title: Beckwith-Wiedemann syndrome and large offspring syndrome involve alterations in methylome, transcriptome, and chromatin configuration [WGBS]

city: Columbia

first: Rocío

middle: Melissa

last: Rivera

state: MO

address: 920 East Campus Drive

department: Division of Animal Sciences

institute: University of Missouri, Columbia

summary: Beckwith-Wiedemann Syndrome (BWS) is the most common epigenetic overgrowth syndrome, caused by epigenetic alterations on chromosome 11p15. In ~50% of patients with BWS, the imprinted region KvDMR1 (IC2) is hypomethylated. Nearly all children with BWS develop organ overgrowth and up to 28% develop cancer during childhood. The global epigenetic alterations beyond the 11p15 region in BWS are not currently known. Uncovering these alterations at the methylome, transcriptome, and chromatin architecture levels are necessary steps to improve the diagnosis and understanding of patients with BWS. Here we characterized the complete epigenetic profiles of BWS IC2 individuals together with the animal model of BWS, bovine large offspring syndrome (LOS). A novel finding of this research is the identification of two molecular subgroups of BWS IC2 individuals. Genome-wide alterations were detected for DNA methylation, transcript abundance, alternative splicing events of RNA, chromosome compartments, and topologically associating domains (TADs) in BWS and LOS, with shared alterations identified between species. Altered chromosome compartments and TADs were correlated with differentially expressed genes in BWS and LOS. Together, we highlight genes and genomic regions that have the potential to serve as targets for biomarker development to improve current molecular diagnostic methodologies for BWS.

overall_design: Whole genome bisulfite sequencing (WGBS) for skin fibroblast primary cells derived from five control and six LOS bovine day 105 fetuses.

first: Yahan

last: Li

first: Ping

last: Xiao

first: Frimpong

last: Boadu

first: Anna

last: Goldkamp

first: Snehal

last: Nirgude

first: Jianlin

last: Cheng

first: Darren

last: Hagen

first: Jennifer

middle: M

last: Kalish


title: Epigenetic profiling of prostate adenocarcinoma and neuroendocrine prostate cancer models to understand DNMT-EZH2 crosstalk. [Cut&Run]

status: Public on Nov 25 2025

city: Boston

first: Varadha Balaji

last: Venkadakrishnan

state: MA

address: 450 Brookline Ave

department: Department of Medical Oncology

country: USA

institute: Dana-Farber Cancer Institute

type: Genome binding/occupancy profiling by high throughput sequencing

summary: EZH2-DNMT crosstalk is diverse across prostate adenocarcinoma and neuroendocrine prostate cancer.

first: Venkadakrishnan

last: Varadha Balaji

first: Beltran

last: Himisha


title: Novel lentiviral constructs for the specification of rare and ectopic cell types in human colonoids

city: Charleston

first: Jorge

last: Munera

state: SC

address: 173 Ashley Avenue

institute: Medical University of South Carolina

type: Expression profiling by high throughput sequencing

summary: In the past 15 years, organoid models have revolutionized the study of the human gastrointestinal (GI) epithelium. Organoids can be derived from patient tissue and grown as strictly epithelial structures referred to as enteroids and colonoids. Despite their utility, rare cell types are often absent in enteroids and colonoids. We hypothesized that induced expression of key transcription factors (TFs) would be sufficient to specify a subset of these cell types. To test this hypothesis, we generated lentiviral constructs that allow the inducible expression of cell-type specifying TFs. We transduced human enteroids and colonoids with these constructs and examined their ability to induce differentiated cell types.

overall_design: To determine the sufficiency of TFs for specifying cell types in enteroids and colonoids, cDNAs for POU2F3 which is required for tuft cell differentiation, SPIB which is required for M-cell differentiation, and KLF4 which is required for goblet cell differentiation, were individually cloned into a doxycycline inducible lentiviral vector. A lentiviral NEUROG3 construct has been previously shown to induce enteroendocrine cells in enteroids and human pluripotent stem cell derived gastrointestinal organoids. NEUROG3 is required for enteroendocrine cell specification in the GI tract. To determine if ectopic duodenal enteroendocrine cells could be generated in colonoids, we generated a NEUROG3-T2A-NKX6-3 construct. NKX6-3 is required for generation of gastrin-producing G cells and somatostatin-producing  D cells. The resultant constructs were packaged into lentiviral particles which were used to transduce human enteroids/colonoids

first: Pritiprasanna

last: Maity

last: Qu

middle: O

last: Múnera


title: Satb2 gene dosage in cortical projection neurons impacts neuronal maturation and somatosensory processing [CUT&RUN]

city: Santa Cruz

first: Bin

last: Chen

state: CA

address: 1156 High St.

department: MCD Biology

institute: University of California Santa Cruz

summary: SATB2 Associated Syndrome (SAS) is characterized by severe intellectual deficiency (ID) with limited or absence of speech, behavioral problems, and craniofacial abnormalities. It is caused by de novo heterozygote mutations in the Special AT-rich sequence-Binding Protein 2 (SATB2) gene, which encodes a DNA-binding protein involved in chromatin remodeling. In the brain, SATB2 expression is mostly restricted to the projection neurons in the cerebral cortex and hippocampus. Much of what is known about SATB2 function in brain development comes from studies using homozygous loss-of-function mutant mice. Nonetheless, the heterozygous loss of SATB2 negatively impacts brain development, as identified in SAS patients. Here, we report a dose-dependent requirement for Satb2 in regulating subtype specification for layer 5 subcerebral and all intra-telencephalic neurons, as a modulator of gene expression, through its binding to gene promoters and enhancers of many ID genes. Consequently, Satb2 haploinsufficiency in humans, and in mice as we show here, results in aberrations of cortical projection axons that impact mouse sensory behavior including discriminative touch and vocalization.  Our findings uncover fundamental mechanisms underlying the etiology of SAS.

overall_design: CUT&RUN using antibodies for: H3K4me3, H3K27me3, H3K27ac at embryonic day (E) 12.5; Nr2f1 at E13.5; Lhx2 at E15.5; Cux1, bcl11a, Satb2 at post-natal day (P) 0, with dissected neocortical tissue from wild-type mice

first: Thomas

last: Finn

first: Jeremiah

last: Tsyporin

first: Gabrielle

last: Servito

first: Sol

last: Katzman


title: Integrative Spatial Multi-Omics Reveal Niche-Specific Inflammatory Signaling and Differentiation Hierarchies in Acute Myeloid Leukemia

city: Houston

first: Hussein

last: Abbas

state: Texas

address: 1515 Holcombe Blvd

institute: The University of Texas at MD Anderson Cancer cent

summary: Acute myeloid leukemia (AML) is a clonal disorder characterized by immature blasts and arrested differentiation that primarily affects the bone marrow (BM) and occasionally presents as extramedullary (EM) disease. EM manifestations highlight AML’s adaptability to distinct microenvironments, which we examined using spatial analyses of medullary and EM tissues. We describe a workflow for Visium-based spatial transcriptomics in medullary and EM AML, revealing insights into cell-cell communication and the spatial organization of AML hierarchies. In BM, monocytes and granulocyte-monocyte progenitors colocalized with leukemic populations, sharing molecular signatures with those in EM sample. CXCL12-CXCR4-mediated communication correlated with PI3K/AKT/mTOR signaling in inflammatory niches. Trans-differentiation signals concentrated in AML-infiltrated regions; committed-like AML cells resided in inflammatory niches distant from trabeculae, while primitive-like cells localized near the endosteal niche. GeoMX digital spatial profiling and Opal multiplex fluorescent immunohistochemistry provided orthogonal validation. Overall, our study offers a valuable multimodal resource for exploring AML spatial biology with potential applications in other BM malignancies.

overall_design: Bone Marrow (BM) and Extramedularry (EM) tissue biopsies were routinely collected before treatment initiation from 2 patients with newly diagnosed AML. The samples were formalin fixed, and BM samples were decalcified using 10% formic acid. Following fixation and decalcification, the samples were embedded in paraffin and stored at room temperature before they were cut into 4-µm sections. Visium Spatial Gene Expression (Direct placement; v1) and Visium CytAssist Spatial Gene Expression (CytAssist-enabled; v2) protocols apllied on each BM and EM samples. Libraries sequenced using Illumina NovaSeq 6000.

middle: A

first: Enes

last: Dasdemir


title: Remote limb ischemic conditioning alleviates nonalcoholic steatohepatitis via extracellular vesicle-mediated muscle-liver crosstalk [hEV_RNAseq]

city: Shanghai

first: Yichao

last: Zhao

state: China

address: #160 Pujian Road, Shanghai,China

department: Renji Hospital

country: China

institute: Shanghai Jiaotong University

summary: Nonalcoholic steatohepatitis (NASH) is an advanced form of nonalcoholic fatty liver disease with adverse outcomes that currently lacks approved medication. Muscle-liver interaction has emerged as one of the critical determinants governing NASH progression. Here, we report that remote limb ischemic conditioning (RIC), a clinically validated therapy for distant ischemic organ protection by transient muscle ischemia, significantly alleviated NASH in different mouse models. The anti-NASH effect of chronic RIC treatment was mediated by the muscle-to-liver transfer of extracellular vesicles (EVs) and their cargo microRNAs, leading to elevation of miR-181d-5p in the liver. Hepatic miR-181d-5p overexpression faithfully mirrored the benefits of RIC treatment via suppression of nuclear receptor 4A3. Furthermore, administration of circulating EVs from human volunteers undergoing RIC treatment improved NASH phenotypes and transcriptomic perturbations in primary human hepatocytes and animal models. Our data underscore the translational potential of RIC treatment for NASH management and extend our understanding of muscle-liver crosstalk.

overall_design: Mice were fed a choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD) to induce nonalcoholic steatohepatitis (NASH), and were intravenously injected with circulating extracellular vesicles (EVs) isolated from human volunteers subjected to remote limb ischemic conditioning (RIC)  or sham treatment . Liver tissues were harvested and subjected to RNA-sequencing.


title: Spatial transcriptome analysis for placental tissue of preeclampsia

city: Nagoya

first: Kosuke

last: Yoshida

state: Aichi

address: 65 Tsuruma-cho

country: Japan

institute: Nagoya University

summary: Preeclampsia (PE) is a major pregnancy complication leading to maternal and neonatal mortality. To investigate the spatial gene expression profile of preeclampsia, we performed the Visium FFPE spatial transcriptome analysis.

overall_design: We used formalin-fixed paraffin embedded (FFPE) tissues of preeclampsia and normal placenta. Accoriding to the Visium FFPE protocol, the spatial transcriptome analysis was conducted by CyberomiX (Kyoto, Japan).

first: Seiko

last: Matsuo

first: Akira

last: Yokoi


title: The gene expression profiles of HUVEC cells treated with LIMCH-overexpressed JAR cells-derived small extracellular vesicles.

summary: Preeclampsia is a major pregnancy complication leading to maternal and neonatal mortality, but the detailed pathogenesis remains unelucidated. Extracellular vesicles (EVs) contain proteins and nucleic acids inside and are responsible for intercellular communication. In this study, we investigated the roles of LIMCH1-containing EVs on vascular endothelial cells in vitro.

overall_design: We established LIMCH-1 overexpressed JAR cells and collected EVs using the ultracentrifuge. Then, HUVEC cells were treated with the EVs, and RNA sequencing was performed.


title: The effect of Bcl6-downstream transcription factors on the liver sex difference

city: Isehara

first: Akihide

last: Kamiya

state: Kanagawa

address: 143 Shimokasuya

institute: Tokai University

summary: To confirm the effect of Bcl6-downstream transcription factors, Cux2, Sall1, Trim24, on the liver sex differrence, we performed the gene expression analysis in male and female Cas9-knockin mice.

