title: Dual targeting of processive transcription for Myc-driven circuitry in medulloblastoma [RNA-seq]

city: Denver

first: Pluto

last: Bio

address: 3575 Ringsby Ct Suite 414

institute: Pluto Bio

summary: Medulloblastoma is the most common malignant brain tumor of childhood. The highest-risk tumors are driven by recurrent Myc amplifications (Myc-MB) and experience poorer outcomes despite intensive multimodal therapy. The Myc transcription factor defines core regulatory circuitry for these tumors and acts to broadly amplify downstream pro-survival transcriptional programs. Therapeutic targeting of Myc directly has proven elusive, but inhibiting transcriptional cofactors may present an indirect means of drugging the oncogenic transcriptional circuitry sustaining Myc-MB. Independent CRISPR-Cas9 screens were pooled to identify conserved dependencies in Myc-MB. We performed chromatin conformation capture (Hi-C) from primary patient Myc-MB samples to map enhancer-promoter interactions. We then treated in vitro and xenograft models with the dual CDK9/7 inhibitor zotiraciclib to evaluate effect on Myc-driven programs and tumor growth. Eight CRISPR-Cas9 screens performed across three independent labs identify CDK9 as a conserved dependency in Myc-MB. Myc-MB cells are susceptible to CDK9 inhibition, which is synergistic with concurrent inhibition of CDK7. The dual CDK9/7 inhibitor zotiraciclib disrupts enhancer-promoter activity in Myc-MB and downregulates Myc-driven transcriptional programs, exerting potent anti-tumor effect.

overall_design:
Myc-amplified D458 or D425 cell lines were established in culture and then treated with either atuveciclib, TG02, or equivalent DMSO control. We then performed gene expression profiling from RNA-seq at the indicated timepoints.

UPDATE: [Jan-11-2025] The titles and treatment information were updated for Samples GSM8267116-GSM8267127.

first: Nathan

last: Dahl

first: Rajeev

last: Vibhakar

first: Lays

last: Martin Sobral


title: Single cell transcriptomic analyses of the dynamic local and systemic response to cardiac injury in mice and zebrafish [zebrafish]

status: Public on Nov 07 2024

city: New York

first: Paul

last: Zumbo

address: 1300 York Ave

department: Applied Bioinformatics Core

country: USA

institute: Weill Cornell Medicine

type: Expression profiling by high throughput sequencing

summary: Mice respond to myocardial infarction (MI) by generating a fibrotic scar in the heart. This is accompanied by systemic inflammation after the MI resolves. In contrast, zebrafish can fully regenerate their heart after heart injury. To further investigate the divergent species’ responses to cardiac injury, single cell transcriptomic analyses were performed on the heart, blood, liver, kidney, and pancreatic islets.

overall_design: Upon heart cryoinjury, cells from multiple organs (heart, liver, pancreas, and whole kidney marrow) were isolated for scRNA-seq at 1 and 7 days post injury together with the uninjured controls.

first: Yao

last: Jun

first: Xia

last: Yu

first: Cao

last: Jingli


title: Single cell transcriptomic analyses of the dynamic local and systemic response to cardiac injury in mice and zebrafish

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

sample_organism:
Danio rerio
Mus musculus

platform_organism:
Danio rerio
Mus musculus

overall_design: Refer to individual Series


title: Single cell transcriptomic analyses of the dynamic local and systemic response to cardiac injury in mice and zebrafish [mouse]

overall_design: Upon LAD ligation or sham surgery, cells from multiple tissues (heart, liver, pancreatic islets, kidney and blood) were isolated for scRNA-seq at 1, 7 and 30 days post infarction

first: Eric

last: Cortada

first: Boris

last: Yang

first: James

middle: C

last: Lo


title: Concomitant loss of TET2 and TET3 results in T cell expansion and genomic instability [BGI-RNA-Seq]

first: Tarmo

last: Äijö

address: 162 5th Avenue

department: Center for Computational Biology

institute: Flatiron Institute

summary: TET proteins mediate DNA demethylation by oxidizing 5-methylcytosine to 5-hydroxymethylcytosine (5hmC) and other oxidative derivatives. While we have previously demonstrated a dynamic enrichment of 5hmC during T cell lineage specification, the impact of TET proteins in CD4 cell lineage differentiation remains largely unknown. Here, we discover that TET proteins exert a fundamental role in regulating the differentiation and maturation of the CD4 lineage by controlling the expression of the lineage specifying factor Th-POK, which is encoded by Zbtb7b.  We demonstrate that TET proteins mediate DNA demethylation of a proximal enhancer, critical for the stability of Th-POK expression. In addition, TET proteins drive the DNA demethylation of site A at the Zbtb7b locus to facilitate GATA3 binding, an event critical for Th-POK expression and subsequent CD4 maturation.  The impaired maturation of Tet2/3 DKO CD4 cells leads to the acquisition of a stemness signature and results in acquisition of malignant traits.

overall_design: We compared gene expression (by bulk RNA-seq) in peripheral  CD4 cells isolated from control (wild type), Tet2/3 DKO mice, as well as from CD45.1+ recipients of CD45.2+ Tet2/3 DKO CD4 cells after one or two rounds of Tet2/3 DKO CD4 cell transplantation

first: Ageliki

last: Tsagaratou


title: EAE_CD3_Sp/Br

status: Public on Nov 06 2024

city: Suita

first: Daisuke

last: Motooka

state: Osaka

address: 3-1, Yamadaoka

country: Japan

institute: NGS core facility, Research Institute for Microbial Diseases, Osaka University

summary: Experimental autoimmune encephalomyelitis (EAE) is the most widely used rodent model for multiple sclerosis. Interferon-γ (IFN-γ) and regulatory T cells (Tregs) are individually well known to play beneficial roles in amelioration of EAE. However, little is known about the relationship between IFN-γ and Tregs during the disease. Here we show that IFN-γ polarizes Tregs into Th1-type Tregs (Th1-Tregs) to recover from EAE. Single-cell RNA sequencing analysis revealed that brain Tregs showed signs of IFN-γ stimulation during EAE. Loss of IFN-γ signaling in Tregs and of T cell-derived IFN-γ impaired the Th1-Treg polarization and worsened the disease. Moreover, selective ablation of Th1-Tregs using an intersectional genetic method promoted pro-inflammatory features of macrophages in the inflamed brains and exacerbated the EAE. Taken together, our study highlights a critical role of T cell-derived IFN-γ for Th1-Treg polarization in inflamed brain to ameliorate EAE.

