supported by the AA-promoted nuclear translocation of SnRK1a, a-subunit of SnRK1 under mitochondrial stress, and the physical interaction of SnRK1a with JMJ15 and CRF6, and high correlation of transcriptome changes resulted from the genetic inactivation of JMJ15 and CRF6 with the ones from the plant overexpressing SnRK1a under normal and AA stress conditions. We show that SnRK1a phosphorylates and destabilize JMJ15 to inhibit its H3K27me3 demethylase activity. While SnRK1a does not phosphorylated CRF6, it promotes CRF6 degradation via proteasome pathway. CRF6 interacts with JMJ15, and prevents JMJ15 from phosphorylation dependent degradation, offering a feedback loop to antagonize the action of SnRK1a. Our findings indicate a new mechanism in which a histone demethylase mediated epigenetic regulation and a transcription factor govern cytokine and oxidative stress signalling are implicated in SnRK1-integrated transcriptional network to mitigate growth-defense trade-offs under mitochondrial dysfunction.

overall_design: mRNA profiles of 10-day old WT, snrk1a, jmj15, crf6 and SnRK1a overexpression plants were generated by deep sequencing, in triplicate.


title: Histological Signatures Reveal Anti-Fibrotic Factors in Mouse and Human Lungs [scATAC-Seq]

city: Stanford

first: Jason

last: Guo

state: CA

address: 257 Campus Dr

institute: Stanford University

summary: Fibrosis, the replacement of healthy tissue with collagen-rich matrix, can occur following injury in almost every organ. Mouse lungs follow stereotyped sequences of fibrogenesis-to-resolution after bleomycin injury, and we reasoned that profiling post-injury histological progression could uncover pro- vs. anti-fibrotic features with functional value for human fibrosis. We mapped spatiotemporally-resolved transformations in lung extracellular matrix (ECM) architecture to spatially-resolved, multi-omic data. First, we charted stepwise trajectories of matrix aberration vs. resolution using unsupervised machine learning, denoting a reversible transition in uniform-to-disordered histological architecture. Single-cell sequencing along these trajectories identified temporally-enriched “ECM-secreting” (Csmd1+) and “pro-resolving” (Cd248+) fibroblasts, for which Visium inferred divergent histological signatures and spatial-transcriptional “neighborhoods”. Critically, pro-resolving fibroblast instillation helped ameliorate fibrosis in vivo. Further, fibroblast neighborhood-associated moieties, Serpine2 and Pi16, functionally modulated human lung fibrosis ex vivo. Spatial phenotyping of idiopathic pulmonary fibrosis further uncovered analogous fibroblast subtypes and neighborhoods in human disease. Collectively, these findings establish an atlas of pro-/anti-fibrotic factors underlying lung matrix architecture and implicate fibroblast-centered moieties in modulating fibrotic progression vs. resolution.

overall_design: scATAC-seq of unsorted cells from bleomycin-injured mouse lung samples using the 10X genomics platform

first: Michelle

last: Griffin

first: Michael

last: Januszyk

first: Derrick

last: Wan

first: Tushar

last: Desai

last: Longaker


title: Histological Signatures Reveal Anti-Fibrotic Factors in Mouse and Human Lungs [scRNA-seq]

overall_design: scRNA-seq of unsorted cells from bleomycin-injured mouse lung samples using the 10X genomics platform


title: Histological Signatures Reveal Anti-Fibrotic Factors in Mouse and Human Lungs

overall_design: Refer to individual Series


title: BMP-dependent patterning of ectoderm tissue material properties modulates lateral mesendoderm cell migration during early zebrafish gastrulation

city: Klosterneuburg

first: Stefania

last: Tavano

address: Am Campus 1

country: Austria

institute: Institute of Science and Technology Austria (ISTA)

summary: Cell migration is a fundamental process during embryonic development. Most studies in vivo have focussed on the migration of cells using the extracellular matrix (ECM) as their substrate for migration. In contrast, much less is known about how cells migrate on other cells, as found in early embryos when the ECM has not yet formed. Here, we show that lateral mesendoderm (LME) cells in the early zebrafish gastrula use the ectoderm as their substrate for migration. We show that the lateral ectoderm is permissive for the animal pole-directed migration of LME cells, while the ectoderm at the animal pole halts it. These differences in the permissiveness depend on the lateral ectoderm being more cohesive than the animal ectoderm, a property controlled by BMP signalling within the ectoderm. Collectively, these findings identify ectoderm tissue cohesion as one critical factor regulating LME migration during zebrafish gastrulation.

overall_design: To investigate the differences in gene expression between animal and lateral ectoderm causing the difference in material properties between the two ectodermal tissues.

first: David

last: Brueckner

first: Saren

last: Tasciyan

first: Xin

last: Tong

first: Roland

last: Kardos

first: Alexandra

last: Schauer

first: Robert

last: Hauschild

first: Carl-Philipp

last: Heisenberg


title: Ectopic expression of DNMT3L in human trophoblast stem cells restores features of the placental methylome [RNA-seq]

first: Sarah

last: Inglesfield

overall_design: To understand determinants of the DNA methylation landscape in hTSCs, we cultured blastocyst- (BTS11) and 1st trimester cytotrophoblast- (CT29) derived hTSCs in 2D conditions (Okae et al., 2018), 2D in organoid media, 3D as organoids (Sheridan et al., 2021), and as organoids cultured in hypoxic conditions. We hypothesized that the absence of DNMT3L from cultured hTSCs results in almost exclusive recruitment of DNMT3B to gene bodies leading to a loss of inter-genic partially methylated domains. To test this, we conducted overexpression (OE) experiments in BTS11 and CT29 hTSCs through lentiviral transduction of DNMT3L, in comparison to a control plasmid. Using stranded mRNA-sequencing, we assayed gene expression in control and DNMT3L-OE hTSCs to understand the impact of differential methylation on gene regulation.

first: Courtney

last: Hanna

first: Georgia

last: Lea

first: Vicente

last: Perez-Garcia


title: Histological Signatures Reveal Anti-Fibrotic Factors in Mouse and Human Lungs [Visium]

first: Jason

last: Guo

overall_design: Visium of unsorted cells from bleomycin-injured mouse lung samples using the 10X genomics platform

last: Griffin


title: ChIP sequencing of wild-type and TRα (-/-) tail with and without T3 treatment.

first: Ryan

last: Dale

address: Rm 10D39, 10 Center Drive

department: National Institute of Child Health and Human Development

institute: National Institutes of Health

summary: Our study represents the first detailed analysis of thyroid hormone binding to the genome in the tail of wild-type and TRα (-/-) tadpoles, with biological replicates, generated by ChIP-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of TR binding profiles.

overall_design: Examination of 3 technical replicate from the same RNA sample which extracted and merged from the several Xenopus tropicalis tadpoles.

first: Yuta

last: Tanizaki

first: Yun-Bo

last: Shi

first: Hongen

last: Zhang


title: AAV-mediated expression of proneural factors stimulates neurogenesis from adult Müller glia in vivo.