overall_design: Male and female Cas9-knockin mice (negative control) and  female Cas9-knockin mice with Cux2, Sall1, Trim24 gRNA or Trim24 gRNA alone. These mouse were dissected and RNA extraction was performed from livers for the microarray analyses.


title: CYP2C29 Monooxygenase Promotes CDAA-HFD-Induced Liver Fibrosis

status: Public on Nov 24 2025

city: Durham

first: Brian

last: Bennett

address: 111 T.W. Alexander Drive

institute: NIEHS

summary: Cytochrome P450 (CYP) monooxygenases (predominately the CYP2C and CYP2J isoforms) catalyze the oxidative metabolism of polyunsaturated fatty acids to generate epoxy fatty acids (EpFAs) and fatty acid diols, which are bioactive lipid signaling molecules that regulate inflammation and other important biological processes. Animal and human studies suggest that this pathway is dysregulated in metabolic dysfunction-associated steatohepatitis (MASH) and related liver diseases; however, its functional role in disease development of MASH, especially liver fibrosis, remains poorly understood. Here, we investigated the role of CYP2C29, the predominant CYP2C/2J isoform in mouse livers, in a choline-deficient, L-amino acid–defined, high-fat diet (CDAA-HFD)-induced liver fibrosis model. Compared with wild-type (WT) mice, CYP2C29 knockout (KO) mice exhibited reduced hepatic concentrations of CYP-derived EpFAs and fatty acid diols and attenuated development of liver fibrosis. Transcriptomic analysis further revealed that Cyp2c29 disruption suppressed the hepatic expression of multiple pro-fibrogenic, pro-inflammatory, and pro-angiogenic genes. Together, these findings demonstrate that the CYP monooxygenase pathway promotes the development of CDAA-HFD-induced liver fibrosis and its associated disorders.

overall_design: 7 WT and 7 CYP2C29 KO mice were fed the CDAA-HFD diet (Research Diets, catalog no. A06071302, New Brunswick, NJ; detailed formulation in Table S2) for 16 weeks. Liver RNA was extracted and analyzed by RNA Sequencing

last: Jing

first: Matthew

last: Edin

first: Fred

last: Lih

first: Yige

last: Wang

first: Karen

last: Matsukuma

first: Joan

last: Graves

first: Fuhang

last: Liu

first: Jiacheng

last: Fang

first: Darryl

last: Zeldin

first: Guodong

last: Zhang


title: Remote limb ischemic conditioning alleviates nonalcoholic steatohepatitis via extracellular vesicle-mediated muscle-liver crosstalk

type: Non-coding RNA profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Beckwith-Wiedemann syndrome and large offspring syndrome involve alterations in methylome, transcriptome, and chromatin configuration [RNA-Seq]

first: Rocío

last: Rivera

overall_design: Total RNA sequencing (ribosomal depletion) for skin fibroblast primary cells derived from five control and six LOS bovine day 105 fetuses.

first: Yahan

last: Li

first: Ping

last: Xiao

first: Frimpong

last: Boadu

first: Anna

middle: K

last: Goldkamp

first: Snehal

last: Nirgude

first: Jianlin

last: Cheng

first: Darren

middle: E

last: Hagen

first: Jennifer

last: Kalish


title: Analysis of Xenopus tropicalis stage NF33 regional gene expression in micro-dissected domains along anterior-posterior axis and Hedgehog, Retinoic Acid, and Rfx6 dependent gene expression in micro-dissected NF33 foregut tissue (mesoderm + endoderm)

city: Cincinnati

first: Aaron

last: Zorn

state: OH

address: 240 Albert Sabin Way, Bld S

department: Developmental Biology

institute: Cincinnati Children's Hospital and Medical Center

summary: To define regional gene expression domains at stage NF33 along the Xenopus tropicalis embryonic anterior-posterior axis and also identify Rfx6, Retinoic Acid, and Hedgehog regulated genes at stage NF33.

overall_design: Retinoic Acid, and Hedgehog regulated genes at stage NF33.  Tissue was micro-dissected from wild-type Xenopus tropicalis embryos at stage NF33 (Adult Xenopus NXR_1018, Xtr.NigerianSt549, obtained from National Xenopus Resource Stock Center, Woods Hole, MA; RRID:SCR_013731).  Pools of n=8 explants (mesoderm+endoderm)s were harvested as 1 biological replicate.  3 such biological replicates per condition were generated.  RNA was extracted using the Machery-Nagel RNA isolation kit (740955) according to manufacturer's instructions. 40uM cyclopamine (Tocris #1623), 100nM BMS493 (Tocris #3509) treatment from NF15-33.   1ng total Rfx6 morpholino oligo or 5bp mismatch oligo  (500pg per cell) was injected at 2-cell stage.

first: Scott

last: Rankin

first: Jacek

last: Biesiada


title: Titin truncating variant associated with atrial fibrillation leads to cell type-specific effects in atrial and ventricular patient hiPSC-CM models

city: Vancouver

first: Kate

last: Huang

state: BC

address: 166-1081 Burrard Street

department: Department of Medicine - Centre for Heart Lung Innovation

country: Canada

institute: University of British Columbia

summary: Protein truncating mutations in the titin gene are associated with increased risk of atrial fibrillation (AF). However, little is known regarding the underlying pathophysiology. We identified a heterozygous titin truncating variant in a patient with unexplained early-onset AF using whole exome sequencing. We used atrial and ventricular patient induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs), CRISPR/Cas9 genetic correction, and engineered heart tissue (EHT) constructs to evaluate the impact of the titin truncating variant on electrophysiology, sarcomere structure, contractility, and gene expression.

overall_design: Atrial-like and ventricular-like patient iPSC-CMs were cultured in a 2D/monolayer format or used to generate EHT constructs. 3 biological replicates per sample type (culture format, cell type, titin genotype) were sequenced to compare the effects of the titin truncating variant on gene expression in atrial and ventricular iPSC-CMs and EHTs.

first: Leili

last: Rohani

first: Yinhan

last: Luo

first: Anne

last: Haegart

first: Stanislav

last: Volik

first: Funda

last: Sar

first: Stéphane

last: LeBihan

first: Colin

last: Collins

first: Zachary

last: Laksman


title: HIF Inhibitor 32-134D Combined with Anti-PD1 Therapy Blocks Murine Hepatocellular Carcinoma Growth and Eradicates Tumors

city: Baltimore

first: Shaima

last: Salman

state: MD

address: 733 N. Broadway

department: Institute for Cell Engineering

institute: JHU

summary: Hepatocellular carcinoma (HCC) is a major cause of cancer mortality worldwide and available therapies, including immunotherapies, are ineffective for many patients. HCC is characterized by intratumoral hypoxia, and increased expression of hypoxia-inducible factor (HIF) 1a in the diagnostic biopsy is associated with patient mortality. Here we report the development of 32-134D, a low-molecular-weight compound that effectively inhibits gene expression mediated by HIF-1 and HIF-2 in HCC cells, and blocks human and mouse HCC tumor growth as monotherapy. In immunocompetent mice bearing Hepa1-6 HCC tumors, addition of 32-134D to anti-PD1 therapy increased the rate of tumor eradication from 25% to 67%. Treated mice showed no changes in appearance, behavior, body weight, hemoglobin or hematocrit. 32-134D altered the expression of a large battery of genes encoding proteins that mediate angiogenesis, glycolytic metabolism, and responses to innate and adaptive immunity. This altered gene expression led to significant changes in the tumor immune microenvironment, including a decreased percentage of tumor-associated macrophages and myeloid-derived suppressor cells, which mediate immune evasion, and an increased percentage of CD8+ T cells and natural killer cells, which mediate anti-tumor immunity. Taken together, these preclinical findings suggest that combining 32-134D with immune checkpoint blockade may represent a breakthrough therapy for HCC.

overall_design: Hep3B cells were seeded into 6-well plates in three biological replicates and exposed to 20% or 1% O2 for 24 hours. Total RNA was isolated using TRIzol and treated with DNase (Qiagen). Library preparation and sequencing using the NovaSeq6000 platform (Illumina) were performed by Johns Hopkins Genetics Resources Core Facility High-Throughput Sequencing Center. The Fastq files were subjected to quality check and analyzed by Genialis (https://www.genialis.com/). Differential expression results with FDR < 0.05 and mRNA fold change > 1.5 were used as a cutoff for further downstream analysis.

first: Gregg

middle: L

last: Semenza


title: Analysis of Xenopus laevis foregut mesoderm+endoderm explants from RA or Rfx6 loss of function at 3 stages: NF25, NF33, NF38.

summary: To define Retinoic Acid and Rfx6 regulated genes in Xenopus laevis foregut tissue, during developmental window of NF15 to NF38, we micro-dissected foregut explants containing both mesoderm and endoderm at stage NF15 from embryos injected with a translation blocking Rfx6 Morpholino Oligo or a control 5bp mismatch oligo, cultured explants, and harvested at NF25, NF33, or NF38 for bulk RNA-seq analysis.  For RA loss of function, foregut explants containing both mesoderm and endoderm were treated with pan RAR antagonist BMS493 from NF15 to NF33, harvested either at NF25 or NF33 for bulk RNA-seq analysis.

overall_design: Foregut mesoderm+endoderm explants were micro-dissected from wild-type Xenopus laevis embryos at stage NF15 and cultured to stage NF25, NF33, or NF38  (Adult Xenopus NXR_0031 obtained from National Xenopus Resource Stock Center,  Woods Hole, MA; RRID:SCR_013731).  Pools of n=8 foregut mesoderm+endoderm explants were harvested at 1 biological replicate.  3 such biological replicates per condition were generated.  RNA was extracted using the Machery-Nagel RNA isolation kit (740955) according to manufacturer's instructions.


title: scRNA-seq of T cells isolated from mouse skin treated with SspA and HDM

city: Charlestown

first: Jin Mo

last: Park

address: 149 Thirteenth Street

department: Cutaneous Biology Research Center

institute: Massachusetts General Hospital

summary: Treatment of mouse skin with the staphylococcal protease SspA and house dust mite extract (HDM) results in dermatitis. Using a 10x Genomics scRNA-seq platform, we analyzed T cells isolated from inflamed skin after treatment with epicutaneous SspA and HDM.

overall_design: T cells isolated from the mouse skin 7 days after SspA and HDM treatment were subjected to fluorescence-activated cell sorting and scRNA-seq.

first: Yanek

last: Jiménez-Andrade


title: Fibroblast histone deacetylase-1 promotes kidney interstitial fibrosis following ischemia-reperfusion injury.

city: Birmingham

first: Kelly

middle: Anne

last: Hyndman

state: Al

address: 1918 University BLVD

department: Medicine

institute: University of Alabama at Birmingham

summary: Histone deacetylases (HDAC) are epigenetic regulators of trascription.  Preclinical studies have determined that HDAC inhibition can prevent kidney interstitial fibrosis in acute kidney injury.  The purpose of this study was to determine if overexpression of HDAC1 in kidney fibroblasts significantly affects the fibroblast transcriptome.  Transforming growth factor beta-1 (TGFB1) causes fibroblasts to differentiate into myofibroblasts leading to extracelluar matrix secretion.  A subset of our cells were treated with TGFB1 for 24 hours to determine if this in combination with HDAC1 overexpression significantly alters the transcriptomes.

overall_design: Normal rat kidney fibroblast cells (NRK49F, ATCC) were transfected with either empty vector (pcDNA3.1) or human HDAC1-Flag plasmids (Addgene #13820, PMCID: PMC19648).  The cells were then treated with 2 ng/ml TGFB1 for 24 hours before RNA isolation and library preparation and sequencing.