overall_design: EAE was induced into WT C57BL/6N mice. CD3+ cells were isolated from the brain and spleen of the same EAE-induced mice at peak of disease, and single cell RNA-seq was performed.

first: Masaaki

last: Okamoto

last: Okuzaki

first: Masahiro

last: Yamamoto


title: Concomitant loss of TET2 and TET3 results in T cell expansion and genomic instability

type: Genome variation profiling by high throughput sequencing


title: Human immunodeficiency virus-1 targets genomic R-loops of the host for integration [IS-seq]

status: Public on Nov 05 2024

city: Seoul

first: Dohoon

last: Lee

address: 1, Gwanak-ro, Gwanak-gu

department: Bioinformatics Institute

country: South Korea

institute: Seoul National University

summary: Although HIV-1 integration sites are considered to favor active transcription units in the human genome, high-resolution analysis of individual HIV-1 integration sites have shown that the virus can integrate in a variety of host genomic locations, including non-genic regions, challenging the traditional understanding of HIV-1 integration site selection. Here, we showed that HIV-1 targets R-loops, a genomic structure made up of DNA–RNA hybrids, for integration. HIV-1 initiates the formation of R-loops in both genic and non-genic regions of the host genome and preferentially integrates into regions of HIV-1-induced R-loops. Using a cell model that can independently control transcriptional activity and R-loop formation, we demonstrated that the presence of R-loops, regardless of transcriptional activity, directs HIV-1 integration targeting sites. We also found that HIV-1 integrase proteins bind to the host genomic R-loops. These findings provide fundamental insights into the mechanisms of retroviral integration and the new strategies of antiretroviral therapy against HIV-1 latent infection.

overall_design: HeLa, CD4+ and Jurkat cells were infected with VSV-G-pseudotyped HIV-1-EGFP and harvested at 5 d post infection (dpi) for HIV-1 integration site sequencing

first: Kiwon

last: Park

first: Sun

last: Kim

first: Kwangseog

last: Ahn


title: EAE_CD4IFNGcKO_CD3

overall_design: EAE was induced into Ifng fl/fl and CD4-Cre/ Ifng fl/fl mice. CD3+ cells were isolated from the brain of the  EAE-induced mice at peak of disease, and single cell RNA-seq was performed.


title: Human immunodeficiency virus-1 targets genomic R-loops of the host for integration [RNA-seq]

overall_design: HeLa cells were infected with VSV-G-pseudotyped HIV-1-EGFP and harvested at 0, 3, 6, and 12 h post infection (hpi) for RNA-seq library construction


title: Intrinsically Disordered Regions Steer the Function Coordination of the Traveling Chromatin Modifier During Pol II Elongation (ChIP-Seq)

status: Public on Nov 13 2024

city: Shanghai

first: Bing

last: Li

address: South Chongqing Road 280

department: Department of Biochemistry and Molecular Cell Biology

country: China

institute: Shanghai Jiao Tong University School of medicine

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The Rpd3S histone deacetylase complex suppresses cryptic initiation and maintains genomic integrity by deacetylating transcribed nucleosomes subsequent to RNA Pol II passage. Recent cryo-EM structure analysis of Rpd3S emphasized the significance of two asymmetrical Rco1/Eaf3 dimers in nucleosome binding. However, the mechanism by which Rpd3S interacts with nucleosomal substrates while traveling with elongating Pol II remains elusive. In this study, we elucidate the crucial role of the Rco1 N-terminal Intrinsically Disordered Region (IDR) in regulating Pol II association. Mutations within the K/R cluster of Rco1 IDR significantly impede Rpd3S binding to CTD, while leaving the overall complex integrity and nucleosome recognition unaffected. Moreover, we identify the Rco1-PHD1 and Eaf3-CHD domains as the minimal module responsible for specific interaction with Ser5-phosphorylated CTD. The Rco1 IDR functions by relieving autoinhibition from its C-terminal regions, enabling the binding of PHD-CHD to CTD. Additionally, we uncover an evolutionarily conserved mechanism governing the distinct roles of the asymmetrical Rco1/Eaf3 dimers, which coordinate nucleosome engagement and Pol II binding. These findings offer valuable new perspectives on the functional dynamics of mobile epigenetic modifying complexes that accompany processive machinery.

sample_organism:
Schizosaccharomyces pombe
Saccharomyces cerevisiae

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for H4 acetylation in wild-type and 13K/R>A mutant yeast strains.