city: Seattle

first: Thomas

last: Reh

state: Washington

address: 1959 NE Pacific Street, UW School of Medicine, Seattle, WA 98195

department: Department of Neurobiology and Biophysics

institute: University of Washington

summary: The lack of regeneration in the human central nervous system (CNS) has major health implications. To address this, we previously used transgenic mouse models to show that neurogenesis can be stimulated in the adult mammalian retina by driving regeneration programs that other species activate following injury. Expression of specific proneural factors in adult Müller glia causes them to re-enter the cell cycle and give rise to new neurons following retinal injury. To bring this strategy closer to clinical application, we now show that neurogenesis can also be stimulated when delivering these transcription factors to Müller glia using adeno-associated viral (AAV) vectors. AAV-mediated neurogenesis phenocopies the neurogenesis we observed from transgenic animals, with different proneural factor combinations giving rise to distinct neuronal subtypes in vivo. Vector-borne neurons are morphologically, transcriptomically and physiologically similar to bipolar and amacrine/ganglion-like neurons. These results represent a key step forward in developing a cellular reprogramming approach for regenerative medicine in the CNS.

overall_design: Viral vectors expressing proneural genes can reliably stimulate glia of the adult mammalian retina to become various subtypes of neurons.

first: Marina

last: Pavlou

first: Marlene

last: Probst

first: Lew

last: Kaplan

first: Elizaveta

last: Filippova

first: Aric

last: Prieve

first: Fred

last: Rieke


title: The Role of TCF7L2 in Hepatic Metabolic Zonation

city: San Antonio

first: Luke

last: Norton

address: 7703 Floyd Curl Dr

institute: UTHSCSA

summary: Background and Aims: The molecular mechanisms regulating the zonal distribution of metabolism in liver are incompletely understood. Here we used multimodal single nuclei genomics techniques to examine the spatial transcriptional function of the transcription factor 7-like 2 (TCF7L2) in rodent liver. We also determined whether inactivation of TCF7L2 disrupted the normal metabolic architecture of the liver and influenced the development of fibrotic liver diseases.      Methods:   Multimodal single nuclei RNA- and ATAC-Seq were used to examine the spatial expression and DNA binding activity of TCF7L2 across the liver lobule. The transcriptional activity of TCF7L2 was targeted by removing exon 11 of Tcf7l2, which encodes part of the DNA binding domain (DBD). The effect of TCF7L2 inactivation on transcriptional regulators of zonation, hepatic metabolism, and the development of fibrosis was investigated in Hep-TCF7L2ΔDBD mice fed the Gubra Amylin Nash (GAN) or choline-deficient amino acid-defined high fat (CDAHFD) diet for 24- and 8-weeks, respectively.  Results:  Tcf7l2 mRNA expression was ubiquitous across the liver lobule, but the presence of the consensus TCF/LEF DNA binding motif in open chromatin was enriched in pericentral (PC) hepatocytes in zone 3, but not periportal (PP) or mid-lobular hepatocytes in zones 1 and 2. Consistent with this, PC hepatocyte gene expression was lost in Hep-TCF7L2ΔDBD mice, which we link to alterations in the transcriptional activity of zonal repressors Tbx3 and Tcf7l1. The absence of PC hepatocyte gene expression resulted in impaired bile acid synthesis and hepatic cholesterol accumulation, and disrupted metabolic pathways linked to ammonia detoxification, most notably glutamine/glutamate homeostasis. Following the CDAHFD, Hep-TCF7L2ΔDBD mice developed more severe hepatic fibrosis. Finally, TCF7L2 expression declined in human livers as the severity of fibrosis progressed.  Conclusion:  TCF7L2 is an important transcriptional regulator of PC hepatocytes in zone 3, and regulates multiple zonated metabolic pathways that may contribute to the development of fibrotic liver diseases.

overall_design: Animal models  Mice with LoxP sites flanking exon 11 of the Tcf7l2 gene, which encodes part of the DNA binding domain of the TCF7L2 protein, have been described previously [23]. Albumin-Cre (Alb-Cre) mice were obtained from The Jackson Laboratory (stock # 003574). Mice were fed either regular chow (Envigo, cat # 7012), a Gubra Amylin NASH (GAN) diet (40% fat, 20% fructose, 2% w/w cholesterol; Research Diets Inc, D09100310) for 24-weeks, or a methionine and choline deficient, 60% HFD diet (CDAHFD, Research Diets cat#: A06071302) for 8-weeks. Tcf7l2 expression was examined in a cohort of rats fed the same CDAHFD diet for 14-weeks, supplemented with NaNO2 (25 mg/kg i.p. three times a week between weeks 4 and 14), and in mice fed a fast food diet (FFD) for 11 months. This consisted of a commercially available high fat, high cholesterol diet (Research Diets Inc, D12079B) and drinking water containing 23.1 g fructose and 17.2 g glucose per 1000 mL of water. The TCF/Lef:H2B-GFP reporter mouse was obtained from The Jackson Laboratory (stock # 013752) and was used to examine TCF/LEF activity in liver sections.   Mice were provided ad-libitum access to acidified reverse osmosis water (pH 2.5 – 3.0) and Nestlets. All mice were housed in ventilated cages (Allentown Caging Equipment Company, cat # 75JAG) with irradiated Sani-Chip bedding in a room maintained at 75 ± 3°F and on a 14 h light/10 h dark cycle. Littermate LoxP/LoxP controls were used in all experiments. All rodent studies, with the exception of those carried out by Gilead Sciences Inc (Mouse FFD and rat CDAHFD), were approved by the UT Health San Antonio Institutional Animal Care and Use Committee (IACUC).   Single-nuclei multiomics  Nuclei were isolated from approximately 20 mg of frozen mouse liver harvested from control (TCF7L2LoxP/LoxP) and liver-specific TCF7L2 mutant mice (Hep-TCF7L2ΔDBD). 10X Genomics Chromium single nuclei multiome gene expression and Assay for Transposase-Accessible Chromatin (ATAC) libraries were prepared from approximately 5000 nuclei/mice at the Genome Sequencing Facility at the University of Texas Health at San Antonio (UTHSA). Liver samples were processed in pairs (one control and one mutant) on three consecutive days to minimize batch effects. Library quality and quantity were assessed using the 2100 Bioanalyzer and were sequenced on a NovaSeq 6000 (Illumina, CA, USA). We performed two distinct analyses that relied on separate clustering. First, we used the control samples to establish the methodology of quantifying zonal gene expression and transcription factor occupancy, including TCF7L2. In a distinct analysis, we then examined the consequences of TCF7L2 transcriptional inactivation on zonation in control vs. mutant livers. A detailed description of the computational methodology used to integrate and analyze the multiomic data described throughout the manuscript is provided in the Supplementary Materials.

first: Iriscilla

last: Ayala

first: Skanda

last: Hebbale

first: Juho

last: Mononen

first: Madelaine

last: Sholto

first: Chris

last: Shannon

first: Ivan

last: Valdez

first: Marcel

last: Fourcadot

first: Terry

last: Bakewell

first: Giovanni

last: Romero

first: Mara

last: Asmis

first: Leandro

last: Freitas

last: Aguiar Vallim

first: Sami

last: Heikkinen


title: Single cell RNA-seq reveals that granulosa cells are a target of phthalate toxicity in the ovary

status: Public on Jan 06 2025

city: Newark

first: Genoa

last: Warner

state: NJ

address: 161 Warren St Tiernan Hall #366 Tiernan Hall #366

institute: New Jersey Institute of Technology

summary: Phthalates are known endocrine disrupting chemicals and ovarian toxicants that are used widely in consumer products. Phthalates have been shown to exert ovarian toxicity on multiple endpoints, altering transcription of genes responsible for normal ovarian function. However, the molecular mechanisms by which phthalates act on the ovary are not well understood. In this study, we hypothesized that phthalates specifically target granulosa cells within the ovarian follicle. To test our hypothesis, we cultured whole mouse antral follicles for 96 hours in the presence of vehicle or 10 ug/mL of a phthalate metabolite mixture. At the end of the culture period, follicles were dissociated into single cell suspensions and subjected to single cell RNA sequencing. We used markers from published studies to identify major cell type clusters, the largest of which were granulosa and theca/stroma cells. We further identified sub-populations of granulosa, theca, and stromal cells and analyzed differentially expressed genes between the phthalate treatment and control. Granulosa cells, specifically mural granulosa cells, had the most differentially expressed genes. Pathway analysis of differentially expressed genes from the overall granulosa cell cluster revealed disruption of cell cycle and mitosis, whereas pathway analysis of the mural granola cell subcluster identified terms related to translation and ribosome. Our findings suggest that phthalates have both broad impacts on major cell types and specific impacts on individual cell types, emphasizing the complexity of phthalate toxicity and highlighting how bulk sequencing can mask effects on vulnerable cell types.