first: Huy

last: Nguyen

first: Kelly A


title: Global analysis of Arabidopsis thaliana DICER-LIKE1 RNA substrates

city: Zürich

first: Alexis

last: Sarazin

address: Universitätstrasse 2

country: Switzerland

institute: ETH

summary: By introducing point mutations in the RNAse III domains of plant DCL1 we generated Catalytically inactive (ci) DCL1. Plant expressing DCL1ci show an increase stability of the RNA/DICER complexes and therefore represent an innovative strategy to perform a direct and systematic analysis of DCL1 substrates.

overall_design: Triplicates of pDCL1-DCL1ci-HA/dcl1-7 and respective dcl1-7 control plants RIP-seq

first: Nicolas

middle: G

last: Bologna

first: Olivier

last: Voinnet


title: Effect of knockdown of PPP2R2A on gene expreesion in NSCLC cells

city: Columbus

first: Zhaojun

last: Qiu

state: Ohio

address: 460 West 12th Ave

institute: The Ohio State University

summary: To investigate the biological roles of PPP2R2A, we conducted bulk RNA-sequencing to assess the impact of PPP2R2A knockdown using two shRNAs in two NSCLC cell lines.

overall_design: To broadly explore the signaling pathways modulated by PPP2R2A, we initially conducted an analysis of global transcriptional changes using RNA-sequencing of NSCLC A549 and H1299 cell lines. These cells were subjected to either shRNA-mediated PPP2R2A KD or a scrambled shRNA control, achieved by using two different shRNAs targeting distinct regions of PPP2R2A.

first: Junran

last: Zhang

first: Deepika

last: Singh


title: Oral bacterial dysbiosis drives autoimmune heart disease via molecular mimicry

city: Toronto

first: Abdelahhad

last: Barbour

address: 124 Edward st

institute: University of Toronto

summary: The link between poor oral health and heart failure progression has long been recognized but remains limited to epidemiological associations without mechanistic understanding. Here, we performed Xenium in situ on heart tissue post-transplantation to detect CD4 T-cells that generated in inflamed gingiva during periodontal diseases.

overall_design: Eight human heart tissue after explantation were used to map the CD4 T-cell profile on them on heart failure patients with oral inflammation and heart failure patients with no oral inflammation.

first: Sina

last: Zargaran

first: Michael

last: Glogauer


title: Regulation of murine follicle-stimulating hormone β subunit transcription by newly identified enhancers [scATAC-seq; Mouse Pituitaries]

city: Los Angeles

first: Michel

last: Zamojski

state: California

address: 8700 Beverly Boulevard

department: Regenerative Medicine Institute

institute: Cedars-Sinai Medical Center

summary: Activin-class ligands of the TGFβ family induce follicle-stimulating hormone (FSH) production by pituitary gonadotrope cells in mice via the actions of the transcription factors SMAD3, SMAD4, and FOXL2, which bind to cis-elements in the FSHβ subunit (Fshb) promoter. An enhancer region for murine Fshb transcription was identified in vitro. However, deletion of the region using CRISPR-Cas9 did not affect FSH synthesis or secretion in mice. Using single-nucleus ATAC-seq of whole pituitaries, we identified three additional chromatin regions upstream of Fshb open exclusively in murine gonadotropes. We found these regions, as well as the Fshb gene, to be fully or partially closed in gonadotropes of FSH-deficient mice with genetically or pharmacologically inactivated activin type II receptors. The data indicate that there may be as many as four activin-sensitive enhancers upstream of murine Fshb. This data is part of a SuperSeries that includes bulk ATAC-seq and ChIP-seq data of gonadotrope-derived LβT2 cells, that substantiate the above conclusions.

overall_design: Two sgRNAs were designed to target a previously identified enhancer region for murine Fshb transcription. Enhancer-knockout mice were generated using CRISPR-Cas9, after electroporation of the two sgRNAs and the Cas9 protein into zygotes. Pituitary glands were dissected from 8- to 10-week-old wild-type and enhancer-knockout mice, and prepared for scATAC-sequencing.  Pituitary glands were similarly collected and processed from wild-type mice treated with either IgG or Bimagrumab, an activin type II receptor antagonist.

first: Yangfan

last: Jin

first: Luisina

last: Ongaro

first: Shinsuke

last: Onuma

first: Hailey

last: Schultz

first: Gauthier

last: Schang

first: Stuart

last: Sealfon

first: Daniel

last: Bernard

first: Frederique

last: Ruf-Zamojski


title: Pyruvate Kinase Muscle (PKM)-2 promotes CD4 T cell survival by regulating pyruvate oxidation during homeostasis and expansion

city: Frederick

first: Ian

last: Bettencourt

address: 1050 Boyles Street Rm 31-16A

department: CIL

institute: NCI

summary: Glycolysis is an essential metabolic pathway for rapidly expanding T cells, but role of Pyruvate Kinase Muscle (PKM) 1 and 2 in regulating this process is underappreciated.  Here, using a pharmacological activator and targeted deletion of PKM2 in T cells we delineated distinct functions of PKM1 and 2 in regulating CD4 T cell survival during homeostasis and expansion.  Expanding PKM2-deficient CD4 T cells increased PKM1 expression with associated mitochondrial ROS-mediated cell death. Examination of T cell compartments revealed PKM2-deficient CD4 T cells were unaltered in the thymus but were significantly reduced in peripheral tissues as mice aged. The failure of PKM1 to protect CD4 T cells in the absence of PKM2 resulted in protection from T cell mediated colitis as pathogenic cell numbers were significantly reduced resulting in less severe disease compared to control cells. This study shows PKM2 is critical for CD4 T cell survival during expansion and homeostasis.

overall_design: Naïve T cells were isolated from spleen and lymph nodes of wild type or Pkm2 knockout mice.  Cells were cultured for 24 hours with IL7 alone (resting) or IL7 plus plate-bound aCD3 and aCD28 antibodies.

first: Jeff

middle: J

last: Subleski

first: Erika

last: Palmieri

first: Jessica

middle: F

last: Walls

first: Steven

last: Hsu

first: Timothy

middle: R

last: Gower

first: Scott

last: Durum

middle: W

last: McVicar


title: RIPK1 regulates beta-cell fate and autoimmunity in type 1 diabetes

city: Indianapolis

first: Madeline

last: Peyton

state: IN

address: 410 W 10th St

institute: Indiana University

summary: Type 1 diabetes (T1D) results from autoimmune destruction of pancreatic beta-cells. RIPK1, a key regulator of cell death and inflammation, has been implicated in beta-cell responses to cytokines and immune attack. To investigate RIPK1's role in beta-cell fate during autoimmune stress, we generated CRISPR/Cas9-mediated Ripk1-deleted NIT-1 beta-cells and exposed them to TNFα+IFNγ or self-reactive diabetic NOD splenocytes. Using RNA-seq, kinome profiling, flow cytometry, and small molecule inhibitors, we demonstrate that RIPK1 deletion or pharmacological inhibition protects beta-cells from cytokine-induced and autoimmune cell death. This study provides insights into RIPK1-dependent pathways in beta-cell vulnerability and suggests potential therapeutic strategies for T1D.

overall_design: 2x2 factorial design: Genotype (Control NIT-1 vs Ripk1-deleted NIT-1) x Treatment (Vehicle vs TNFα+IFNγ). NIT-1 beta-cells from NOD mice; CRISPR-engineered Ripk1Δ vs control cells. Cells treated with TNFα (40 ng/mL) + IFNγ (100 ng/mL) or vehicle. Six biological replicates per condition (24 samples total). Multi-modal analyses: RNA-seq, kinome profiling, flow cytometry, small molecule inhibitor testing.


title: Chidamide and Anlotinib act synergistically in Jurkat cells by inhibiting the Hippo signaling pathway

city: luzhou

first: Tao

last: Ma

state: si chuan

address: taiping jie

department: Department of Hematology

institute: Affiliated Hospital of Southwest Medical University

summary: T-cell Acute Lymphoblastic Leukemia (T-ALL) is an aggressive hematologic malignancy characterized by drug resistance, relapse and poor prognosis. Chidamide, a histone deacetylase inhibitor, has shown epigenetic therapeutic potential in T-ALL, but its efficacy is limited and acquired drug resistance exists. To overcome these limitations, we systematically evaluated the synergistic Combination effects of three drug candidates, OTX-015 (BET inhibitor), Metformin (metabolic regulator), and Anlotinib (multitargeted tyrosine kinase inhibitor), with Chidamide.

overall_design: The human T-ALL Jurkat cell line was treated under three conditions for 48 hours: 1) Chidamide (5 μM) alone, 2) Anlotinib (1 μM) alone, and 3) Combination of Chidamide (5 μM) and Anlotinib (1 μM). Untreated cells served as control. Total RNA was extracted from biological replicates for each group and subjected to RNA sequencing (RNA-seq) on an Illumina platform with 2×150 bp paired-end reads. Differential gene expression analysis was performed to identify genes and pathways modulated by the treatments.

first: Chunli

last: Zhang


title: Microglial reactivity and neuroinflammation-driven in motivational behaviors are regulated by Orai1 channels

city: Chicago

first: Rogan

last: Grant

state: IL

address: 303 E Superior St, Simpson Querrey Biomedical Research Center 5-522

department: Pulmonary and Critical Care Medicine

institute: Northwestern University

summary: Microglia are the primary immune cells of the central nervous system (CNS) and act as the first line of defense against disease and injury in the brain by releasing inflammatory molecules and clearing dead cells. Microglial functions are tightly regulated by dynamic transitions between homeostatic and reactive cell states, which can either exacerbate or resolve CNS inflammation. While Ca2+ signaling is known to regulate specific microglial functions such as phagocytosis, its role in controlling these critical state transitions remains poorly understood. Here, we identify the Orai1 Ca2+ channel as a key regulator of microglial plasticity and inflammation-induced behavioral dysfunction in mice. Transcriptomic and metabolomic analyses revealed that conditional Orai1 deletion in microglia suppressed proinflammatory programs linked to immunity and inflammation. Conversely, Orai1 knockout increased expression of growth factors and anti-inflammatory mediators, including BDNF, ARG1, and the mitochondrial metabolite, itaconate. In a model of CNS inflammation induced by peripheral inflammatory challenge with lipopolysaccharide (LPS), Orai1 deficiency attenuated microglial and astrocyte reactivity and reduced hippocampal levels of the cytokines IL-1β and IL-6. Consistent with these cellular changes, microglial Orai1 knockout mice were protected against LPS-induced changes in motivational, depression-like behaviors, including impairments in reward-seeking and escape responses. These findings establish Orai1 channels as central regulators of microglial cell plasticity and neuroimmune responses, highlighting their therapeutic potential for neuroinflammatory disorders.

overall_design: Primary microglia from Orai1fl/fl or Orai1fl/fl Cx3CR1-cre/ERT2 were cultured from neonates or FACS purified from adult animals and  treated with Tg+PDBu, LPS, or vehicle control  for 6h or 24h.