first: Yixuan

last: Pan

first: Meiyang

last: Liu

first: Qi

last: Zhang


title: Human immunodeficiency virus-1 targets genomic R-loops of the host for integration [DRIPc]

overall_design: HeLa, CD4+ and Jurkat cells were infected with VSV-G-pseudotyped HIV-1-EGFP and harvested at 0, 3, 6, and 12 h post infection (hpi) for DRIPc-seq library construction


title: Targeting Ketone Body Metabolism in Mitigating Gemcitabine Resistance

status: Public on Jan 02 2025

city: Los Angeles

first: Krizia

last: Rohena Rivera

state: California

address: 8750 Beverly Blvd

institute: Cedars Sinai Medical Center

summary: Chemotherapy is often combined with surgery for muscle invasive and non-muscle invasive bladder cancer. However, 70% of the patients recur within 5 years. Metabolic reprogramming is an emerging hallmark in cancer chemoresistance. Here, we report a gemcitabine resistance mechanism which promotes cancer reprogramming via the metabolic enzyme, OXCT1. This mitochondrial enzyme, responsible for the rate-limiting step in β-hydroxybutyrate (βHB) catabolism, was elevated in muscle invasive disease and in chemo-resistant bladder cancer patients. Resistant orthotopic tumors presented an OXCT1-dependent rise in mitochondrial oxygen consumption rate, ATP, and nucleotide biosynthesis. In resistant bladder cancer, knocking out OXCT1 restored gemcitabine sensitivity, and administering the non-metabolizable βHB, enantiomer (S-βHB) only partially restored gemcitabine sensitivity. Suggesting an extra-metabolic role for OXCT1, multi-omics analysis of gemcitabine sensitive and resistant cells revealed an OXCT1-dependent signature with the transcriptional repressor, OVOL1, as a master regulator of epithelial differentiation. The elevation of OVOL1 target genes was associated with its cytoplasmic translocation and poor prognosis in a chemotherapy-treated BCa patient cohort. The knockout of OXCT1 restored OVOL1 transcriptional repressive activity by its nuclear translocation. Orthotopic mouse models of bladder cancer supported OXCT1 as a mediator of gemcitabine sensitivity through ketone metabolism and regulating cancer stem cell differentiation.

overall_design: Three independent clones of 5637 parental  and 5637 gemcitabine resistant cells were analyzed for gene expression differences associated with gemcitabine resistance.

last: Rohena-Rivera

first: Sungyong

last: You

first: Minhyung

first: Sandrine

last: Billet

first: Johanna

last: ten Hoeve

first: Gabrielle

last: Gonzales

first: Chengqun

last: Huang

first: Ashley

last: Heard

first: Keith

last: Chan

first: Neil

middle: A

last: Bhowmick


title: Loss of XIST lncRNA promotes stemness and cellular plasticity in ovarian cancer

city: Irvine

first: Sha

last: Sun

state: CA

address: 4232 McGaugh Hall

department: Development and cell biology

institute: University of california Irvine

summary: Cancer cell plasticity is an emerging hallmark of the disease where cells can transition into different cellular states, contributing to tumor heterogeneity. Cellular plasticity is linked to cancer progression, treatment resistance, and relapse. Cancer Stem Cells (CSCs) play an important role in cancer cell plasticity, and molecular factors contributing to stemness maintenance are still poorly understood. Here, we investigate the role of XIST (X-inactive specific transcript) long noncoding RNA (lncRNA) in ovarian cancer cellular plasticity. We perform in silico and in vitro analyses to better understand its function. Our findings demonstrate a significant downregulation of XIST in ovarian tumors.  cancer patients.Tumors with low XIST expression have higher stemness index and decreased overall survival. This reduction is associated with an increased stemness in tumors with XIST  index and decreased overall survival rates. XIST knockdown (KD) in ovarian cancer cell lines increases stem cell population. Transcriptomic data show that low XIST expression correlates with the upregulation of the stem cell-specific pathways MYC and MTOR, as well as the metabolic pathway OXPHO. Our study also demonstrates that XIST KD increases specifically mesenchymal cancer stem cell (MCSC) population. Finally, ovarian cancer cells with XIST KD show higher plasticity under hypoxic conditions by transitioning into Epithelial Cancer Stem CellsCell (ECSC). Our data reveal that XIST loss induces CSC enrichment, leading to higher cellular plasticity. These findings highlight the important role of XIST in protecting cell identity in ovarian cancer and open the door to new therapeutic approaches for patients with low XIST expression

overall_design: To better understand the function of XIST lncRNA in ovarian cancer, we generate OVCAR3 cell lines that stably express a sgRNA targeting XIST. We also look how OVCAR3 behave in hypoxic enviromnent af XIST knockdown. We generate transcriptomic data  by RNA-Sequencing of two diffenrents sgRNA again XIST after and before hypoxic treatment.

first: Ikrame

last: Naciri

first: Minzhi

last: Liang

first: Ying

last: Yang

first: Heather

last: Karner

first: Benjamin

last: Lin

first: Maria

last: De Lourdes Andrade Ludena

first: Eric

last: Hanse

first: Alfredo

last: Lebron

first: Olga

middle: V

last: Razorenova

first: Dequina

last: Nicholas

first: Mei

last: Kong


title: Non-classical action of Ku70 promotes Treg suppressive function through a FOXP3-dependent mechanism in lung adenocarcinoma

status: Public on Nov 04 2024

first: Qianru

address: 227 South Chongqing Road Shanghai

institute: Shanghai Jiao Tong University School of Medicine

summary: To explore the role of Ku70 in Treg cell biology, we isolated conventional T cells (Tconv) and  regulatory T cells (Treg) from Foxp3Yfp-cre mice and performed Ku70 ChIP-seq. Furthermore, to investigate the impact of Ku70 deficiency on Foxp3 binding patterns, we sorted Treg cells from Foxp3Cre (WT) and Xrcc6fl/fl;Foxp3Cre (KO) mice and conducted Foxp3 ChIP-seq. These analyses revealed that Ku70 forms a complex with FOXP3, which supports FOXP3 transcriptional activity.

overall_design: Using chromatin immunoprecipitation DNA-sequencing (ChIP-seq), we performed a comparative analysis of Ku70 binding peaks in Treg and Tconv cells in mice, as well as examined the impact of Ku70 deficiency on Foxp3 binding sites.


title: Regulatory T cells crosstalk with tumor and endothelium through lymphotoxin signaling

city: Baltimore

first: yang

last: song

state: MD

address: HSF-3, 3rd Floor; 670 W Baltimore St.