overall_design: Antral follicels from young adult CD-1 mice were cultured for four days in the presence of DMSO (vehicle contro) or 10 ug/mL of phthalate metabolie mixture (n=3 each, 30 follicles/sample). After culture, follicles were subjected to single cell sequencing

first: Erik

last: Mattson


title: Time series of host responses to symbiotic T6SS disruption in an amoeba-bacteria symbiosis

city: Waterville

first: Suegene

last: Noh

state: ME

address: 5717 Mayflower Hill

department: Biology

institute: Colby College

summary: In order to understand host responses to symbiont strains with and without an intact symbiotic type 6 secretion system, we infected Dictyostelium discoideum with wildtype vs. ∆tssH Paraburkholderia bonniea and collected samples in a time series to capture the early stages of symbiotic association.

overall_design: Two host strains of Dictyostelium discoideum amoebas were each infected with wildtype (wt) and ∆tssH (delta) Paraburkholderia bonniea bacteria strain bb433. We collected samples at 90, 360, and 2250 minutes post-infection (mpi). The 2250 mpi sample was intended to represent a longer-term infection and was called the 'positive' sample. Additional samples include uninfected 'negative' controls.


title: Spatial Transcriptomics of IPMN Reveals Divergent Indolent and Malignant Progenitor Phenotypes

city: Durham

first: Peter

last: Allen

state: NC

address: 203 Research Dr., MSRB1 Rm. 475

department: Surgery

institute: Duke Medical Center

summary:
Purpose: Intraductal papillary mucinous neoplasms (IPMN) occur in 5-10% of the population, but only a small minority progress to pancreatic ductal adenocarcinoma (PDAC). The lack of accurate predictors of high-risk disease leads both to unnecessary operations for indolent neoplasms as well as missed diagnoses of PDAC. Digital spatial RNA profiling (DSP-RNA) provides an opportunity to define and associate transcriptomic states with cancer risk.

Results: Our analysis uncovered three distinct epithelial transcriptomic states - "normal-like" (cNL), "low-risk" (cLR), and "high-risk" (cHR) - which were significantly associated with pathologic grade. Furthermore, the three states were significantly correlated with the exocrine, classical, and basal-like molecular subtypes described in PDAC. Specifically, exocrine function diminished in cHR, classical activation distinguished neoplasia (cLR and cHR) from cNL, and basal-like genes were specifically upregulated in cHR. Intriguingly, markers of cHR were detected in NL and LGD regions from specimens with PDAC but not low-grade IPMN.

Conclusions: DSP-RNA of IPMN revealed low-risk (indolent) and high-risk (malignant) expression programs that correlated with the activity of exocrine and basal-like PDAC signatures, respectively, and distinguished pathologically low-grade from malignant specimens. These findings contextualize IPMN pathogenesis and have the potential to improve risk stratification.

sample_organism:
Homo sapiens
blank sample

platform_organism:
Homo sapiens
blank sample

overall_design: Whole-transcriptome DSP-RNA profiling of 10 patients with IPMN was conducted using NanoString GeoMx. Epithelial (PanCK+) areas of interest (AOIs) were annotated as normal duct (NL), low-grade dysplasia (LGD), high-grade dysplasia (HGD), or invasive carcinoma (INV). Unique molecular identifier (UMI) encoded gene expression count data was generated by Illumina sequencing. The processed count data was then analyzed using R/Bioconductor.

first: Matthew

middle: K

last: Iyer

first: Ashley

last: Fletcher

first: Chanjuan

last: Shi


title: Targeting ferroptosis resistance re-sensitizes metastatic HR+ HER2- breast cancer to palbociclib-hormone therapy

city: Liège

first: Nor Eddine

last: Sounni

address: avenue de l'Hôpital 11

department: Medical oncology

country: Belgium

institute: Liège University (ULG - ULiège)

summary: HR+HER2- breast cancers resistant to palbociclib-hormone therapy displayed increased lipid uptake and expression of stress response proteins (GPX4, PSMA7), by adding ferroptosis inducers to palbocilib-fulvestrant combination therapy, tumor response was enhanced in three different pre-clinical models: resistant cells, xenografts and PDX.

overall_design: For the generation of resistant cells, T47D and CAMA-1 were treated with 0.3 µM of Palbociclib (P) and 30 nM of Fulvestrant (F) for 1 and 2 year(s) respectively. Resistance was assessed by proliferation assay with Incucyte. RNAseq was performed with 1) T47D parental, T47D treated with PF for 5 and 20 days and T47D resistant to PF, obtained after 1 year and 2) CAMA-1 parental cells and CAMA-1 treated with PF for 5 and 20 days.

first: Charles

last: Pottier

first: Nor

first: Laetitia

last: Ruiz


title: Single-cell joint analysis of DNA oxidative damage and transcriptome [Cortex paired-damage]

city: New York

first: Dongsheng

last: Bai

state: NY

address: 101 Avenue of the Americas

institute: New York Genome Center

summary: We used a new technology, named Paired-Damage, to perform simultaneous profiling of DNA oxidative damage and transcriptome on Hela cells and mouse cortex at single-cell level. We also performed matched damagenome, ATAC-seq and RNA-seq of Hela cells at bulk level.

overall_design: We develop Paired-Damage for simultaneous profiling the genome-wide DNA oxidative damage and transcriptome, in highly scalable single-cell manner. Paired-Damage measures all the major oxidative DNA damage including 8-oxoG, AP sites and single-strand breaks,  together with transcriptome from one same cell. To demonstrate the utility of Paired-Damage, we applied it in Hela cells treated with oxidative stress condition and mouse brain cortex cells. We also performed matched damagenome, ATAC-seq and RNA-seq of Hela cells at bulk level.

first: Zhenkun

last: Cao

first: Chenxu

last: Zhu


title: IL-23 tunes inflammatory functions of human MAIT cells

city: Paris

first: Hélène

last: Lopez-Maestre

address: 25-28 rue du Dr Roux

institute: Institut Pasteur

summary: IL-23 signaling plays a key role in the pathogenesis of chronic inflammatory and infectious diseases, yet the cellular targets and signaling pathways affected by this cytokine remain poorly understood. We show that IL-23 receptors are expressed on the large majority of human MAIT, but not of conventional T cells, suggesting that these innate-like T cells are critical mediators of IL-23 functions. In this study, we investigated the effects of IL-23 on human MAIT cell transcriptional and chromatin accessibility profiles, using RNA- and CITE-seq and ATAC-seq, respectively. Protein and transcriptional profiling at the population and single cell level demonstrates that stimulation with IL-23 or the structurally related cytokine IL-12 drives distinct functional profiles, revealing a high level of plasticity of MAIT cells. IL-23, in particular, affects key molecules and pathways related to autoimmunity and cytotoxic functions. Integrated analysis of transcriptomic and chromatin accessibility shows that AP-1 transcription factors constitute a key regulatory node of the IL-23 pathway in MAIT cells.