first: Kaitlyn

last: DeMeulenaere

first: Megan

last: Martin

first: Hiam

last: Valencia

first: Jelena

last: Radulovic

first: Michael

last: Salter

first: Murali

last: Prakriya


title: Downregulation of motility during stress conditions requires stressosome input in Listeria monocytogenes

city: Berlin

first: Dmitriy

last: Ignatov

address: Charitéplatz 1

country: Germany

institute: Max Planck Unit for the Science of Pathogens

summary: Motility in Listeria monocytogenes is negatively regulated within the mammalian host in response to temperature through a complex interplay between positively acting and negatively acting regulators. Motility genes are expressed under saprophytic conditions where the temperature is 30ºC or less. Motility is costly on cellular resources due to the large molecular structures that need to be synthesized, and the proton motive force required to produce flagellar rotation. Under stressful environmental conditions there is a trade off between motility and energy conservation and bacteria typically repress motility when adverse conditions are encountered. Here we investigated the impact of the SigB-mediated general stress response on the regulation of motility in L. monocytogenes and sought to elucidate the regulatory steps involved. We show that an rsbX mutation that compromises the ability to inactivate the stressosome, the sensory hub at the top of the SigB activation pathway, results in motility repression. Escaping from this repressed state occurred through the acquisition of mutations that decreased SigB activity. Flagellar expression was abolished in these mutants and a transcriptomic analysis revealed that the entire flagellar operon is strongly repressed. The motility anti-repressor gene gmaR was also downregulated in an ΔrsbX background. These effects can be reversed by providing functional copies of rsbX or gmaR in trans. Deletion of an internal SigB dependent promoter that produces an antisense RNA for the large motility operon (including gmaR) restores the ability to produce flagellin in the ΔrsbX background. A similar phenotype is observed by deleting MogR, which acts as a negative regulator of flagella motility. Stressosome mutations that negatively affect SigB activity   result in derepressed motility whereas those that increase SigB activity result in a decreased motility phenotype similar to the ΔrsbX strain. Overall, the data indicate that stress sensing via the stressosome negatively impacts motility in L. monocytogenes and shows that the general stress response can take priority over motility when L. monocytogenes encounters harsh environmental conditions.

overall_design: RNA-seq profiles of the wild-type and ΔrsbX strains of L. monocytogenes EGD-e.  Bacteria  were grown in BHI medium at 23°C under normal laboratory light conditions until OD600 = 0.85 when bacteria were harvested, RNA was isolated and subjected to RNA-seq.

first: Teresa

last: Tiensuu

first: Jörgen

last: Johansson


title: RNA-seq analysis of human cells overexpressing a novel short XIST transcript isoform

city: Bronx

first: Kevin

middle: Sean

last: O'Leary

state: NY

address: 1301 Morris Park Ave

department: Genetics

institute: Albert Einstein College of Medicine

summary: We identified a novel short XIST isoform (sXIST) that is expressed in both male and female peripheral glia specifically. To study its functions, we overexpressed a fragment of it in the human HEK293T cell line and performed RNA-seq analysis of the sXIST overexpressing cells and GFP controls. We found that the up-regulated genes were enriched in pathways related to cilia functions and many share miRNA binding sites with the sXIST.

overall_design: HEK-293T cells were transfected with a sXIST construct or GFP. mRNA from the virus-infected cells was iisolated and analyzed by RNA-seq.

first: Meng-Yen

last: Li

first: Elena

last: Ezhkova

first: Deyou

last: Zheng


title: Differentially expressed microRNAs between WT and AtC3H3 OX, and WT and AtC3H26 OX

city: Busan

first: Yong-Hwan

last: Moon

address: Busandaehak-ro 63Beon-gil 2, Geumjeong-gu

department: Molecular Biology

country: South Korea

institute: Pusan National University

summary: CCCH-type zinc finger (ZF) proteins comprise a diverse family of RNA- and DNA-binding regulators that play crucial roles in plant growth, development, and responses to stress. While tandem CCCH zinc finger (TZF) proteins have been extensively studied, the molecular functions of non-TZF members remain less understood. Here, we characterized Arabidopsis thaliana AtC3H26, a non-TZF CCCH protein and paralog of AtC3H3, and demonstrated its role as a ribonuclease (RNase) in salt stress tolerance and phosphate (Pi) homeostasis. In vitro assays confirmed that AtC3H26 degrades RNA substrates in a dose-dependent manner. mRNA sequencing of OX lines revealed a significant upregulation of phosphate starvation–responsive genes, including SPX1, PS2/PECP2, and SRG3/GDPD1, whereas small RNA sequencing identified the downregulation of miR399 and miR827, which are involved in the Pi starvation response. AtC3H26 OX plants accumulated higher Pi levels than wild type, suggesting that AtC3H26 modulates Pi accumulation. Collectively, these findings establish AtC3H26 as a novel cytoplasmic RNase that coordinates RNA turnover with stress and nutrient signaling in Arabidopsis.

overall_design: Differential gene expression between WT and AtC3H3 OX, as well as between WT and AtC3H26 OX, was examined at 10 days after germination under SD conditions (8-hour-light and 16-hour-dark cycle) on a microRNA level.

first: Hye-Yeon

last: Seok

first: Yunseong

last: Jang

first: Sun-Young

last: Lee

first: Dahyun

last: Kim


title: Cardio-immune Interaction via a Clusterin-TLR4 Axis Suppresses Inflammation and Triggers Regeneration

city: Shanghai

first: Qi

last: Tang

address: 320 Yue Yang Road

institute: CAS Center for Excellence in Molecular Cell Science

summary: This study aimed to investigate the transcriptional effects of BMP2 treatment on neonatal mouse cardiomyocytes. Primary cardiomyocytes were treated with either PBS or BMP2 and subjected to bulk RNA sequencing to profile BMP2-induced changes in gene expression. To further dissect the underlying signaling mechanisms, cells were co-treated with the BMP2 receptor inhibitor Dorsomorphin, allowing assessment of the contribution of the canonical SMAD pathway to BMP2-mediated transcriptional regulation. Comparative transcriptomic analysis across treatment groups provides new insights into the role of the BMP2–SMAD signaling axis in cardiomyocyte gene regulation.

overall_design: To investigate the transcriptional effects of BMP2 treatment on neonatal mouse cardiomyocytes, we performed bulk RNA-seq on CMs treated with either BMP2 or PBS. In addition, cells were treated with the BMP2 receptor inhibitor Dorsomorphin to assess the contribution of the downstream canonical SMAD signaling pathway to gene expression.

first: Lei

last: Fan

first: Yutong

last: Wang


title: Bulk RNA sequencing analysis of human airway basal stem cells stimulated by BALF from mouse fibrotic lungs and normal lungs

first: Wei

last: Zuo

state: Shanghai

address: No. 500 Zhennan Road, Putuo District, Shanghai

institute: University

summary: To understand how the microenvironment of fibrotic lungs affects the behavior of airway basal stem cells (BCs), we constructed a mouse model of pulmonary fibrosis using bleomycin induced injury, and collected bronchoalveolar lavage fluid (BALF) from the control group and bleomycin damaged mice on day 14 after injury. Then, human BCs were stimulated with control or bleomycin BALF in culture and subjected to bulk RNA sequencing. Transcriptomic analysis showed that basal stem cells stimulated with bleomycin BALF upregulated genes related to chemotaxis and migration, and activated the TGF-β pathway.

overall_design: Human BCs were stimulated with PBS or bleomycin BALF in culture for 48 hours and subjected to bulk RNA sequencing. Total RNA was extracted from Human basal cells using Trizol (15596018CN, Invitrogen), and RNA extraction was performed according to the manufacturer’s instructions. Subsequently, the extracted RNA was treated with Dnase I (Invitrogen, Life Technologies, USA) to remove any contaminating DNA. Utilize the Novogene platform for cDNA library construction and sequencing.

first: Tingting

last: Zou

first: Shiyu

last: Zhang

first: Mingzhe

last: Liu


title: A deep-time landscape of plant cis-regulatory sequence evolution

city: Amherst

first: Kirk

last: Amundson

state: MA

address: 240 Thatcher Rd

department: Biology

institute: University of Massachusetts Amherst

summary: Developmental gene function is conserved over deep time, but similar cis-regulatory sequence conservation is rarely found. However, rapid sequence turnover, paleopolyploidy, structural variation, and limited phylogenomic sampling have impeded conserved non-coding sequence (CNS) discovery. Using Conservatory, an algorithm that leverages microsynteny and iterative alignments to map CNS-gene associations over evolution, we uncovered ~2.3M CNSs,  including over 3,000 predating angiosperms, from 284 plant species spanning 400 million years of diversification. Ancient CNSs were enriched near developmental regulators, and mutagenizing those near HOMEOBOX genes produced strong phenotypes. Tracing CNS evolution uncovered key principles: CNS spacing varies, but order is conserved; genomic rearrangements form new CNS-gene associations; and ancient CNSs are preferentially retained among paralogs but often are lost as cohorts or evolve into lineage-specific CNSs.

overall_design: Six replicates of ATAC-seq data from young leaves of Solanum lycopersicum cv. M82 were generated on an Illumina MiSeq SY-410-1003 instrument. Two replicates of Hi-C data from Solanum lycopersicum cv. M82 anantha inflorescences (meristematic enriched tissue) were generated on an Illumina NovaSeq 6000 with SP flow cell configuration.

first: Zachary

middle: B

last: Lippman


title: Heterogeneous therapy-resistant cancer cells have distinct and exploitable drug sensitivity profiles

city: Philadelphia

first: Gianna

middle: T

last: Busch

state: Pennsylvania

address: 415 Curie Blvd, 415 CRB

department: Bioengineering

institute: University of Pennsylvania

summary: Resistance to targeted therapies is a significant clinical problem, but eliminating resistant cancer cells has proven difficult. One potential reason for this difficulty is heterogeneity in the resistant population: even genetically homogeneous cancer cell populations can give rise to many resistant subtypes, each potentially with specific second-line drug vulnerabilities. Using high-throughput drug screening of clonally-derived resistant clones with varying transcriptomes and morphologies, we show that each clone had a distinct drug sensitivity profile. These results suggested that there are drugs that are effective against only subsets of resistant populations but in combination eliminate a large proportion of the resistant population. Using the individual clone sensitivity profiles, we prospectively identified combinations that were highly effective at eliminating most of the resistant population. Our results demonstrate the effectiveness of “subpopulation-directed synergy”, showing that considering population heterogeneity can reveal therapeutic opportunities otherwise masked by population averages, offering new strategies to combat therapy resistance.

overall_design: RNA-seq profiling of clonal, drug-naïve melanoma cells (WM989 A6-G3) and resistant clones generated by isolating and expanding colonies generated by treating this parental cell line with vemurafenib or combination dabrafenib/trametinib

first: Ryan

middle: H

last: Boe

first: Jingxin

last: Li

first: Robert

last: Gruener

first: Miles

last: Arnett

first: Pavithran

last: Ravindran

first: Meenhard

last: Herlyn

first: R Stephanie

last: Huang

first: Arjun

last: Raj


title: Remote limb ischemic conditioning alleviates nonalcoholic steatohepatitis via extracellular vesicle-mediated muscle-liver crosstalk [EV miRNA]

first: Yichao

last: Zhao

address: #160 Pujian Road, Shanghai,China

institute: Shanghai Jiaotong University

summary: Nonalcoholic steatohepatitis (NASH) is an advanced form of nonalcoholic fatty liver disease with adverse outcomes that currently lacks approved medication. Muscle-liver interaction has emerged as one of the critical determinants governing NASH progression. Here, we report that remote limb ischemic conditioning (RIC), a clinically validated therapy for distant ischemic organ protection by transient muscle ischemia, significantly alleviated NASH in different mouse models. The anti-NASH effect of chronic RIC treatment was mediated by the muscle-to-liver transfer of extracellular vesicles (EVs) and their cargo microRNAs, leading to elevation of miR-181d-5p in the liver. Hepatic miR-181d-5p overexpression faithfully mirrored the benefits of RIC treatment via suppression of nuclear receptor 4A3. Furthermore, administration of circulating EVs from human volunteers undergoing RIC treatment improved NASH phenotypes and transcriptomic perturbations in primary human hepatocytes and animal models. Our data underscore the translational potential of RIC treatment for NASH management and extend our understanding of muscle-liver crosstalk.

overall_design: Mice were subjected to Remote limb ischemic conditioning (RIC) or sham treatment. Then, circulating extracellular vesicles (EVs) were isolated and subjected to miRNA-sequencing.