department: Department: Institute for Genome Sciences

institute: University of Maryland in Baltimore

summary:
Tregs with multifaceted functions suppress anti-tumor immunity by signaling surrounding cells. Tregs use the surface LTa1b2 to preferentially stimulate LTbR nonclassical NFkB signaling on both tumor and LECs to accelerate tumor growth and metastasis. Selectively targeting LTβR nonclassical NFkB pathways on both tumors and LECs cocultured with Tregs, inhibits tumor growth and migration in vitro. Leveraging in vivo Treg LTa1b2 interactions with LTβR on tumor and LECs, transfer of WT but not LTa-/- Tregs promotes  B16F10 melanoma growth and tumor cell-derived chemokines in LTβR-/- mice; and increases SOX18 and FLRT2 in lymphatic vessels of LTbR-/-melanoma. Blocking the nonclassical pathways suppresses tumor growth and lymphatic metastasis by reducing chemokine production, restricting Treg and MDSC recruitment to tumors, and retaining intratumoral IFNg+ CD8 T cells. Our data reveals that Treg LTa1b2 promotes LTbR nonclassical NFkB signaling in tumor cells and LECs providing a rational strategy to prevent Treg promoted tumor growth and metastasis.
Best regards

overall_design: 10 groups of total RNA sample from B16F10 melanoma with differential treatment. First 5 sets: WT B16F10 pretreated with classical LTbR-NFkB blocking peptide ciLT (20uM) for 30 min, wash and stimulated with anti-LTbR agonist Ab (3C8) for 1 hour, CRISPR-Cas9 IKKb KO B16F10 used as a control;  2nd 5 sets: WT B16F10 pretreat with  nonclassical NFkB blocking peptide nciLT (20uM) for 30 min,wash then stimulated with 3C8 for 6 hours. CRISPR-Cas9 NIK KO used as a control.

first: Wenji

last: Piao

first: Yanbao

last: Xiong

first: Lushen

last: Li

first: Jonathan

middle: S

last: Bromberg


title: Molecular logic for cellular specializations that initiate the auditory parallel processing pathways

city: Houston

first: Junzhan

last: Jing

state: Texas

address: 1250 Moursund St., Suite 0925

department: Neuroscience

institute: Baylor College of Medicine

summary: The cochlear nuclear complex (CN), the starting point for all central auditory processing, comprises a suite of neuronal cell types that are highly specialized for neural coding of acoustic signals, yet molecular logic governing cellular specializations remains largely unknown. By combining single-nucleus RNA sequencing and Patch-seq analysis, we reveal a set of transcriptionally distinct cell populations encompassing all previously observed types and discover multiple new subtypes with anatomical and physiological identity. The resulting complete cell-type taxonomy reconciles anatomical position, morphological, physiological, and molecular criteria, enabling determination of molecular basis of the remarkable cellular phenotypes in the CN. In particular, CN cell-type identity is encoded in a transcriptional architecture that orchestrates functionally congruent expression across a small set of gene families to customize projection patterns, input-output synaptic communication, and biophysical features required for encoding distinct aspects of acoustic signals. This high-resolution account of cellular heterogeneity from the molecular to the circuit level illustrates molecular logic for cellular specializations and enables genetic dissection of auditory processing and hearing disorders with unprecedented specificity.

overall_design: Here we employed a massively parallel 10X genomics single nucleus RNA-sequencing (snRNA-seq) analysis of the CN to derive a molecular classification and then related each molecularly distinct cell type to well-established CN cell types via Patch-seq.

first: Xiaolong

last: Jiang


title: Identification of LRP1 and CD13 expressing human periosteal cells for stem cell-mediated bone repair

first: Dongsu

last: Park

state: TX

address: One Baylor Plaza

department: Molecular and Human Genetics

summary: Human periosteal skeletal stem cells (P-SSCs) are critical for cortical bone maintenance and repair. However, their in vivo identity, molecular characteristics, and specific markers remain unknown. Here, single-cell sequencing revealed human periosteum contains SSC clusters  expressing known SSC markers, PDPN and PDGFRA. Notably, human P-SSCs, but not bone marrow SSCs (BM-SSCs), selectively expressed newly identified markers, LRP1 and CD13. Single-cell analysis of mouse periosteum further supported the preferential expression of LRP1 and CD13 in Prx1 P-SSCs. These LRP1CD13 human P-SSCs were perivascular cells with high osteochondrogenic but minimal adipogenic potential. In addition, LRP1CD13 human P-SSCs are maintained in vitro and self-renew in vivo upon transplantation into bone injuries in mice. When Lrp1 was conditionally deleted in Prx1-lineage cells, it led to severe bone deformity, short statue, and periosteal defects. By contrast, local treatment with a LRP1 agonist at the injury sites induced early P-SSC proliferation and bone healing. Thus, human periosteum contains unique osteochondrogenic stem cell subsets, and these P-SSCs express specific markers, LRP1 and CD13, with regulatory mechanism through LRP1 that enhances P-SSC function and bone repair.

overall_design: Non-hematopoietic and non-endothelial cells from mouse long bone periosteum were isolated by Fluorescence-activated cell sorting and analyzed by 10x scRNA-seq

first: Youngjae

last: Jeong


title: Spatiotemporal functions of leukemia inhibitory factor in embryo attachment and implantation chamber formation

city: Bunkyo-ku

first: Shizu

last: Aikawa

state: Tokyo

address: 7-3-1 Hongo

institute: The University of Tokyo

summary: RNA-seq analyses using embryos and uterine luminal layers collected from Lif-floxed, Lif uterine epithelial-specific KO (eKO), and recombinant Lif (rLif)-treated Lif eKO females on 8pm of day 4 of pregnancy.