overall_design: PBMC were isolated from healthy donors. MAIT cells were activated for 6 days in PBMC cultures using MR1 tetramers loaded with 5-OP-RU (MR1/5-OP-RU), in the absence or presence of IL-23, or IL-12. After 6 days, MAIT cells were sorted and we performed RNA-seq, ATAC-seq, and CITE-seq. To identify the effects of IL-23 or IL-12 on MAIT cells, we compared cells activated in the presence of IL-23 or IL-12 with those stimulated with MR1/5-OP-RU only.

last: Camard

first: Tharshana

last: Stephen

first: Hanane

last: Yahia-Cherbal

first: Vincent

last: Guillemot

first: Sébastien

last: Mella

first: Victoire

last: Baillet

first: Daniele

last: Capocefalo

first: Laura

last: Cantini

first: Claire

last: Leloup

first: Julie

last: Marsande

first: Juan

last: Sienes Bailo

first: Ambre

last: Dangien

first: Natalia

last: Pietrosemoli

first: Milena

last: Hasan

first: Huimeng

last: Wang

first: Sidonia

last: Eckle

first: Anne

last: Fourie

first: Carrie

last: Greving

first: Barbara

last: Shaikh

first: Raphaelle

last: Parker

first: Daniel

last: Cua

first: Elisabetta

last: Bianchi

first: Lars

last: Rogge


title: Single-cell joint analysis of DNA oxidative damage and transcriptome [bulk damage]

first: Dongsheng

last: Bai

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

first: Zhenkun

last: Cao

first: Chenxu

last: Zhu


title: Identification of Bronchial Epithelial Genes Associated with Type-2 Eosinophilic Inflammation in Asthma

first: Kimberly

last: Dill-McFarland

state: WA

address: 750 Republican St

department: Allergy and Infectious Diseases

summary: Airway inflammation has a critical role in asthma pathogenesis and pathophysiology. Yet, the molecular pathways contributing to airway inflammation are not fully known, particularly Type-2 (T2) inflammation characterized by both eosinophilia and FeNO levels. Here, we seek to identify genes whose level of expression in epithelial brushing samples is associated with both bronchoalveolar lavage (BAL) eosinophilia and generation of FeNO. We used a segmental allergen bronchoprovocation (SBP-Ag) procedure in asthma subjects, and RNA-sequencing (RNA-seq) analyses of BAL cells and brushing samples before and 48 h after SBP-Ag. Allergen bronchoprovocation increased FeNO levels which correlated with eosinophilia but not neutrophilia. Thirteen genes were identified in brushing samples, whose expression changed in response to SBP-Ag and correlated with both airway eosinophilia and FeNO levels after SBP-Ag. Among these 13 genes, the epithelial cell product, CDH26 was a candidate to contribute to the amplification of T2 inflammation as reflected by eosinophilia and FeNO, and causal mediation analyses with pro-T2 and pro-eosinophilic cytokine mediators in BAL fluids. Among the genes associated with reduced eosinophila and FeNO, HEY2 is known to enhance cell proliferation, migration, invasion, and EMT, as well as to reduce apoptosis. This unbiased RNA-seq analysis in subjects with allergic asthma has revealed several epithelial cell genes that may be critical for the development or augmentation of T2 inflammation in asthma, particularly CDH26.

overall_design: Segmental bronchial provocation with allergen (SBP-Ag) was used to provoke airway inflammation and retrieve airway cells to characterize cellular responses by RNA-sequencing to determine gene-expression profiles. This sample set includes bronchoalveolar brushings (BE). Differential gene-expression was assessed by linear modeling of post vs pre SBP-Ag gene expression as well as post SBP-Ag expression associated with FeNO, FEV1, eosinophils, and neutrophils.

first: Stephane

last: Esnault

first: Matthew

middle: C

last: Altman

first: Melissa

last: Rosenkranz

first: Nizar

last: Jarjour

first: William

last: Busse


title: Transcriptional acclimation to warming temperatures in the Australian alpine herb Wahlenbergia ceracea

city: Poznan

first: Rocco

middle: Fabrizio

last: Notarnicola

state: Wielkopolskie

address: ul, Uniwersytetu Poznańskiego 6

department: Evolutionary Biology

institute: Adam Mickiewicz University

summary: Understanding the molecular basis of plant heat tolerance helps to predict the consequences of climate warming on plant performance, particularly in vulnerable environments. Our current understanding comes primarily from studies in Arabidopsis thaliana (Arabidopsis) and select crops exposed to short and intense periods of heat stress. In this study we sought to characterise the transcriptomic responses of Wahlenbergia ceracea (W. ceracea), an Australian alpine herb, to growth under sustained moderate warming. We compared responses of pre-defined tolerant and sensitive lines, based on measures of photosynthetic thermostability, to growth under cool (24/15 ºC day/night) or warm (30/20 ºC day/night) temperatures to identify the pathways involved in heat tolerance and acclimation. Under warmer growth temperatures, W. ceracea up-regulated genes involved in RNA metabolism, while down-regulating those involved in photosynthesis and pigment metabolism. In tolerant lines, genes related to photosystem II and light-dependent photosynthetic reactions were more strongly down-regulated. This suggests that the regulation of electron transport and its components may be involved in thermal acclimation. Our results also highlight the importance of hormonal gene networks, particularly those responsive to ethylene, during longer-term moderate warming. In conclusion, our results point to post-transcriptional processes and the stabilisation of the electron transport chain as candidate mechanisms for thermal acclimation in heat-tolerant W. ceracea. The study also revealed many temperature-responsive genes with no detectable ortholog in Arabidopsis, whose characterization may enhance our understanding of physiological acclimation and have relevance for the biotechnological improvement of threatened species and crops.

overall_design:
To investigate transcriptional responses potentially involved in physiological acclimation to warming, we grew plants ofWahlenbergia ceraceaunder either cool (current) or warmer (future) temperatures
We then performed differential gene expression analysis using RNA-seq data from leaf tissues harvested after two weeks of acclimation under experimental conditions

middle: F

first: Piter

last: Arnold

first: Zhi-Ping

last: Feng

first: Jemimah

last: Hamilton

first: Ashley

last: Jones

first: Stella

last: Loke

first: Benjamin

last: Schwessinger

first: Diep

last: Ganguly

first: Adrienne

last: Nicotra


title: The GIP receptor activates futile calcium cycling in white adipose tissue to increase energy expenditure and drive weight loss in mice

first: Christine

last: Kusminski

address: 5323 Harry Hines Boulevard

department: Internal Medicine, Touchstone Diabetes Center

institute: The University of Texas Southwestern Medical Center

summary: Obesity is a chronic disease that contributes to the development of insulin resistance, type 2 diabetes (T2D), and cardiovascular risk. GIP receptor (GIPR) and GLP-1 receptor (GLP-1R) co-agonism provide an improved therapeutic profile in individuals with T2D and obesity when compared with selective GLP-1R agonism. While the metabolic benefits of GLP-1R agonism are established, whether GIPR activation impacts weight loss through peripheral mechanisms is yet to be fully defined. Here, we generated a mouse model of GIPR induction exclusively in the adipocyte. We show that GIPR induction in the fat cell protects mice from diet-induced obesity and triggers profound weight loss (~35%) in an obese setting. Adipose GIPR further increases lipid oxidation, thermogenesis and energy expenditure. Mechanistically, we demonstrate that GIPR induction activates SERCA-mediated futile calcium cycling in the adipocyte. GIPR activation further triggers a metabolic memory effect, which maintains weight loss after the transgene has been switched off, highlighting a unique aspect in adipocyte biology. Collectively, we present a mechanism of peripheral GIPR action in adipose tissue, which exerts beneficial metabolic effects on body weight and energy balance.