title: A late cytoplasmic surveillance pathway ensures ribosome integrity

city: Austin

first: Can

last: Cenik

state: Texas

address: 2500 Speedway

department: Department of Molecular Biosciences

institute: University of Texas

summary: Errors in ribosome assembly can produce defective subunits that can lead to aberrant translation events. How faulty ribosomes are recognized and whether they are recognized during assembly or translation remains poorly understood. We utilized a mutation in the ribosomal protein uL16 to track defective 60S subunits through the biogenesis and translation pathways. This mutation deletes a critical loop in the P site and arrests pre-60S particles during late cytoplasmic maturation. However, simultaneous mutations in the late biogenesis factors Nmd3 and Tif6 bypass this block, releasing defective ribosomes into the translational pool. Cryo-EM and selective ribosome profiling reveal that these ribosomes can form peptide bonds, but stall predominantly at the first few codons. We show that the uL16 mutant ribosomes are detected and targeted for degradation during biogenesis and that they escape degradation if they enter translation. We identify Reh1 as a non-canonical ribosome assembly factor that is required for this surveillance pathway.

overall_design: Selective ribosome profiling of  uL16-loop mutant ribosomes in S.cerevisiae.

first: Ruta

last: Chitale

first: Kaoling

last: Guan

first: Shilpa

last: Rao

first: Caroline

last: Wang

first: David

last: Taylor

first: Arlen

last: Johnson


title: Bulk RNA sequencing analysis of Ctrl/PROTAC human airway basal stem cells stimulated by TGF-β

first: Wei

last: Zuo

summary: To evaluate how TGF-β regulates human airway basal stem cells (BCs), we used PROTAC to inhibit TGF-β signaling in BCs and exposed Ctrl-BCs and PROTAC-BCs to recombinant TGF-β1 for 24 hours, followed by bulk RNA sequencing analysis. We found that multiple cell cycle related and proliferation related genes were upregulated in PROTAC-BCs compared to Ctrl-BCs, while the expression levels of cell migration related genes were reduced in the PROTAC-BCs group, and genes related to barrier formation and stem cell differentiation were also downregulated.

overall_design: Ctrl-BCs and PROTAC-BCs were stimulated with 10 ng/ml recombinant TGF-β1 in culture for 24 hours and subjected to bulk RNA sequencing. Total RNA was extracted from BCs using Trizol (15596018CN, Invitrogen), and RNA extraction was performed according to the manufacturer’s instructions. Subsequently, the extracted RNA was treated with Dnase I (Invitrogen, Life Technologies, USA) to remove any contaminating DNA. Utilize the Novogene platform for CDNA library construction and sequencing.

first: Tingting

last: Zou

first: Shiyu

last: Zhang

first: Mingzhe

last: Liu


title: Regulation of murine follicle-stimulating hormone β subunit transcription by newly identified enhancers [ATAC-seq; LβT2 cells]

first: Michel

last: Zamojski

summary: Activin-class ligands of the TGFβ family induce follicle-stimulating hormone (FSH) production by pituitary gonadotrope cells in mice via the actions of the transcription factors SMAD3, SMAD4, and FOXL2, which bind to cis-elements in the FSHβ subunit (Fshb) promoter. An enhancer region for murine Fshb transcription was identified in vitro. The region, however, did not significantly alter basal or activin-stimulated murine Fshb promoter-reporter activity in LβT2 cells. In contrast, three other open chromatin regions enhanced basal and activin A-stimulated Fshb promoter-reporter activity in LβT2 cells, with the two most distal showing the greatest effects. Using ATAC-seq we show that these two regions were open in response to activin A in LβT2 cells.  This data is part of a SuperSeries that includes ChIP-seq data further showing that these regions exhibited enrichment of the enhancer mark H3K27ac, and were bound by SMAD2/3 and FOXL2 in response to activin A in LβT2 cells. The SuperSeries also includes scATAC-seq data of whole mouse pituitaries indicating that there may be as many as four activin-sensitive enhancers upstream of murine Fshb.

overall_design: LβT2 cells were cultured in 12-well plates, and then treated with either vehicle or activin A, after which cells were collected, trypsinized and prepared for ATAC-sequencing.

first: Frederique

last: Ruf-Zamojski

first: German

last: Nudelman

first: Hailey

last: Schultz

first: Natalia

last: Mendelev

first: Daniel

last: Bernard

first: Stuart

last: Sealfon


title: Aldosterone-induced myotube to fibroblast signals

first: Jill

last: Rafael-Fortney

address: 460 W. 12th Ave

department: Physiology & Cell Biology

summary: Here we report the gene expression data following bulk RNA sequencing used to identify the aldosterone-mediated cellular communication from cultured myotubes to fibroblasts isolated from neonatal hind limbs of wild-type (C57) and dystrophic mice (mdx). We found distinct transcriptional changes induced by aldosterone in myotubes, which in turn resulted in distinct transcriptional changes in the CM-treated fibroblasts.

overall_design: Bulk RNA sequencing was performed on vehicle (DMSO, 0.001% v/v) or aldosterone (1 µM)-treated C57 and mdx myotubes following 48 hrs of treatment, and conditioned media (CM)-treated C57 and mdx fibroblasts. CM was collected from myotubes following 15 hrs of incubation with fresh media after 48 hrs of vehicle or aldosterone treatment. Primary myogenic progenitor cells (which were differentiated to myotubes) and fibroblasts were isolated from hindlimbs of neonatal (<5-day-old) C57 and mdx mice.

first: Swathy

last: Krishna

first: Chetan

last: Gomatam

first: Reva

last: Reddy

first: Jeovanna

last: Lowe

first: Christoph

last: Lepper


title: Single-Cell Mapping of Alternative Splicing Linked to Checkpoint Immunotherapy Response

city: Leuven

first: Jieyi

last: Xiong

address: Herestraat 49, box 912

country: Belgium

institute: VIB-KULeuven Center for Cancer Biology

summary: Evidence suggests that alternative RNA splicing (AS) plays a critical role in tumor biology and may contribute to the generation of tumor antigens. Here, we develop a method to detect AS in short-read single-cell 5’-RNA-sequencing data, allowing us to uniquely characterize the heterogeneity and dynamic changes in AS in individual cell types within the tumor microenvironment. We identify numerous splicing events specific to either cancer cells or stromal cell types, or for triple-negative versus estrogen receptor-positive breast cancers. By correlating these splice events with expression of splicing regulators in individual cells, we also identify their potential mediators. For instance, we identify and functionally validate the Epithelial Splicing Regulatory Protein-1 (ESRP1) to drive AS in breast cancers responding to immune checkpoint blockade (ICB). Prioritization of splicing events based on their likelihood to represent tumor antigens reveals that their aggregated load also correlates with high immune activity in multiple cancers, while also predicting expansion of T-cells in breast cancers receiving ICB and prolonging long-term survival of cancer patients treated with ICB. Collectively, our method provides a framework for analyzing AS in single-cell data and defines a key role for AS in the response to ICB.

overall_design: RNA-seq profiling of widetype MCF-7 cells and their ESRP1 knockdown and overexpression derivatives, as well as widetype BT-549 cells and their NOVA1 knockdown derivatives.

first: Chen

last: Gu

first: Diether

last: Lambrechts


title: Structural and molecular insights into specialized translation mediated by the ribosome mRNA-binding channel [Ribo-Seq]

city: Rehovot

first: Ariel

last: Ogran

address: 234 Herzl Street

department: Bimolecular Sciences

country: Israel

institute: Weizmann Institute of Science

summary: The ribosome mRNA channel is central for translation, but its involvement in specific regulatory mechanisms remains unexplored. Employing Cryo-EM on ribosomal complexes bound to Kozak and TISU mRNAs from wild-type and RPS26 mutant cells (RPS26dC), we demonstrate that RPS26/eS26 adopts distinct conformations, providing a structural basis for its opposing effect on their activity. Translatome studies of wild-type and RPS26dC revealed AUG-context-dependent rearrangements in key intermediates of the 48S and full 80S initiation complexes and slower scanning. Downregulated mRNAs are enriched with AUG upstream nucleotides and a C at position -1 that contacts 18S rRNA nucleotide G1207, a contact lost in the mutant structure. The top downregulated mRNAs are replication-dependent histones, which, despite having short 5’UTRs and suboptimal Kozak, exhibit robust translation activity that is RPS26/eS26-dependent. We identified a translational enhancer in H2B 5’UTR, spanning nucleotide -16 to -9, overlapping the predicted RPS26/eS26-binding sites. Intriguingly, the H2B mRNA-ribosome complex structure adopts a conformation distinct from the Kozak and TISU. Exploiting these features, we designed a high-efficiency translational cassette with minimal leaky scanning, improving efficacy and safety for mRNA-based therapy. These findings underscore the central importance of the ribosome’s mRNA-binding channel in selective translation regulation and its potential for therapeutic applications.

overall_design: Ribo-seq profling of WT human embryonic kidney cells (HEK293T) and RPS26dC HEK293T cells

first: Davide

last: Fraticelli

first: Disha-Gajanan

last: Hiregange

first: Benjamin

last: Weiss

first: Tal

last: Havkin-Solomon

first: Irene

last: Martinez-Roman

first: Anat

last: Bashan

first: Ada

last: Yonath

first: Rivka

last: Dikstein


title: Regulation of murine follicle-stimulating hormone β subunit transcription by newly identified enhancers

first: Michel

last: Zamojski

address: 8700 Beverly Boulevard

summary: Activin-class ligands of the TGFβ family stimulate follicle-stimulating hormone (FSH) synthesis in pituitary gonadotrope cells through SMAD3/SMAD4 and FOXL2-mediated transcriptional regulation of Fshb. This SuperSeries integrates chromatin accessibility and transcription factor binding datasets to identify activin-sensitive regulatory regions controlling Fshb expression. It includes single-nucleus ATAC-seq of mouse pituitaries, and bulk ATAC-seq and ChIP-seq of gonadotrope-derived LβT2 cells. Together, these datasets indicate that there may be as many as four activin-sensitive enhancers upstream of murine Fshb.

overall_design: Mice were engineered with a previously identified Fshb enhancer knocked out.  Single-nucleus ATAC-seq was performed on whole pituitaries from wild-type, and enhancer knockout mice, as well as from IgG- and Bimagrumab-, an activin type II receptor antagonist, treated mice.  Bulk ATAC-seq and ChIP-seq for SMAD2/3, FLAG-FOXL2 and H3K27ac was performed on vehicle- and activin A-treated LβΤ2 cells.

first: Yangfan

last: Jin

first: Hailey

last: Schultz

first: German

last: Nudelman

first: Louise

last: Bilezijkian

first: Stuart

last: Sealfon

first: Frederique

last: Ruf-Zamojski

first: Daniel

last: Bernard


title: Hic-5 drives epithelial mechanotransduction promoting a feed-forward cycle of bronchoconstriction.

city: Boston

first: Adam

last: Haber

state: Massachusetts

address: 665 Huntington Ave, HSPH Building 1, Room 305

institute: Harvard T. H. Chan School of Public Health

summary: Mechanical forces are essential for organ function, but excessive or dysregulated forces can promote pathologic conditions. In asthma, bronchoconstriction narrows the airway, compressing the airway epithelium and activating mechanotransduction, yet key regulators of mechanotransduction remain unclear. Here we show that Hic-5, a focal adhesion adaptor, is a key regulator of epithelial mechanotransduction. In human airway epithelial cells at air–liquid interface exposed to mechanical compression that mimics bronchoconstriction, we find that compression induces Hic-5 expression in airway basal cells. We further validated these in vitro findings by reanalyzing single-cell RNA-seq data from patients with asthma undergoing bronchoconstriction after allergen challenge, which revealed increased Hic-5 expression in airway basal cells. Hic-5 knockdown in human airway epithelial cells markedly attenuates mechanoresponses to compression, including stress fiber formation, differential gene expression, and increased secretion of endothelin-1 (ET-1). Through secretion of ET-1, a potent bronchoconstrictor, Hic-5 drives epithelial mechanotransduction and promotes a feed-forward cycle of bronchoconstriction, thereby highlighting dysregulated mechanical forces as active drivers of human disease.