overall_design: Lif-loxP/loxP, Ltf-iCre, and Pgr-Cre mice were used in this study. Lactoferrin (Ltf) is expressed in the uterine epithelium, whereas progesterone receptor (Pgr) is expressed in the entire uterus (i.e., epithelium, stroma, and myometrium). Lif-loxP/loxP females were crossed with Ltf-iCre and Pgr-Cre males to generate mice with deletion of Lif in the epithelium (Lif eKO mice) or the whole uterus (Lif uKO mice), respectively. The day when the vaginal plug was detected was considered day 1 of pregnancy. Pregnant mice were euthanized by cervical dislocation on designated days of pregnancy for the evaluation of pregnancy phenotypes and sample collection. On day 4, one uterine horn was flushed with saline to confirm the presence of blastocysts. To determine the function of Lif, female mice received rLif (20 µg/head, i.p.) at 10 am and 6 pm of day 4 of pregnancy or pseudopregnancy. For RNA-seq analyses, mice were dissected 2 h after the last injection. Uterine tissues were collected from Lif-floxed, Lif eKO, and rLif-treated Lif eKO mice at 8 pm on day 4. One side of the uterine horn was flushed with PBS to collect blastocysts and confirm pregnancy. The other side of the uterine horn was snap-frozen for cryosectioning. Laser microdissection was performed to collect luminal layers.

first: Takehiro

last: Hiraoka

first: Yasushi

last: Hirota


title: Regulatory T cells and IFN-γ-producing Th1 cells play a critical role in the pathogenesis of Sjögren’s Syndrome

first: Maxwell

last: McDermott

state: NY

address: 550 1st Ave

department: Pathology

institute: NYU Langone Health

summary: Sjogren’s Disease (SjD) is an autoimmune disorder characterized by progressive salivary and lacrimal gland dysfunction, inflammation, and destruction, and extra glandular manifestations. The etiology and pathophysiology of SjD are incompletely understood. Regulatory T (Treg) cells are critical for immunological tolerance and although Treg abnormalities have been implicated in the pathogenesis of several autoimmune diseases, their role in SjD remains ambiguous.  We here show that Stim1fl/flStim2fl/flFoxp3-Cre (Stim1/Stim2 Foxp3cre) mice develop a SjD-like phenotype, including severe sialadenitis with hyposalivation, lacrimal gland (LG) inflammation with dry eye and corneal damage, increased anti-Ro and anti-La autoantibody levels and interstitial lung disease. At the cellular level, salivary gland inflammation in Stim1/2Foxp3 mice is characterized by infiltration with CD4+ and CD8+ T cells, which is dominated by a T helper (Th) 1 and Th2 immune response. The changes in gene expression in SGs of mice correlate strongly with transcriptional and epigenetic dysregulation found in SjD patients. Our findings in Stim1/Stim2 Foxp3cre mice provide new evidence for a critical role of Treg cells and IFNγ producing Th1 cells in mediating SjD progression.

overall_design: Whole submandibular glands (SMG) of unstimulated Stim1/Stim2 Foxp3cre and Cre negative littermate control mice were isolated. Differential gene expression between the two conditions was determined by bulk RNA sequencing.

first: Yin-Hu

last: Wang

first: Li

last: Wenyi

first: Stefan

last: Feske


title: Bacterial single-cell RNA sequencing captures biofilm transcriptional heterogeneity and differential responses to immune pressure

city: Omaha

first: Lee

middle: Erik

last: Korshoj

state: NE

address: 985900 Nebraska Medical Center

department: Pathology, Microbiology, and Immunology

institute: University of Nebraska Medical Center

summary: Biofilm formation is an important mechanism of survival and persistence for many bacterial pathogens. These multicellular communities contain subpopulations of cells that display vast metabolic and transcriptional diversity along with high recalcitrance to antibiotics and host immune defenses. Investigating the complex heterogeneity within biofilm has been hindered by the lack of a sensitive and high-throughput method to assess stochastic transcriptional activity and regulation between bacterial subpopulations, which requires single-cell resolution. We have developed an optimized bacterial single-cell RNA sequencing method, BaSSSh-seq, to study Staphylococcus aureus diversity during biofilm growth and transcriptional adaptations following immune cell exposure.

overall_design: BaSSSh-seq was first applied to compare transcriptional profiles of S. aureus biofilm and planktonic growth. BaSSSh-seq was next applied to S. aureus biofilms following direct co-culture with the major leukocyte infiltrates associated with biofilm infection: macrophages, granulocytic myeloid-derived supressor cells, and neutrophils.

middle: E

first: Tammy

last: Kielian


title: Effect of depletion of Ku70 on gene expression profiles of  murine regulatory T cells in lung tumor microevnronment

first: Qianru

last: Huang

summary: The functions of the DNA repair protein Ku70 in immune homeostasis and tumor immunity remain unexplored. Here, we conducted deep sequencing of mRNA profiles from tumor-infiltrating Treg cells (from Foxp3Cre and Xrcc6fl/fl;Foxp3Cre mice injected with Lewis lung carcinoma cells). By comparing gene expression profiles, we found that Ku70-absent Treg displayed activated pathways in response to inflammatory factors and upregulated expression of genes negatively regulated by Foxp3. These findings strongly indicate that Ku70 plays a supportive role in enhancing Foxp3 transcriptional activity.

overall_design: To investigate the role of Ku70 in the regulating of Treg function, we generated Treg cell-specific knockout mice (Xrcc6flox/floxFoxp3Yfp-Cre, cKO) and Foxp3Yfp-Cre (control, WT) mice. To establish lung metastasis model, LLC cells (1 million per mouse) were tail-intravenously injecting into 8- to 10-week-old WT and cKO mice. Tumors were excised at 21 days, and Tumor-infiltrating Treg cells were isolated for RNA sequencing.