overall_design: To examine the in vivo effects of inducing the GIPR in white fat on weight loss and energy balance, we performed RNA sequencing on subcutaneous adipose tissue from wild-type (WT) control mice and inducible adipose tissue-specific TRE-GIPR-Adip (GA) mice. This was to evaluate transcriptional changes in adipose tissue following feeding of Doxycycline-diet containing high fat-diet (Dox-HFD) to all mice for 2 weeks (i.e., GIPR transgene induction in adipose tissue).

first: Xinxin

last: Yu


title: High Shear Stress Reduces ERG Causing Endothelial-Mesenchymal Transition and Pulmonary Arterial Hypertension

first: Jan-Renier

last: Moonen

address: 269 Campus Drive, CCSR 1220

summary: Computational modeling indicated that a pathological level of high shear stress (HSS, 100 dyn/cm2) is generated in distal pulmonary arteries (PA) (100-500 um) in a congenital heart defect with increased PA blood flow causing PA hypertension (PAH), and in idiopathic PAH with occlusive vascular remodeling.  The response of human PA endothelial cells (EC) to HSS compared to physiologic laminar shear stress (LSS, 15 dyn/cm2), was therefore assessed.  Endothelial-mesenchymal transition (EndMT), a feature of PAH not previously attributed to HSS was observed. HSS did not alter induction of the transcription Krüppel-like factors (KLF) 2/4, but H3K27ac peaks containing motifs for an ETS-family transcription factor (ERG) were reduced, as was the interaction between ERG and KLF2/4 and ERG expression.  In PAEC under LSS, reducing ERG by siRNA caused EndMT related to decreased bone morphogenetic protein receptor 2 (BMPR2), cadherin 5 (CDH5) and platelet and endothelial cell adhesion molecule 1 (PECAM1), and increased Snail/Slug (SNAI1/2) and smooth muscle alpha α-2 actin (ACTA2).  In PAEC under HSS, transfection of ERG prevented EndMT.  We induced HSS in mice by an aorto-caval shunt that causes a progressive increase in PAH over eight weeks and used an adeno-associated viral vector (AAV2-ESGHGYF) to replenish ERG selectively in PAEC.  Elevated PA pressure and resistance, EndMT and vascular remodeling assessed by muscularization of peripheral arteries were markedly reduced by ERG delivery in the aorto-caval shunt mice. Thus, agents that restore ERG in the pulmonary vasculature will be of therapeutic benefit in overcoming the adverse effect of HSS on progressive PAH.

overall_design: Pulmonary artery endothelial cells were pre-conditioned by exposure to 15 dyn/cm2 of unidirectional uniform LSS for 24 h. Cells were then exposed to an additional 24 h of LSS, or 24 h of 100 dyn/cm2 unidirectional uniform HSS. Shear stress was generated using the Ibidi Perfusion System (Ibidi). Static (ST) controls were performed simultaneously with the shear stress experiments, cultured on 0.4 mm chamber slides using the same EC media. CUT&RUN was performed using the CUTANA CUT&RUN kit (#14-1048, EpiCypher, USA) following manufacturer’s instructions.

first: Tsutomu

last: Shinohara

first: Ramesh

last: Nair

first: Katherine

last: Guo

first: Jesse

last: Engreitz

first: Marlene

last: Rabinovitch


title: Spatio-temporal X-linked gene reactivation in the mouse germline reveals site-specific retention of epigenetic silencing

first: Laurène

last: Syx

address: 11 rue Pierre et Marie Curie

department: Centre de recherche

institute: Institut Curie

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Ectopic expression of DNMT3L in human trophoblast stem cells restores features of the placental methylome [ChIP-seq]

city: Cambridge

first: Sarah

last: Inglesfield

address: Babraham Research Campus

institute: The Babraham Institute

summary: The placental DNA methylation landscape is unique, with widespread partially methylated domains (PMDs) and hundreds of placental-specific imprinted domains. Furthermore, the placental ‘methylome’ has been the focus of extensive study for links with pregnancy complications. Human trophoblast stem cells (hTSCs) offer exciting potential for functional epigenetic studies; however, whether the hTSC epigenome recapitulates primary cytotrophoblast, remains poorly described. In this study, we demonstrate that hTSCs exhibit an atypical methylome, with DNA methylation present over transcribed gene bodies but a complete loss of placental PMDs. Using single-cell RNA-seq from human embryogenesis, hTSCs display a notable absence of DNMT3L expression. Ectopic expression of DNMT3L in hTSCs restored placental PMDs. DNMT3L-expressing hTSCs showed comparable stemness but failed to syncytialise in organoid culture, associated with hypermethylation of STB transcription factor motifs. These findings reveal that DNMT3L is essential in establishing the human placental methylome and that DNMT3L downregulation is necessary for successful trophoblast differentiation.

overall_design:
To understand determinants of the DNA methylation landscape in hTSCs, we cultured blastocyst- (BTS11) and 1st trimester cytotrophoblast- (CT29) derived hTSCs in 2D conditions (Okae et al., 2018), 2D in organoid media, 3D as organoids (Sheridan et al., 2021), and as organoids cultured in hypoxic conditions. We assayed histone modifications using ChIP_seq to evalue the localisation of DNA methylation in hTSCs.

Please note that the following published datasets were re-analyzed as replicates (as detailed in the re-analyzed_sample_details.txt).

The accessions of the public datasets used in the current study:

Sample GSM accns Series SRX accns Biosample accns
================================================
GSM3633664 GSE127288 SRX5443225 SAMN11029824
GSM3633665 GSE127288 SRX5443226 SAMN11029822
GSM3633672 GSE127288 SRX5443233 SAMN11029813
GSM3633673 GSE127288 SRX5443234 SAMN11029848
GSM3633676 GSE127288 SRX5443237 SAMN11029842
GSM3633677 GSE127288 SRX5443238 SAMN11029846
GSM5862947 GSE196221 SRX14060225 SAMN25690350
GSM5862950 GSE196221 SRX14060228 SAMN25690347
GSM5862951 GSE196221 SRX14060229 SAMN25690346
GSM6043305 GSE200760 SRX14845721 SAMN27567865
GSM6043306 GSE200760 SRX14845722 SAMN27567864
GSM6043307 GSE200760 SRX14845723 SAMN27567863
GSM6043317 GSE200761 SRX14845739 SAMN27567878
GSM6043318 GSE200761 SRX14845740 SAMN27567877
GSM1436879 GSE59434 SRX652746 SAMN02911906
GSM1436880 GSE59434 SRX652747 SAMN02911907
GSM1436881 GSE59434 SRX652748 SAMN02911908
GSM1436882 GSE59434 SRX652749 SAMN02911909
GSM2232855 GSE84382 SRX1945295 SAMN05389843
GSM2232856 GSE84382 SRX1945296 SAMN05389844
GSM2232857 GSE84382 SRX1945297 SAMN05389845
GSM2232858 GSE84382 SRX1945298 SAMN05389846
GSM2232859 GSE84382 SRX1945299 SAMN05389847
GSM2232860 GSE84382 SRX1945300 SAMN05389848