overall_design: To mimic bronchoconstriction, well-differentiated HBE cells were exposed to apical-to-basal pressure with a magnitude of 30cm H2O for 3 hrs, as previously described (11, 12, 49, 53). Time-matched control cells received 0cm H2O pressure. Cells and conditioned media were collected between 3 and 72 hrs. In experiments using recombinant human (rh) TGF-β1 (Cell Signaling Technologies, Danvers, MA), we spiked 10 ng/ml into the basolateral media of HBE cells in ALI cultures. Cells were collected between 24 and 72 hrs for further analysis. To determine the signaling pathways that regulate mechanical compression-induced Hic-5 protein expression, we used pharmacological inhibitors of the TGF-β receptor (SB431542; 10 µM, Tocris, Bristol, UK), MEK (U0126; 10 µM, Tocris), and EGFR (AG1478; 10 µM, Tocris). Each inhibitor was added to the basolateral media of HBE cells 1 hr prior to exposure to mechanical compression or TGF- β. As a control, 0.1% DMSO was used.To knock down Hic-5 expression in HBE cells, we used antisense oligonucleotides designed and provided by Ionis Pharmaceuticals. We initially tested five ASOs targeting TGFB1I1(denoted as Hic-5 ASO). Then, we  selected the most effective ASO and used for subsequent experiments. HBE cells maintained in ALI culture were treated with 10µM of either a non-targeting control (scrambled) ASO or Hic-5 ASO between ALI days 9 and 20 prior to the application of mechanical compression. We confirmed the knockdown efficiency of the Hic-5 ASO in well-differentiated HBE cells by RT-qPCR and western blot analysis.

first: Chimwemwe

last: Mwase

first: Wenjiang

last: Deng

first: Hyo-Jin

last: Kim

first: Jennifer

last: Mitchel

first: Thien-Khoi

last: Phung

first: Michael

last: O’Sullivan

first: Joel

last: Mathews

first: Jeffrey

last: Crosby

first: Christopher

last: Turner

first: Jin-Ah

last: Park


title: RNA-binding protein PTBP1 mediates HSV-1 infection through splicing regulation of HS 3-O-sulfotransferase genes

city: Nanjing

first: Erguang

last: Li

address: Hankou Road 22, Nanjing, Jiangsu Province, China

department: Medical School

institute: Nanjing University

summary: Polypyrimidine tract-binding protein 1 (PTBP1) is an RNA-binding protein (RBP) that plays an instrumental role in regulating alternative splicing events by manipulating RNA folding. Here we report a novel role of PTBP1 in heparan sulfate (HS) biosynthesis. Specifically, we found that genetic ablation of PTBP1 resulted in loss of HS3ST3A1 and HS3ST3B1 (also known as 3OST3A1 and 3OST3B1, respectively) expression. Both the HS3ST3A1 and HS3ST3B1 genes, which encode heparan sulfate -glucosamine 3-O-sulfotransferases for heparan sulfate modification of proteoglycans (HSPGs), are unique in that each pre-mRNA is composed of 2 exons separated by an extraordinarily long intron. Analyses of global iCLIP-seq and CRIC-seq revealed that the intron region of HS3ST3A1/HS3ST3B1 pre-mRNAs interacted with the PTBP1 dimer, creating contacts to impact splicing events. Ectopic expression of WT, but not a dimerization defective mutant (C23S) of PTBP1, restored functional 3OST3 expression. Cell-surface HSPGs are the most abundant proteoglycans that can act as co-receptors modulating cytokine activities as well as for viruses to attach and to enter cells. We showed that loss of 3OST3 expression markedly reduced HSV-1 infection and viral spreading. This study demonstrates that PTBP1 mediates 3OST3 expression for HS biosynthesis and plays a role in HSV-1 infection.

overall_design: Utilize RNA-seq to comprehensively analyze the effect of PTBP1 konckout and HSV-1 infection on host gene expression in HeLa cells

first: Weikang

last: Sun

first: Mengyu

last: Zhang

first: Renjie

last: Luo

first: Ruilin

last: Wang

first: Jie

last: Yang

first: Zhiqiang

last: Huang

first: Xiangdong

last: Liu

first: Peng

last: Cao


title: Xenium In Situ analysis of mouse skin mock-treated or treated with SspA and HDM

city: Charlestown

first: Jin Mo

address: 149 Thirteenth Street

institute: Massachusetts General Hospital

summary: Treatment of mouse skin with the staphylococcal protease SspA and house dust mite extract (HDM) results in dermatitis. Using a 10x Genomics Xenium In Situ platform, we analyzed spatial features in mock-treated skin and inflamed skin treated with epicutaneous SspA and HDM.

overall_design: Mouse skin sections prepared 7 days after mock or SspA/HDM treatment were subjected to Xenium v1 analysis using the Mouse Tissue Atlassing Panel.

first: Yanek

last: Jiménez-Andrade


title: Unbiased assessment of 2-Arachidonoylglycerol in cardiovascular inflammation and myocardial remodeling

city: Bonn

first: Susanne

middle: Viktoria

last: Schmidt

address: Venusberg Campus 1

institute: Institute of Clinical Chemistry and Clinical Pharmacology

summary: 2-arachidonoylglycerol (2-AG) is an endogenous modulator of inflammatory processes and ligand for the endocannabinoid system. Recent scientific investigations accumulated evidences for its key role in inflammatory cardiovascular diseases, like atherosclerosis and left ventricular remodeling. However, molecular mechanisms set in motion by 2-AG are still poorly understood and its role in cardiovascular inflammation is the matter of controversial debate. Yet, there is a dramatic need to understand local effects of 2-AG at the site of heart inflammation, and tissue injury and reconstruction to develop novel therapeutic strategies which modulate both processes for the better and a fast recovery. Myeloid cells, like tissue-resident macrophages and monocytes are central mediators of inflammatory processes and mediate cardiac adverse remodelling after an insult by myocardial infarction, myocarditis or chronic hypertension. In this study, we investigated the impact of 2-AG on the transcriptional programming of peripheral monocytes as model system for resident macrophages in heart tissue or monocytes which are recruited to the site of cardiac tissue remodelling. Data of RNA sequencing analysis and quantification cytokine expression we seeked to clarify the question, if 2-AG induces rather an inflammatory or a wound healing phenotype of monocytes recruited to the site of cardiac tissue injury.

overall_design: CD14+ primary human monocytes were isolated from peripheral blood of healthy donors and treated with 2-AG up to 24h. Genes differentially expressed between 2-AG treated monocytes and respective controls were determined by mRNA sequencing and used for bioinformatics analysis of changes in myeloid cell functions and cell signaling pathways.

first: Moritz

last: Nöthel

first: Susanne V.

first: Julian

last: Jehle

first: Franziska

last: Dorer

first: Alexandru

last: Odainic

first: Marie

last: Rüthing

first: Georg

last: Nickenig


title: Activated T cells degrade extracellular proteins to enhance effector functions

city: shanghai

first: yunxue

last: yin

address: 825 Zhangheng Road

institute: Fudan unversity

summary: Proteins are the most abundant source of amino acids in body fluids. However, the potential contribution of extracellular protein catabolism to the regulation of T cell immunity remains poorly understood. In this study, we show that endocytosed extracellular proteins function as an amino acid source in activated T cells, maintaining mTORC1 activity and sustaining cytokine production following T cell activation. Genetic ablation of Tfe3 impairs the activation-induced upregulation of lysosomal genes and disrupts extracellular protein catabolism, resulting in attenuated mTORC1 signaling and compromised anti-viral and anti-tumor T cell responses. The TFE3-protein-mTORC1 signaling axis demonstrates clinical relevance. CD8+PD-1+ tumor-infiltrating T cells from older patients with lung cancer display reduced lysosomal degradation capacity and impaired cytokine secretion compared to their middle-aged counterparts. This functional defect is rescued by treatment with Vismodegib, a TFE3-inducing drug. Our findings reveal lysosome-mediated extracellular protein catabolism as an important metabolic pathway supporting T cell immunity.

overall_design: Sorted tumor-derived total CD8+ T cells from three Tfe3fl/fl and three Cd4-Cre+Tfe3fl/fl mice were multiplexed using sample tags to facilitate simultaneous processing.


title: CDK9 is a Dependency in GATA-3 driven and MCL-1 independent T-cell Lymphomas

city: Ann Arbor

first: Chenguang

state: Michigan

address: 1500E Medical Center Dr, Room 4411

department: Department of Internal Medicine

institute: University of Michigan

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

overall_design: Refer to individual Series


title: Single-cell RNA sequencing of MCF-7 cells treated with 17-β estradiol or fulvestrant, and fulvestrant resistant derivatives of MCF-7 cells

city: Darlinghurst

first: Leila

last: Eshraghi

state: NSW

address: 384 Victoria Street

country: Australia

institute: Garvan Institute of Medical Research

summary: Single-cell RNA sequencing (scRNA-seq) on MCF-7 breast cancer cells treated with fulvestrant or 17-β estradiol, and MCF-7 cell line derivatives made resistant to fulvestrant.

overall_design: High throughput single cell RNA sequencing (RNA-seq) using the Drop-seq method was used to analyse single cell transcriptomes of MCF-7 cells that are treated with fulvestrant, and then released from fulvestrant arrest by the addition of 17-β estradiol. Fulvestrant resistant cell lines were also examined.

first: Catherine

last: Caldon


title: Decoding the endometrial landscape at single-cell resolution from implantation preparation to early pregnancy [bulk RNA-seq]

city: Newark

first: Nataki

middle: C.

last: Douglas

state: NJ

address: 185 S Orange Ave, MBS E569

department: Ob-Gyn and Women's Health

institute: Rutgers-New Jersey Medical School

summary: Embryo implantation relies on the development of a receptive endometrium during the secretory phase of the menstrual cycle. Prior studies have attempted to define the window of embryo implantation within the secretory phase based on patterns of differentially expressed genes, but consensus surrounding this signature has not been achieved.  We aim to examine the cell types that characterize the purported receptivity signature and identify new transcriptomic markers that define the mid-secretory endometrium through integrated bulk and single-cell RNA sequencing.

overall_design: Bulk RNA-seq profiling of endometrial biopsies from the proliferative, early secretory, mid secretory, and late secretory menstrual phases.

first: Gregory

last: Burns

first: Emmanuel

middle: N

last: Paul

first: Rong

last: Li

first: Manish

last: Persaud

first: Kristin

last: Blackledge

first: Jessica

last: Garcia de Paredes

first: Qingshi

last: Zhao

first: Ripla

last: Arora

first: Anat

last: Chemerinski


title: Cas9-mediated in vivo miRNA silencing

city: Marseille

first: Eduardo

last: GASCON

address: 27 bd Jean Moulin

country: France

institute: INT Timone

summary: Although the advent of Cas9 technology has expanded our ability to precisely edit the genome, manipulating micro RNAs in vivo has been shown to be particularly challenging, especially in the brain. Here, we sought to generate novel tools aiming at targeting and efficiently down regulating defined microRNAs species in a cell-specific manner so that their function in discrete neuronal networks could be investigated. Focusing on miR-124, a micro RNA highly expressed in the mammalian brain and transcribed from three independent chromosomal loci, we designed and validated different gRNAs directed against this miRNA. In vitro, our Cas9 designs show not only a significant reduction in miR-124 levels but also a functional effect on miR-124 silencing. Similarly, when packed into AAV vectors and injected into the mouse cortex, miR-124-Cas9 vectors strongly down regulate miR-124 levels without affecting the expression of other miRNAs. In parallel, levels of endogenous miR-124 targets exhibit a significant increase supporting the release of its silencing activity. To functionally validate our tools, we provide evidences that deletion of miR-124 in the sub ventricular zone altered migration of newly generated neurons into the olfactory bulb. Finally, we also showed that our vectors also modified the Ca2+ permeability of AMPA receptors, a robust functional output downstream of miR-124. These tools are expected to help elucidating miRNA function in complex experimental settings such as brain networks in vivo.