title: Chimeric protein EWS::FLI1 drives cell proliferation in Ewing Sarcoma via aberrant expression of KCNN1/SK1 and dysregulation of calcium signaling

city: Nantes

first: Anaïs

last: Postec

address: 1 Rue Gaston Veil

country: France

institute: Nantes University

summary: Ewing sarcoma (ES) is characterized by EWS::FLI1 or EWS::ERG fusion proteins. Knowing that ion channels are involved in tumorigenesis, this work aimed to study the involvement of the KCNN1 gene, which encodes the SK1 potassium channel, in ES development. Bioinformatics analyses from databases were used to study KCNN1 expression in patients and cell lines. Molecular approaches and in vitro assays were used to study the transcriptional regulation of KCNN1 and its involvement in the regulation of ES cell proliferation. KCNN1 is overexpressed in ES patient biopsies, and its expression is inversely correlated with patient survival. EWS::FLI1, like EWS::ERG, promotes KCNN1 and SK1 expression, binding to GGAA microsatellites near the promoter of KCNN1 isoforms. KCNN1 is involved in the regulation of ES cell proliferation, with its silencing being associated with a slowing of the cell cycle, and its expression modulates membrane potential and therefore calcium flux. These results highlight that KCNN1 is a direct target of EWS::FLI1 and EWS::ERG and demonstrate that KCNN1 is involved in the regulation of intracellular calcium activity and ES cell proliferation, making it a promising therapeutic target in ES.

overall_design: Estimated transcripts expression between several Ewing Sarcoma cell lines (A673, EW3, EW24, MHHES1, RDES, SKES1, TC71) and several Osteosarcoma cell lines (KHOS, G292, MG63, HOS-MNNG, U2OS, SAOS2).

first: Maryne

last: Dupuy

first: Maxime

last: Gueguinou

first: Régis

last: Brion

first: Robel

last: Tesfaye

first: Mathilde

last: Mullard

first: Laura

last: Regnier

first: Jérôme

last: Amiaud

first: Clémence

last: Hubsch

first: Marie

last: Potier-Cartereau

first: Aurélie

last: Chantôme

first: Bénédicte

last: Brounais-Le Royer

first: Marc

last: Baud’huin

first: Steven

last: Georges

first: François

last: Lamoureux

first: Benjamin

last: Ory

first: Natacha

last: Entz-Werlé

first: Olivier

last: Delattre

first: Françoise

last: Rédini

first: Christophe

last: Vandier

first: Franck

last: Verrecchia


title: Screening and molecular mechanism research on bile microRNAs associated with chemotherapy efficacy in perihilar cholangiocarcinoma

status: Public on Nov 02 2024

city: Beijing

first: Haizhen

last: Du

state: Beijing

address: No.52 Fucheng Road, Haidian District

institute: Peking University Cancer Hospital and Institute

summary: The efficacy of hepatic arterial infusion chemotherapy (HAIC) with oxaliplatin (OXA) and 5-fluorouracil (5-Fu) for treating advanced perihilar cholangiocarcinoma (pCCA) has been demonstrated, yet the survival benefits of HAIC for pCCA patients vary. Here, we aimed to screen out HAIC resistance-related bile microRNAs (miRNAs) and explore the functions of specific bile miRNAs in pCCA based on high-throughput sequencing. Levels of bile miR-532-3p,miR-1250-5p, and miR-4772-5p were related to the survival of advanced pCCA patients after HAIC. However, only overexpression of miR-532-3p promoted OXA/5-Fu resistance, and downregulation of its expression improved sensitivity to OXA/5-Fu. Mechanistic investigations revealed secreted protein acidic and rich in cysteine (SPARC) as the direct target of miR-532-3p. Our study reveals that bile miR-532-3p, miR-1250-5p, and miR-4772-5p may serve as survival biomarkers in advanced pCCA patients after HAIC and that bile miR-532-3p promotes resistance to HAIC with OXA and 5-Fu via negatively regulating SPARC expression.

overall_design: A retrospective review of the Hospital Information System identified a total of 31 advanced pCCA patients who received first-line HAIC treatment from June 2015 to June 2016 at our center, of which bile samples were obtained from 18 patients prior to HAIC treatment. According to the complete survival follow-up, all of the patients died, with the following survival distribution: OS < 6 months, n = 7; OS between 6 months and 12 months, n = 4; and OS > 12 months, n = 7. We ultimately selected the bile samples of four patients among the seven with better prognosis (OS > 12 months; group A) and four patients among the seven with dismal prognosis (OS < 6 months; group B) from the biobank according to the patients’ baseline characteristics and results of RNA quality control. The bile samples (15 mL) were obtained by percutaneous transhepatic cholangial drainage (PTCD) within 7 days before the first cycle of the HAIC procedure. Centrifugation of the fresh bile at 2,000 rpm for 10 minutes yielded a supernatant, which was then stored in a freezer at −80°C. Patients signed informed consent forms for clinical specimen collection prior to PTCD. The sequencing of mRNAs was performed by CloudSeq Biotech Inc. (Shanghai, China). After removing rRNA, a NEBNext® Ultra™ II Directional RNA Library Prep Kit (New England Biolabs, Inc., MA, USA) was used to construct RNA libraries. High-throughput sequencing was performed using an Illumina HiSeq instrument. Differentially expressed mRNAs were filtered by |Fold change| ≥ 2.0 and P value < 0.05. Finally, Gene Ontology (GO) and pathway enrichment analyses were also performed for differentially expressed mRNAs.


title: Tumor stage-driven disruption of NK cell maturation and cytotoxic functions in the lung tumor microenvironment

status: Public on Nov 01 2024

city: Paris

first: Jules

last: Russick

address: 1 avenue Claude Vellefaux

institute: Commissariat à l'Energie Atomique

summary: Natural Killer (NK) cells play a pivotal role against cancer, both by direct killing of malignant cells and by promoting adaptive immune response though cytokine and chemokine secretion. In the lung tumor microenvironment (TME), NK cells are scarce and dysfunctional, but the underlying mechanisms are still unclear. By conducting single-cell transcriptomic analysis of lung tumors, and exploring pseudotime, we uncovered that the TME disrupts the maturation trajectory of NK cells in a tumor stage-dependent manner, ultimately resulting in the selective exclusion of the cytotoxic subset. Using functional assays, we observed intratumoral NK cell death and a reduction in cytotoxic capacities depending the tumor stage. Finally, our analyses of human public dataset on lung cancer corroborate these findings, revealing a parallel dysfunctional maturation process of NK cells during tumor progression. These results highlight novel mechanisms by which tumor cells escape from NK cell cytotoxicity, therefore paving the way for tailored therapeutic strategies.