first: Courtney

last: Hanna

first: Georgia

last: Lea

first: Vicente

last: Perez-Garcia


title: The m6A Demethylase ALKBH5 Maintains Stemness and Therapeutic Resistance of Glioblastoma Stem Cells [Polysome-seq]

city: Xiamen

first: Tianzhi

last: Huang

address: No. 4221, Xiang'an South Road, Xiang'an District

department: State Key Laboratory of Cellular Stress Biology

institute: Xiamen University

summary: One prevalent modification, N6-methyladenosine (m6A), is a dynamic and reversible process regulated by methyltransferases (the "writers"), demethylases (the "erasers"), and m6A-binding proteins (the "readers"). Its abnormal changes are closely associated with the development and progression of human cancers. m6A demethylases, such as FTO and ALKBH5, eliminate the m6A modification from RNA, thus regulating the overall levels of this modification. Here, we show that the m6A demethylase ALKBH5 is highly expressed after ionizing radiation treatment. We conducted polysome profiling alongside RNA sequencing in both control and ALKBH5-KO GSC 83 cells and identified genes with differential translational efficiency due to ALKBH5 KO. We further elucidated that genes, whose translational efficiencies (TEs) were down-regulated after ALKBH5 knockout, were enriched in stemness properties and DNA repair. Overall, our findings imply that ALKBH5 facilitates the selective translation of mRNAs encoding pathways related to stemness and resistance to therapeutic interventions.

overall_design: Polysome-seq profiling of wildtype and ALKBH5-KO GSC 83 cells

first: TianZhi

first: Xiaobing

last: Jiang

first: Xuan

last: Wang


title: Spatio-temporal X-linked gene reactivation in the mouse germline reveals site-specific retention of epigenetic silencing [RNA-seq]

summary: Random X-chromosome inactivation (XCI) is a hallmark of female mammalian somatic cells. This epigenetic mechanism, mediated by the long non-coding RNA Xist, occurs in the epiblast and is stably maintained to ensure proper dosage compensation of X-linked genes during life. However, this silencing is lost during primordial germ cell (PGC) development. Using a combination of single-cell allele-specific RNA sequencing and low-input chromatin profiling on in vivo developing mouse PGC, we provide unprecedented detailed maps of gene reactivation. We demonstrate that PGC still carry a fully silent X chromosome at embryonic day (E) 9.5, despite the loss of Xist expression. X-linked genes are then gradually reactivated outside the Xist first-bound regions. At E12.5, a significant part of the inactive X chromosome (Xi) still resists reactivation, carrying an epigenetic memory of its silencing. Late-reactivated genes are enriched in repressive chromatin marks, including DNA methylation and H3K27me3 marks. Our results define the timing of reactivation of the silent X chromosome a key event in female PGC reprogramming with direct implications for reproduction.

overall_design:
After dissection of embryos at the location of the PGC, according to embryonic stage, and sexing of the embryos, samples were resuspended in 200 µL of 0.25 % trypsin and incubated at 37 °C for 3 min. Trypsin was inactivated with serum and a single-cell solution was obtained by vigorous up-and-down.
For scRNA-seq experiments from E10.5, cells were manually picked based on their GFP and size and washed in PBS-acetylated BSA. For the earlier developmental stages, cells were collected by FACS and processed quickly for cDNA amplification.
Poly(A)+ mRNA extracted from each single cell was reverse-transcribed from 3’-UTRs and amplified according to Tang et al, 2010 and Borensztein et al, 2018.

first: Clara

last: Roidor

first: Nicolas

last: Servant

first: Maud

last: Borensztein


title: Genome and transcriptome analysis of the lignite-degrading Trichoderma cf. simile WF8 strain highlights potential degradation mechanisms

status: Public on Jan 05 2025

city: Nanjing

first: Paul

last: Daly

state: Jiangsu

address: 50 Zhongling St

institute: Jiangsu Academy of Agricultural Sciences

summary:
The biodegradation of lignite (brown coal) by microorganisms has the potential for bioremediation of contaminated mining sites and to generate alternative ways to valorize lignite, such as by producing humic acids or building block chemicals. Previously, a lignite-degrading strain of Trichoderma was isolated, but the genomic and transcriptomic basis of its lignite-degrading ability remained unknown. Here we report that the sequenced genome of the T. cf. simile WF8 strain encoded for enzymes with roles in the degradation of lignite, and potentially tolerance to lignite-breakdown products. There was only a small number of annotated unique genes in the T. cf. simile WF8 genome compared to other fungi, and likely the expression of gene families shared with other fungi is a key factor in lignite biosolubilization by T. cf. simile. The transcriptomes were analyzed of T. cf. simile cultured at two time-points with the lignite-breakdown model compounds 4-phenoxybenzoic acid (which was growth inhibitory), and phenetole and 9-10-dibutoxyanthracene (neither of which inhibited growth), and showed ~20% of genes up-regulated by one or more of these compounds. The analysis highlights candidates for characterization and engineering enzyme over-expressing T. cf. simile strains with potentially improved degradation capacity, e.g., laccases and peroxidases, or tolerance and catabolism of breakdown products, e.g., cytochrome P450s, and ring cleavage dioxygenases.

Published in International Biodeterioration & Biodegradation (https://doi.org/10.1016/j.ibiod.2025.105997)

overall_design: Transcriptome of mycelia from shake-flask cultures from two time points (1 day or 3 day) with one of three lignite model compounds (4-phenoxybenzoic acid, phenetole or 9-10-dibutoxyanthracene) or a control without a lignite model compound.

first: Jinghua

last: Yao

first: Yajuan

last: Chen

first: Deyu

last: Zhuo

first: Siqiao

first: Baichao

last: Xu

first: Congwei

last: Yan

first: Wanrong

last: Li

first: Hui

last: Feng

first: Sheng

last: Deng

first: Feng

last: Cai

first: Andrei

last: Steindorff

first: Irina

last: Druzhinina

first: Lei

last: Xiao

first: Lihui

last: Wei


title: Ly6ChiCD62LhiLy6GhiCx3CR1low Circulating Monocyte subsets are altered in a Murine Diabetic Gastroparesis Model

city: Rochester

first: Madhusudan

last: Grover

state: MN

address: 200 First St SW

institute: Mayo Clinic

summary: Circulating monocytes (Mo) are precursors to a subset of gastric resident-muscularis-macrophages (MMs). Changes in MMs are associated with delayed gastric emptying (DGE) in diabetic gastroparesis. Investigating the dynamics of Mo in an animal model of DGE using CyTOF and computational approaches, we show a high heterogeneity within the Mo-population. In DGE mice, via unbiased clustering we identified two reduced Mo clusters which exhibit migratory phenotype (Ly6ChiCD62LhiLy6GhiCD45RhiCCR2hi-intMERTEKhi-intCx3CR1low) resembling classical-Mo (CMo-like). All markers enriched in these clusters are known to regulate cell differentiation, proliferation, adhesion, and migration. Trajectory inference analysis predicted these Mo as precursors to subsequent Mo-lineages. In gastric muscle tissue, we demonstrated an increase in the gene expression level of chemokine receptor Ccr2, suggesting possibly increased trafficking of classical monocytes. These findings contribute to better understanding of the heterogenicity of the Mo-population and establish a link between two CMo-like clusters and the development of DGE phenotype.

overall_design: In this study, we determined the state of Mo in the setting of diabetes with and without DGE in a STZ-induced-diabetes mouse model using cytometry by time-of-flight (CyTOF) and computational analysis.