overall_design: The objective is to identify and quantify mutations introduced by CRISPR/Cas9 in the 3 miR-124 gene loci and the 2 miR-128 loci, as well as possible mutations introduced outside these target genes. AAVs encoding CRISPR/Cas9 were injected into the motor cortex of C57BL/6 WT mice. We tested three guides (inoperative Sa1, Sa4 targeting miR-124 and Sa2 targeting miR-128) and one construct without a guide. After 4 weeks, tissues were collected, dissociated and Cas9-positive cells were sorted by FACS.

last: Gascon

first: David

last: Roura-Martinez

first: Florence

last: Jaouen

first: Cynthia

last: Rombaut

first: Ana

last: Borges

first: Natalia

last: Popa

first: Dipankar

last: Bachar


title: Clonal CD8+ T Cells Populate the Leptomeninges and Coordinate with Immune Cells in Human Degenerative Brain Diseases

city: New York

first: Ryan

last: Hobson

state: New York

institute: Columbia University

summary: Meningeal immune cells monitor the central nervous system (CNS) and influence neuroinflammation in mice, but the human leptomeningeal immune landscape and the changes that occur in this immunological niche in neurodegeneration remain underexplored. We performed single-cell RNA and T cell receptor (TCR) sequencing of 99,625 high-quality immune cells from 57 leptomeninges and brain samples from donors with Alzheimer’s disease (AD), amyotrophic lateral sclerosis, and Parkinson’s disease and found that while the leptomeninges are home to highly clonally expanded CD8 tissue-resident memory (TRM) T cells, the maximal level of clonal expansion was decreased in AD in comparison to non-neurodegenerative controls. Intra-patient paired tissue analysis further revealed that brain and leptomeningeal TCR repertoires share significant similarities, but tissue-specific clones emerge in AD. Finally in AD, the degree of CD8 TRM clonal expansion was positively correlated with microglial TGFB2, suggesting that brain and leptomeningeal immune cells coordinate their activities in AD. In addition to identifying key inflammatory dynamics in the human degenerating CNS, this study establishes a foundational resource for future studies that could inform treatment for AD and other neuroinflammatory diseases.

overall_design: scRNA/TCR sequencing of CD45+ cells isolated from human postmortem leptomeninges and brain

first: Samuel

last: Levy

first: Delaney

last: Flaherty

first: Harrison

last: Xiao

first: Benjamin

last: Ciener

first: Hasini

last: Reddy

first: Nick

last: Zabinyakov

first: Chitra

last: Singal

first: Christine

middle: Y

last: Kim

first: Andrew

last: Teich

first: Neil

middle: A

last: Shneider

first: Elizabeth

middle: M

last: Bradshaw

first: Wassim

last: Elyaman


title: The organization principles of P. falciparum translation initiation features and their potential as drug targets.

city: Petah Tikva

first: Rom

last: Barzilay

address: Harav Unterman 10

department: Biomolecular Sciences

institute: Weizmann Institute of Science, Israel

summary: Plasmodium falciparum, the deadliest malaria-causing parasite, challenges eradication efforts due to drug resistance. To date, mRNA translation remains an unexplored therapeutic target. Most mRNAs contain unusually long 5’UTR and multiple upstream AUGs. How the parasite overcomes scanning distance and uORF constraints is unknown. Using Ribo-seq, TIS-seq, and long-read data, we mapped translation initiation sites, determined translation efficiency, and re-annotated the 5’UTRs. We identified actively translated uORFs in 81% of mRNAs, predominantly initiated by unspecified AUG, while inactive uORFs are enriched with inhibitory AUG contexts. Notably, initiation within CDS through leaky scanning is also highly prevalent. Surprisingly, mRNAs with long 5’UTR and active uORFs exhibit higher translation efficiency compared to those lacking uORFs. This is facilitated by a specific spacing of active uORFs and peptide length, optimizing scanning distance, ribosome density, and reinitiation. Remarkably, one-hour exposure to the eIF1-eIF4G1 inhibitor i14G1-12 reduced parasite cell viability and caused translational repression by enhancing leaky scanning and disturbing the unique arrangement of uORFs. Collectively, our findings uncovered the unusual translation regulatory features of P. falciparum and highlighted the therapeutic potential of targeting these mechanisms

overall_design:
To obtain a genome-wide, quantitative view of translation in P. falciparum, and under treamtent with the eIF1-eIF4G1 inhibitor i14G1-12, we performed Ribosome Profiling on DMSO and i14G1-12 treated cultures of trophozoite-stage parasites.
To assess translation efficiency (TE), we used cycloheximide (CHX), a translation elongation inhibitor that stalls all translating ribosomes. Measurement of total mRNA abundance from the same samples was used for normalization.
Two independent replicates were assigned for each library (CHX, HAR, total RNA) in each treatment group (DMSO, i14G1-12), adding up to a total of 12 samples.

first: Anastasia

last: Lev

first: Ariel

last: Ogran

first: Rivka

last: Dikstein


title: Single cell mapping of the human endometrium and first trimester decidua identifies distinct epithelial and stromal cell contributions to fertility [single cell RNA-seq]

first: Nataki

last: Douglas

overall_design: Single cell transcriptome profiling of endometrial biopsies from the proliferative, early secretory, mid secretory, and late secretory menstrual phases, and decidua from the first trimester of pregnancy.

first: Gregory

last: Burns

first: Emmanuel

last: Paul

first: Manisha

last: Persaud

first: Qingshi

last: Zhao

first: Rong

last: Li

first: Kristin

last: Blackledge

first: Jessica

last: Garcia de Paredes

first: Pratibha

last: Shukla

first: Ripla

last: Arora

first: Anat

last: Chemerinski

middle: C


title: A Workflow for Transcriptome-wide Assessment of Antisense Oligonucleotide Selectivity

city: Carlsbad

first: Sagar

last: Damle

state: CA

address: 2855 Gazelle Ct

institute: Ionis Pharmaceuticals

summary: Antisense oligonucleotides (ASOs) designed to recruit RNase H1 (gapmer ASOs) have been used successfully to downregulate the expression of therapeutic targets. Gapmer ASOs can be identified that selectively reduce the expression of transcripts containing the perfectly complementary intended ASO target site without affecting the expression of unintended transcripts (selective ASOs). However, ASOs can also be identified that reduce the expression of unintended transcripts with target sites that are not perfectly complementary to the ASO (non-selective ASOs). Currently, the understanding of in silico rules for predicting off-targets is suboptimal. In order to determine the selectivity of gapmer ASOs, we therefore developed an experimental workflow called Concentration-Response Digital Gene Expression (CR-DGE). In CR-DGE, ASO treatment is performed at increasing concentrations and the effect on the transcriptome is measured using 3’Tag-Seq. Expression data are then analyzed to identify genes with concentration-responsive knockdown. We demonstrate that CR-DGE identifies ASO concentration-responsive genes with high reproducibility and greater sensitivity than conventional single-concentration assays.  Applying CR-DGE to a panel of gapmer ASOs identifies ASOs with a range of selectivity. These results demonstrate that CR-DGE can be used effectively to assess the selectivity of gapmer ASOs, offering a valuable tool for research and therapeutic development.

overall_design: Concentration-response 3'Tag-Seq data for cultured A431 cells treated with ACTN1 ASO and non-targeting control ASO

first: Andy

last: Watt

first: Steven

last: Kuntz

first: Amanda

last: Crutchfield

first: Emma

last: Carlborg

first: Judy

last: Webb

first: Clare

last: Quirk

first: Dorde

last: Relic

first: Scott

last: Donovan


title: Gene expression and chromatin accessibility in human embryonic stem cells with partial deletion of PDHB during DE differentiation [ATAC-seq]

city: WuHan

first: liming

last: meng

address: No. 115, Donghu Road

institute: WuHan university

summary: We established a PDHB heterozygous cell line in human embryonic stem cell HUES8. Then we performed definitive endoderm (DE) differentiation using both WT and PDHB partial deletion cell lines, and assessed gene expression and chromatin accessibility changes during this process by RNA-seq and ATAC-seq. Differentially expressed genes and regions with altered chromatin accessibility showed that in the PDHB-deleted cell line, the efficiency of endoderm differentiation was significantly inhibited. This alteration primarily occurred through changes in chromatin accessibility at BAF complex binding sites on DNA, which is a major ATP-dependent chromatin remodeling complex. And we also detected the SMAD2/3 binging by ChIP-seq.

overall_design: We performed the DE differentiation in WT and PDHB partial deletion cells. The DE differentiation medium was based on DMEM and supplemented with 100 ng/mL Activin A, 1x penicillin/streptomycin, and 0.2% bovine serum albumin. After the differentiation, we performed the RNA-seq and ATAC-seq in both cells. Besides that, we performed the SMAD2/3 ChIP-seq in WT cells. Detection was carried out after 3 days of induction using the same system.

first: Liming

last: Meng

first: Jie

last: Yang

first: Wei

last: Jiang


title: Structural and molecular insights into specialized translation mediated by the ribosome mRNA-binding channel [RNA-Seq]

first: Ariel

last: Ogran

first: Davide

last: Fraticelli

first: Disha-Gajanan

last: Hiregange

first: Benjamin

last: Weiss

first: Tal

last: Havkin-Solomon

first: Irene

last: Martinez-Roman

last: Bashan

first: Ada

last: Yonath

first: Rivka

last: Dikstein


title: Meningeal-derived retinoic acid regulates neurogenesis via suppression of Notch and Sox2 [ATAC-seq]

city: Aurora

first: Christina

last: Como

state: CO

institute: University of Colorado

summary: The meninges act as a regulator of brain development by secreting ligands that act on neural cells to regulate neurogenesis and neuronal migration. Meningeal-derived retinoic acid (RA) promotes neocortical neural progenitor cell cycle exit; however, the underlying molecular mechanism is unknown. Here, we used spatial transcriptomics and profiling of retinoic-acid receptor-a (RARa) DNA binding in Foxc1-mutant embryos that lack meninges-derived ligands to identify the neurogenic transcriptional mechanisms of RA signaling in telencephalic neural progenitors. We determined that meningeal-derived RA promotes neurogenesis by suppressing progenitor self-renewal pathways Notch signaling and the transcription factor Sox2. We show that RARα binds in the Sox2ot promoter, a long non-coding RNA that regulates Sox2 expression, and RA promotes Sox2ot expression in neocortical progenitors. Our findings elucidate a previously unknown mechanism of how meningeal RA coordinates neocortical development and insight into how defects in meningeal development may cause neurodevelopmental disorders.

overall_design: Bulk ATAC-seq data from Prom1+ magnetically separated E14 forebrain neural progenitors

first: Julie

last: Siegenthaler


title: Transcriptomic profiles of the maternal-fetal interface in wild-type and C1qtnf6-KO mice

city: Yangzhou

first: Yuchen

last: Tang

state: Jiangsu

address: Wenhui Road Yangzhou Citr No.48

institute: College of Veterinary Medicine Yangzhou University

summary: To understand the function of CTRP6 protein during gestation, we performed RNA-seq experiments for the maternal-fetal interface in wild-type and C1qtnf6-KO mice.