overall_design:
Mice with small or large tumors (see Material and Methods) were sacrificed. Tumoral, and non-tumoral counterpart, parts of the lungs were analyzed in this single-cell experiment

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Submitter states that missing raw files are due to file loss.
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first: Carine

last: Torset

first: Dan

last: Sun

first: Solenne

last: Marmier

first: Nicolas

last: Merle

first: Elodie

last: Voilin

first: Nathalie

last: Josseaume

first: Maxime

last: Meylan

first: Isaïas

last: Hernandez

first: Pierre-Emmanuel

last: Foy

first: Marco

last: Alifano

first: Audrey

last: Lupo

first: Sophie

last: Siberil

first: Niklas

middle: K

last: Björkström

first: Diane

last: Damotte

first: Isabelle

last: Cremer


title: Decoding Aging in the Heart via Single Cell Dual Omics of Non-Cardiomyocytes

city: Chapel Hill

first: Yiran

last: Song

state: NC

address: 111 Mason Farm Road

institute: The University of North Carolina

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

overall_design: Refer to individual Series

first: Li

last: Wang


title: Connecting Transcriptomics with Computational Modeling to Reveal Developmental Adaptations in Pediatric Human Atrial Tissue

city: Washington

first: Nikki

last: Posnack

state: DC

address: 111 Michigan Avenue, NW

institute: Children's National Research Institute

summary: Nearly 1% of babies are born with congenital heart disease (CHD) – many of whom will require heart surgery within the first few years of life. A detailed understanding of cardiac maturation can help to expand our knowledge on cardiac diseases that develop during gestation, identify age-appropriate drug therapies, and inform clinical care decisions related to surgical repair and postoperative management. Yet, to date, our knowledge of the temporal changes that cardiomyocytes undergo during postnatal development is limited. In this study, we collected right atrial tissue samples from pediatric patients (n=117) undergoing heart surgery. Patients were stratified into five age groups. We measured age-dependent adaptations in cardiac gene expression, and used computational modeling to simulate action potential and calcium transients. Enrichment of differentially expressed genes (DEGs) revealed age-dependent changes in several key biological processes (e.g., cell cycle, structural organization), cardiac ion channels, and calcium handling genes. Gene-associated changes in ionic currents exhibited age-dependent trends, with changes in calcium handling (INCX) and repolarization (IK1) most strongly associated with an age-dependent decrease in the action potential plateau potential and increase in triangulation, respectively. We observed a shift in repolarization reserve, with lower IKr expression in younger patients, a finding potentially tied to an increased amplitude of IKs that could be triggered by elevated sympathetic activation in pediatric patients. Collectively, this study provides valuable insights into age-dependent changes in human cardiac gene expression and electrophysiology, shedding light on molecular mechanisms underlying cardiac maturation and function throughout development.

overall_design: Gene expression compared between (5) age groups: neonate, infant, toddler, school-age, adolescent/young adult (considered baseline). FDR<0.05 cutoff


title: Type III interferons induce pyroptosis in gut epithelial cells and impair tissue repair upon intestinal injury.

city: Boston

first: Ivan

last: Zanoni

address: 1 Blackfan Circle

institute: Boston Children's Hospital and Harvard Medical School

summary: Tissue damage and repair are hallmarks of the inflammatory process. Despite a wealth of information on the mechanisms that govern tissue damage, mechanistic insight on how inflammatory mediators affect repair is lacking. Here, we investigated how interferons influence tissue repair after damage to the intestinal mucosa driven by inflammation or genotoxicity. We found that type III, but not type I or II, interferons delay epithelial cell regeneration by inducing the upregulation of Z-DNA-binding protein 1 (ZBP1). Z-nucleic acids formed during the damage and repair process are then sensed by ZBP-1, leading to Caspase-8 activation, and cleavage of Gasdermin C (GSDMC). Cleaved GSDMC drives epithelial cell death by pyroptosis and delays the re-epithelialization of the large or small intestine after colitis or irradiation, respectively. We also found that the type III interferon/ZBP1/Caspase-8/GSDMC pathway is activated in patients with inflammatory bowel disease (IBD). Our findings highlight a molecular signaling cascade initiated by type III interferons that delays intestinal tissue repair, which has important implications for IBD patients or individuals exposed to radiation therapies.

overall_design:
The goal of this study was to identify differentially expressed genes in the colonic mucosa between IBD patients with an acute flare and compare it to uninflamed healthy controls. We performed IonTorrent sequencing from RNA isolated from colonic tissue samples derived from inflamed tissue samples from IBD patients or uninflamed tissue samples from healthy controls.