first: Shefaa

last: AlAsfoor

first: Erik

last: Jessen

first: Suraj

last: Pullapantula

first: Jennifer

last: Voisin

first: Linda

last: Hsi

first: Kevin

middle: D

last: Pavelko

first: Samera

last: Farwana

first: Sihan

last: Ji

first: Michael

last: Strausbauch

first: Gianluca

last: Cipriani

first: Lai

first: David

last: Linden

first: Ruixue

last: Hou

first: Richard

last: Myers

first: Yogesh

last: Bhattarai

first: Jill

last: Wykosky

first: Alan

last: Burns

first: Gianrico

last: Farrugia

first: Surendra

last: Dasari


title: The effects of sorafenib treatment on the regulatory T cell (Treg) population

city: Hangzhou

first: Shuowang

last: Wang

address: Yuhang Tang Street No.866

institute: Zhejiang University

summary: To investigate the impact of sorafenib treatment on gene expression in Treg cells, we performed RNA sequencing on Treg cells treated with sorafenib and on control Treg cells that were not treated with sorafenib. Specifically, our study focused on the expression of immune-suppressive related genes, and we observed an upregulation in the expression of Tgf-β1 (Transforming Growth Factor-β1), Prf1 (Perforin 1), Gzma (Granzyme A), and Il10 (Interleukin 10).

overall_design: Comparative gene expression profiling analysis of RNA-seq data for sorafenib treated Treg cell and non-sorafenib treated cell.


title: The effects of sorafenib treatment on the Hep1-6 tumor tissue

summary: This study aimed to investigate the transcriptomic changes in subcutaneous Hepa1-6 tumor tissues of mice following intraperitoneal administration of sorafenib, a tyrosine kinase inhibitor, versus a control solvent. Mice were injected with either sorafenib or the control solvent four times, according to the experimental design. After the treatment period, tumor samples were collected and subjected to RNA sequencing (RNA-seq) to analyze the gene expression profiles. The results from this RNA-seq analysis provide insights into the molecular mechanisms underlying the response of Hepa1-6 tumors to sorafenib treatment, potentially identifying novel therapeutic targets and biomarkers for hepatocellular carcinoma.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for sorafenib treated  and non-sorafenib treated Hep1-6 tissue.


title: Single-cell joint analysis of DNA oxidative damage and transcriptome [Hela RNA]

first: Dongsheng

last: Bai

first: Zhenkun

last: Cao

first: Chenxu

last: Zhu


title: Single-cell joint analysis of DNA oxidative damage and transcriptome [Hela paired damage]


title: IMMUNE-RESPONSIVE GENE 1 REGULATES ATHEROSCLEROTIC PLAQUE INFLAMMATION IIII

status: Public on Jan 03 2025

last: Gildea

state: New York

address: 550 1st avenue

institute: NYU Langone Health

summary: Atherosclerosis is the major underlying cause of cardiovascular disease (CVD) mortality and is fueled by a failure to resolve inflammation within the vessel wall1. The mechanisms underpinning sustained inflammation within this microenvironment are not fully elucidated. A more comprehensive understanding of the immunoregulatory pathways modulating inflammation in this context holds the potential to inform the development of more specific therapies. Immune responsive gene 1 (Irg1) is an enzyme that diverts cis-aconitate from the TCA cycle and produces the metabolite itaconate, which elicits immunological tolerance in the context of microbial infection2–4. We hypothesized that the IRG1-itaconate axis restricts inflammation in the atherosclerotic plaque.

overall_design: Formation of atherslerotic plaque was induced in mice by placing them on Western Diet. Aorta's were removed and single-cell suspensions of the aortic arch were made and CD45+ immune cells were isolated by staining (anti-CD45 PerCp-Cy5.5, BioLegend) and flow sorting viable (fixable viability dye e780, eBioscience) single cells on a Facs Aria II cytometer (BD) equipped with a 100 µm nozzle. Next scRNA seq was performed and analyzed.

first: Fazli

last: Bozal

first: Yannick

last: Cyr

first: Jose

last: Barcia Duran

first: Kathryn

last: Moore


title: Loss of aryl hydrocarbon receptor reduces pancreatic tumor growth by increasing immune cell infiltration

city: Oslo

first: Jason

last: Matthews

address: Sognsvannsveien 9

department: Nutrition

country: Norway

institute: University of Oslo

summary: Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease which despite large research efforts remains poorly understood. Many studies provide evidence that the aryl hydrocarbon receptor (AHR) plays a critical role in the pathogenesis of several cancers and represents new anti-cancer therapeutic target. The role of AHR in PDAC is, however, unclear since AHR has been reported to exhibit both pro- and anti-tumor activities. In this study we evaluated the role of AHR in murine PDAC cells in vitro and in vivo. Loss of AHR did not affect cell proliferation compared with wildtype (WT). Consistent with these findings, no differences in tumor development between WT and AhrKO cells were observed in immunocompromised mice. However, tumors from AhrKO cells grew more slowly than tumors from WT cells in immune competent mice. RNA sequencing of the tumors resulted in 1009 genes upregulated and 477 genes downregulated in AhrKO tumors compared with WT tumors. Pathway analysis identified immunoregulatory interactions, interferon signaling, interleukin signaling and PD1 signaling among the top upregulated pathways. Increased infiltration of CD45+ cells and higher numbers of CD8+ T-cells and F4/80+ cells were observed in AhrKO tumors. Analysis of the F4/80+ cell populations revealed a lower percentage of anti-inflammatory (M2) macrophages in AhrKO tumors. Ahr deficiency in macrophages (LysMCre) or lymphocytes (RorcCre) did not alter tumor development of WT cells compared with WT C57BL/6 mice. AhrKO tumors in RorcCre mice, but not in LysMCre mice had significantly lower tumor weights normalized to body weights compared with AhrKO tumors in WT mice. These findings show that tumors derived from AhrKO pancreatic cancer cells exhibit a proinflammatory gene responses that contribute to increased immune cell infiltration and reduced tumor growth. They also suggesting that loss of AHR in lymphocytes together Ahr loss in cancer cells further reduces tumor growth.

overall_design: CR705 cells were plated at a density of 1 × 105 cells/well in a 6-well plate and treated with test compounds the following day. After 24 h incubation, cells were lysed and harvested using 10 mM Tris-Cl buffer containing 1 mM EDTA and 1% SDS, with pH of 8.0, and sonicated 2 × 30 seconds on/off with Bioruptor (Diagenode, Denville, NJ, USA). Pierce™ BCA Protein Assay Kit (Thermo Fischer Scientific, Waltham, MA, USA) was used to determine protein concentration.  CR705 mice AHRKO cells were compared relative to WT cells

first: Vinicius

last: Kannen

first: Ninni

last: Olafsen

first: Siddhartha

last: Das

first: Paolo

last: Giulana

first: Fauzia

last: Izzati

first: Hani

last: Choksi

first: Samaneh

last: Åhrling

first: Paola

last: Cappello

first: Indrek

last: Teino

first: Toivo

last: Maimets

first: Kristaps

last: Jaudzems

first: Antanas

last: Gulbinas

first: Zilvinas

last: Dambrauskas

first: Landon

last: Edgar

first: Denis

last: Grant


title: effect of RNA uptake on gene expressions in human NK and CD8-positive T cells

city: Matsumoto

first: Sachie

last: Hiratsuka

address: 3-1-1 Asahi

department: Medicine

institute: Shinshu University

summary: In the blood, non-vesicular extracellular RNA functions as a cell-cell communication molecule. We have identified that IL1b-mRNA fragment binds ZC3H12D protein on the cell surface. This protein expresses on NK and T cells.  When extracellular RNA binds ZC3H12D on the cell surface, the RNA-ZC3H12D complex goes inside the cell to modulate cell character, and as a result, the RNA stimulated-immune cell became anti-tumor rather than pro-tumor. To decipher the molecular mechanisms to govern the extracellular RNA-mediated signaling, we carried out NGS analysis to find out the difference in gene expression profiles after external RNA stimulation.