overall_design: We collected the maternal-fetal interface samples from wild-type and C1qtnf6-KO mice at four different gestation stages, includinggd10.5, gd14.5, gd16.5 and gd19.5. Three replicates per sample were used for RNA-seq experiments.

first: Hairui

last: Fan

first: Xiaolei

last: Chen

first: Cui

last: Du

first: Ming-an

last: Sun


title: The gene expression of primitive macrophage progenitors derived from Down syndrome model mouse embryonic stem cells on differentiation day 4

city: Kyoto

first: Keiichi

last: Ishihara

state: Kyoto

address: 5 Misasagi Nakauchi-cho, Yanashina-ku

institute: Kyoto Pharmaceutical University

summary: The Ts1Cje mouse model of Down syndrome (DS) has partial triplication of mouse chromosome 16, of which a large portion is homologous to human chromosome 21. We have shown that the number of microglia was decreased in the brain of Ts1Cje fetus, thereby we analyzed the effects of Ts1Cje trisomic region on differentiation of embryonic stem cells (ESCs) with genotype of Ts1Cje mice into primitive macrophage progenitors.

overall_design:
To elucidate the reason why the number of microgila is decreased in Ts1Cje embryos, we induced primitive macrophage progenitors from ESCs established from the Ts1Cje mice.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 3 differnt cells of 2 genotypes (wild-type and Ts1Cje).
Comparative gene expression profiling analysis of RNA-seq data for primitive macrophage progenitors derived from wild-type and Ts1Cje mouse ESCs.

first: Koki

last: Harada


title: Transcriptome analysis of quiescent livers and regenerating livers after partial hepatectomy (PH) in young and aged mice

city: Fukuoka

first: Atsushi

last: Suzuki

state: Fukuoka

address: 3-1-1 Maidashi, Higashi-ku

department: Medical Institute of Bioregulation

institute: Kyushu University

summary: To elucidate the effects of aging and the regeneration process on the liver, we conducted transcriptome analyses on young (11–14 weeks old), old (60 weeks old), and very old (100 weeks old) C57BL/6 mice (Clea Japan) before and after partial hepatectomy (PH), as well as on sham control mice.

overall_design: We also used RNA-seq analysis data with the following conditions for batch correction; RNA-seq analyses for 11-14 weeks old male mouse 52 hours after sham surgery (YM_Sh2d_1_mix18, YM_Sh2d_2_mix18 and YM_Sh2d_3_mix21), 11-14 weeks old male mouse 4 days after sham surgery (YM_Sh4d_1_mix19, YM_Sh4d_2_mix21 and YM_Sh4d_3_mix21), 60 weeks old male mouse 52 hours after sham surgery (OM_Sh2d_1_mix20, OM_Sh2d_2_mix17, OM_Sh2d_3_mix23 and OM_Sh2d_3X_mix17), 60 weeks old male mouse 4 days after sham surgery (OM_Sh4d_1_mix19, OM_Sh4d_2_mix20 and OM_Sh4d_3_mix20), 100 weeks old male mouse 52 hours after sham surgery (VOM_Sh2d_1_mix9, VOM_Sh2d_2_mix9 and VOM_Sh2d_3_mix9), 100 weeks old male mouse 4 days after sham surgery (VOM_Sh4d_1_mix13, VOM_Sh4d_2_mix13, VOM_Sh4d_3_mix23 and VOM_Sh4d_3X_mix17), 11-14 weeks old female mouse that did not undergo surgery (YF_Ctrl_1_mix2, YF_Ctrl_2_mix2, YF_Ctrl_3_mix2 and YF_Ctrl_4_mix18), 11-14 weeks old female mouse 52 hours after PH (YF_52h_1_mix3, YF_52h_2_mix3 and YF_52h_3_mix18), 11-14 weeks old female mouse 4 days after PH (YF_4d_1_mix4, YF_4d_2_mix4 and YF_4d_3_mix19), 11-14 weeks old female mouse 52 hours after sham surgery (YF_Sh2d_1_mix3, YF_Sh2d_1X_mix11, YF_Sh2d_2_mix3, YF_Sh2d_2X_mix10 and YF_Sh2d_3_mix17), 11-14 weeks old female mouse 4 days after sham surgery (YF_Sh4d_1_mix4, YF_Sh4d_1X_mix11, YF_Sh4d_2_mix4, YF_Sh4d_2X_mix10 and YF_Sh4d_3_mix19), 60 weeks old female mouse that did not undergo surgery (OF_Ctrl_1_mix12, OF_Ctrl_1X_mix1, OF_Ctrl_1X_mix10, OF_Ctrl_2_mix12, OF_Ctrl_2X_mix1, OF_Ctrl_3_mix12 and OF_Ctrl_3X_mix1), 60 weeks old female mouse 52 hours after PH (OF_52h_1_mix14, OF_52h_2_mix14 and OF_52h_3_mix19), 60 weeks old female mouse 4 days after PH (OF_4d_1_mix14, OF_4d_2_mix15 and OF_4d_3_mix15), 60 weeks old female mouse 52 hours after sham surgery (OF_Sh2d_1_mix14, OF_Sh2d_2_mix22, OF_Sh2d_2X_mix14 and OF_Sh2d_3_mix21), 60 weeks old female mouse 4 days after sham surgery (OF_Sh4d_1_mix13, OF_Sh4d_2_mix15, OF_Sh4d_3_mix22 and OF_Sh4d_3X_mix15), 100 weeks old female mouse that did not undergo surgery (VOF_Ctrl_1_mix2, VOF_Ctrl_2_mix2 and VOF_Ctrl_3_mix2), 100 weeks old female mouse 52 hours after PH (VOF_52h_1_mix5, VOF_52h_2_mix5 and VOF_52h_3_mix11), 100 weeks old female mouse 4 days after PH (VOF_4d_1_mix6, VOF_4d_2_mix6, VOF_4d_3_mix22 and VOF_4d_3X_mix11), 100 weeks old female mouse 52 hours after sham surgery (VOF_Sh2d_1_mix5, VOF_Sh2d_2_mix5 and VOF_Sh2d_3_mix13) and 100 weeks old female mouse 4 days after sham surgery (VOF_Sh4d_1_mix6, VOF_Sh4d_2_mix6 and VOF_Sh4d_3_mix15).

first: Ryo

last: Murayama

first: Kenichi

last: Horisawa


title: Transcriptome analysis of HUVECs and those transduced with hiHepPC-inducing factors in culture with or without CHIR99021 [RNA-Seq]

summary: We previously showed that a specific combination of three transcription factors (FOXA3, HNF1A, and HNF6) can induce direct reprogramming of human umbilical vein endothelial cells (HUVECs) and peripheral blood-derived endothelial cells into human induced hepatic progenitor cells (hiHepPCs; PMID: 33087715). However, the low induction efficiency of hiHepPCs will be a problem when using them in research and clinical applications. Here, we show that activation of the canonical Wnt signaling pathway increase the reprogramming efficiency of HUVECs to hiHepPCs by rapidly inducing chromatin remodeling and gene expression changes in the transduced HUVECs.

overall_design: hiHepPCs were induced from HUVECs by forced expression of the three transcription factors (FOXA3, HNF1A, and HNF6). Total RNA was extracted from transduced HUVECs 4, 8, 9, 16, and 44 days after retrovirus infection, and transcriptome analyses were performed using a 3’UTR-seq method (PMID: 27121950). Two conditions, with and without CHIR99021, were examined during induction of hiHepPCs from HUVECs. Additionally, the same analysis was conducted for parental HUVECs.

first: Shizuka

last: Miura

first: Hiroki

last: Inada


title: Chromatin state analysis of HUVECs and those transduced with hiHepPC-inducing factors in culture with or without CHIR99021 [ATAC-seq]

overall_design: hiHepPCs were induced from HUVECs by forced expression of the three transcription factors (FOXA3, HNF1A, and HNF6). ATAC-seq was performed 9, 16, and 44 days after retrovirus infection. Two conditions, with and without CHIR99021, were examined during induction of hiHepPCs from HUVECs. Additionally, the same analysis was conducted for parental HUVECs.


title: A Transcriptional Hub for Endochondral ossification adn Fracture Repair.

city: Pamplona

first: Asier

last: Ullate-Agote

state: Navarra

address: Av. de Pío XII, 55

department: Hemato-Oncology

country: Spain

institute: CIMA

summary: After injury, bone tissue initiates a reparative response that will recover its structure and function, failure to initiate or delay this response results in fracture nonunion. The molecular mechanisms underlying fracture nonunion appearance are not yet stablished. We proposed that the signaling pathways derived from hypoxia and mediated by reactive oxygen species (ROS) homeostasis control Bmp2 expression and fracture healing initiation, having potential to induce fracture nonunion when excessive ROS develops into oxidative stress. Here, we silenced Apex1, the final transducer of ROS signaling and responsible of the activation of key transcription factors by oxidoreduction of cysteines, evaluating its role during endochondral ossification and fracture repair. Silencing Apex1 in limb bud mesenchyme results in transient metaphyseal dysplasia derived of impaired chondrocyte differentiation. During bone regeneration, silencing Apex1 generates a fracture nonunion phenotype characterized by delayed fracture healing initiation with impaired periosteal response and reduced chondrocyte and osteoblasts differentiation. In addition, compromised chondrocyte differentiation hampers fracture healing progression and callus vascularization. Our data supports a molecular mechanism of fracture healing initiation dependent of hypoxia derived ROS signaling in mesenchymal progenitors.

overall_design: Bulk RNA-seq for fracture calluses of wild-type and Apex1 KO mice at different days post fracture(-dpf)  3-, 6-, 10- and 14-dpf)

first: Froilán

last: Granero-Moltó

first: Emma

last: Muiños-López

first: José

last: Valdés-Fernández


title: Insights into Intramuscular Adipose-Muscle Signaling in the Diabetic Lower Extremity

city: St. Louis

first: Gretchen

last: Meyer

state: MO

address: 507 S Euclid Ave

department: Department of Physical Therapy

institute: Washington University School of Medicine in Saint Louis

summary: Intramuscular adipose tissue (IMAT) has been proposed to directly contribute to diabetic myofiber dysfunction through paracrine signaling. The impacts of this signaling beyond contributing to myofiber insulin resistance are largely unknown. This study aims to explore the human IMAT transcriptome, with a focus on its potential role in diabetic muscle regenerative deficits. Using a within-subjects design, we compared IMAT to subcutaneous (SQ) fat in individuals with and without diabetes. We hypothesized that diabetic IMAT would exhibit an inflammatory profile, similar to diabetic SQ, and that inflammatory secreted factors from IMAT progenitors would impair cultured myoblast fusion. Instead, we found that the diabetic IMAT transcriptome exhibited reduced enrichment of inflammatory pathways compared with SQ and less transcriptional evidence for immune cell infiltration. While diabetic IMAT featured a mostly anti-myogenic transcriptional profile for secreted cytokines, media conditioned by diabetic IMAT progenitors did not uniquely impair fusion of cultured myoblasts compared with non-diabetic IMAT or SQ. Surprisingly, the diabetic status of the myoblast donor predicted myoblast fusion, with reduced fusion rates in diabetic myoblasts exposed to conditioned media from all adipose sources. This suggests that IMAT-myoblast signaling may be detrimental to regeneration in diabetes, but that the effect is driven in part by changes in myoblast sensitivity to IMAT secreted factors, rather than altered IMAT secretome alone. This emphasizes the insight that can be gained from disease-state matched and mismatched culture models and highlights the need to better understand how diabetes impacts myoblasts and their interaction with the disease environment.

first: Chang

last: Gui

first: Dakota

last: Kamm

first: Jeremie

last: Ferey

first: Kathryn

last: Bohnert

first: Jeremy

last: McCormick

first: Mary

last: Hastings