overall_design: Human PBMC was purchased from Roche. Thawed PBMC was maintained in HPLM (Thermo) medium supplemented with 10% FCS. The following day, FITC labeled IL1b-mRNA  (10ng/mL) was applied in the media, and 0.5h or 6h later cells were FACS-sorted to isolate NK cells (CD56+CD3-) and CD8+T cells. Isolated cells were immediately placed into Trizol (Thermo) reagent to obtain total RNA.

first: Takeshi

last: Tomita

first: Masayoshi

last: Kato

first: Hikaru

last: Hayashi

first: Sayaka

last: Seki


title: Effects of nitrate on Vibrio cholerae transcription (RNASeq)

city: boston

first: jamaurie

last: james

state: ma

address: 300 longwood ave

institute: Boston Childrens Hospital

summary: Within this study, we aimed to investigate how enviornmental nitrate effects the  transcription of the Vibrio cholerae genome. Here we performed RNASeq using static cultures of WT, dCrp, dHapR, & dhapR/Crp.

overall_design: To evaluate the transcriptional regulation of Vibrio cholerae, we performed RNASeq on mutant strains of  V. cholerae O1 El Tor strain C6706. The individual samples were wild type, and those that contained in frame deletions of cyclic AMP receptor protein(CRP), hapR, and a double mutant CRP/hapR. In triplicate, we performed the experiment statically in two conditions at the same time point: those grown in LB Media, and those grown in LB media supplemented with 5mM Sodium Nitrate. Differential gene expression analysis was perfomed to obtain more data.

first: Jamaurie

last: James

first: Renato

last: Santos

first: Paula

last: Watnick


title: Single-cell joint analysis of DNA oxidative damage and transcriptome [ATAC-seq]

first: Dongsheng

last: Bai

summary: We used a new technology, named Paired-Damage, to perform simultaneous profiling of DNA oxidative damage and transcriptome on Hela cells and mouse cortex at single-cell level. We also performed matched damagenome, ATAC-seq and RNA-seq of Hela cells at bulk level.

overall_design: We develop Paired-Damage for simultaneous profiling the genome-wide DNA oxidative damage and transcriptome, in highly scalable single-cell manner. Paired-Damage measures all the major oxidative DNA damage including 8-oxoG, AP sites and single-strand breaks,  together with transcriptome from one same cell. To demonstrate the utility of Paired-Damage, we applied it in Hela cells treated with oxidative stress condition and mouse brain cortex cells. We also performed matched damagenome, ATAC-seq and RNA-seq of Hela cells at bulk level.

first: Zhenkun

last: Cao

first: Chenxu

last: Zhu


title: The Guinea Pig Serves as an Alternative Model to Study Human Preimplantation Development

city: Stockholm

first: Cheng

last: Zhao

state: Stockholm

address: H9 Klinisk vetenskap,171 77 Stockholm

department: Clinical Science

institute: Karolinska institute

summary: Preimplantation development is a pivotal phase in human embryogenesis, establishing fundamental lineages and being crucial for overall health. Ethical constraints with human embryos necessitate a model organism, and the guinea pig, with its physiological similarities to humans, emerges as a valuable alternative. This rodent mirrors human preimplantation timing, placental features, and fetal development. Importantly, it serves as a model for studying long-term consequences of prenatal insults. Comparative studies across species, including guinea pigs, are essential for unraveling pluripotency mechanisms, offering insights into conserved and divergent aspects and understanding evolutionary adaptations. Addressing the lack of guinea pig embryonic transcriptomic data, this research employs single-cell RNA sequencing, immunofluorescence, and epigenetic analyses to explore gene expression, metabolic requirements, and X-chromosome inactivation dynamics. The study advances our understanding of early embryogenesis, emphasizing the guinea pig's relevance as a model for developmental and pluripotency research.

overall_design: To obtain a comprehensive overview of the spatial-temporal gene expression and lineage dynamics across guinea pig preimplantation development, we sequenced the transcriptomes of individual blastomeres isolated from E3-E6 embryos.

first: Jesica

last: Canizo

first: Sophie

last: Petropoulos


title: An epigenetic timer of gestation length in mice [CUT&RUN]

city: San Francisco

first: Adrian

last: Erlebacher

address: 513 Parnasssus

institute: UCSF

summary: Current efforts investigating parturition timing mechanisms have focused on the proximal triggers of labor onset generated in late gestation. By studying the delayed parturition phenotype of mice with uterine fibroblast deficiencies in the histone H3K37me3 demethylase KDM6B, we provide evidence that parturition timing is regulated by events that take place in early pregnancy. Immediately after copulation, uterine fibroblasts engage in a locus-specific, epigenetic program that abruptly adjusts H3K27me3 levels across their genome. In the absence of KDM6B, many of the adjusted loci over-accumulate H3K27me3. This over-accumulation leads to nearby genes being misexpressed in mid-to-late gestation, a delayed effect partly attributable to a second locus-specific but KDM6B-independent process initiated within uterine fibroblasts soon after implantation. This second process employs progressive H3K27me3 loss to temporally structure post-midgestational patterns of gene induction. Further dissection of the ways uterine programming controls parturition timing may have relevance to human pregnancy complications such as preterm labor.

overall_design: H3K27me3 CUT&RUN was performed on myometrium/endometrium fibroblasts from control and Pgr-cre Kdm6b floxed mice on GD0.5, GD5.5, GD8.5, GD14.5, and GD18.5. Additionally, H3K27me3 CUT&RUN was performed on myometrium/endometrium fibroblasts from B6CBAF1/J from GD0.5, GD5.5 after natural mating, or GD5.5 after mating with a vasectomized male. Data also includes H3K27me3 CUT&RUN on B6CBAF1/J non-pregnant mice treated with progesterone for 3 days prior to sacrifice.

first: McIntyre

last: Tara

first: Erlebacher

last: Adrian


title: mRNA-seq analysis of mouse cytotoxic T lymphocytes (CTLs) over-expressing Akt isoforms

city: Miaoli County

first: Li-Rung

last: Huang

address: No.35, Keyan rd. Zhunan town

country: Taiwan

institute: National Health Research Institutes

summary: PI3K/Akt pathway plays a central role in T-cell activation and is impaired in Ag-specific CTLs during chronic viral infections. In this study, we genetically engineered HBV-specific CTLs and adoptively transferred these Akt-expressing viral Ag-specific CTLs into HBV-infected mice and monitored the virological and immunological responses. In the meantime, we also analyzed the mRNA profiles of control-CTLs and CTLs-overexpressing Akt1 and Akt2, respectively before adoptive transfer and after in vitro restimulation. We expect to elucidate the potential mechanisms about how the Akt signaling in CTLs prevents T-cell exhaustion and enhance T-cell expansion in the liver during persistent HBV infection from this mRNA-seq analysis.

overall_design: Examination of mRNA profiles of mouse CTLs over-expressing mouse Akt1, Akt2 and control reporter protein, respectively.

first: Su-Fang

last: Lin