***************************************************************
Submitter cannot make the raw sequencing data publicly available due to patient privacy concerns.
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first: Kautilya

last: Jena

first: Julien

last: Mambu

first: Daniel

last: Boehmer

first: Benedetta

last: Sposito

first: Virginie

last: Millet

first: Joshua

last: de Sousa Casal

first: Hayley

middle: I

last: Muendlein

first: Roberto

last: Spreafico

first: Romain

last: Fenouil

first: Lionel

last: Spinelli

first: Sarah

last: Wurbel

first: Chloé

last: Riquier

first: Franck

last: Galland

first: Philippe

last: Naquet

last: Chasson

first: Megan

last: Elkins

first: Vanessa

last: Mitsialis

first: Natália

last: Ketelut-Carneiro

first: Katlynn

last: Bugda Gwilt

first: Jay

middle: R

last: Thiagarajah

first: Hai-Bin

last: Ruan

first: Zhaoyu

last: Lin

first: Egil

last: Lien

first: Feng

last: Shao

first: Janet

last: Chou

first: Alexander

last: Poltorak

first: Jose

last: Ordovas-Montanes

first: Katherine

last: Fitzgerald

first: Scott

middle: B

last: Snapper

first: Achille

last: Broggi


first: Ivan

last: Zanoni

state: MA

address: 300 Longwood Ave

institute: Boston Children's Hospital

summary: Tissue damage and repair are hallmarks of inflammation. Despite a wealth of information on the mechanisms that govern tissue damage, mechanistic insight on how inflammation affects repair is lacking. Here, we investigated how interferons influence tissue repair after damage to the intestinal mucosa. We found that type III, not type I or II, interferons delay epithelial cell regeneration by inducing the upregulation of Z-DNA-binding protein 1 (ZBP1). Z-nucleic acids formed following intestinal damage are sensed by ZBP1, leading to Caspase-8 activation, and cleavage of Gasdermin C (GSDMC). Cleaved GSDMC drives epithelial cell death by pyroptosis and delays repair of the large or small intestine after colitis or irradiation, respectively. The type III interferon/ZBP1/Caspase-8/GSDMC axis is also active in patients with inflammatory bowel disease (IBD). Our findings highlight the capacity of type III interferons to delay gut repair, which has important implications for IBD patients or individuals exposed to radiation therapies.

overall_design: Age-matched (8–12-weeks old) cohorts of male and female mice were given 2.5% DSS drinking water for 7 days, followed by 3 days of recovery. On day 10, three mice per condition and genotype were sacrificed and  colons were harvested. Single-cell RNA-sequencing was performed using 10x Genomics 3' v3.1 transcriptomics kit with mouse replicates (per tissue region) combined together and labeled with biotinylated antibodies, followed by oligo-conjugated streptavidin to differentiate between samples (using Biolegend TotalSeqB reagents).

first: Kautilya

last: Jena

first: Daniel

last: Boehmer

first: Joshua

last: de Sousa Casal


title: Regulatory T cells crosstalk with tumor and endothelium through lymphotoxin signaling [array]

first: yang

last: song

summary: Tregs with multifaceted functions suppress anti-tumor immunity by signaling surrounding cells. Tregs use the surface LTa1b2 to preferentially stimulate LTbR nonclassical NFkB signaling on both tumor and LECs to accelerate tumor growth and metastasis. Selectively targeting LTβR nonclassical NFkB pathways on both tumors and LECs cocultured with Tregs, inhibits tumor growth and migration in vitro. Leveraging in vivo Treg LTa1b2 interactions with LTβR on tumor and LECs, transfer of WT but not LTa-/- Tregs promotes  B16F10 melanoma growth and tumor cell-derived chemokines in LTβR-/- mice; and increases SOX18 and FLRT2 in lymphatic vessels of LTbR-/-melanoma. Blocking the nonclassical pathways suppresses tumor growth and lymphatic metastasis by reducing chemokine production, restricting Treg and MDSC recruitment to tumors, and retaining intratumoral IFNg+ CD8 T cells. Our data reveals that Treg LTa1b2 promotes LTbR nonclassical NFkB signaling in tumor cells and LECs providing a rational strategy to prevent Treg promoted tumor growth and metastasis.

overall_design: primary mouse lymphatic endothelial cells( mLECs) pretreated with or without  peptides nciLT or ciLT were activated with LTbR agonist Ab (3C8) with crosslinking (CL)  for total RNA extraction and hybridization on Affymetrix microarrays

first: Wenji

last: Piao

first: Jonathan

last: Bromberg


title: Transposable element activity captures human pluripotent cell states.

first: Florencia

last: Levin

address: 1 Baylor Plaza


title: Transposable element activity captures human pluripotent cell states [ATAC-seq]

summary: Human pluripotent stem cells (hPSCs) serve as powerful in vitro models to elucidate the molecular underpinnings of embryonic cell fate transitions. hPSCs can be maintained in two distinct states: a naïve state, corresponding to the pre-implantation epiblast, and a primed state, mirroring the post-implantation epiblast. Our research demonstrates that transposable elements act as sensitive indicators of these pluripotency states. We engineered hPSCs with fluorescent reporters that capture the temporal expression dynamics of two transposable elements, LTR5_Hs and MER51B. This dual reporter system facilitates real-time monitoring and isolation of stem cells as they transition from naïve to primed pluripotency and further towards differentiation. Unexpectedly, we identified a rare, metastable cell population within primed hPSCs, marked by transcripts associated with pre-implantation embryo development and triggered by DNA damage. Additionally, our system uncovered novel transcriptional regulators involved in pluripotency, naïve reprogramming, and differentiation. Our study provides key insights into the dynamic regulation of transposable elements during embryonic development and introduces a novel system for investigating and exploiting cellular plasticity.

overall_design: Primed hESC were sorted for GFP+ and GFP+/RFP+ expression and sequenced. Two replicates were sequenced for each sample.

first: Yingzhi

last: Cui

first: Alison

last: Swearingen


title: Whole genome methylome and transcriptome study of tree shrew brains [BiSulfite-seq]

city: Santa Barbara

first: Soojin

last: Yi

address: UC Santa Barbara

department: EEMB

institute: University of California, Santa Barbara

type: Methylation profiling by high throughput sequencing

summary: To understand the regulatory impact of DNA methylation on regulation of gene expression and X chromosome, we examined genome-wide DNA methylationand gene expression from female and male tree shrew brains.

overall_design: Whole genome bisulfite sequencing and RNA sequencing of Tupaia belangeri: adult male and female tree shrew prefrontal cortex

first: Lei

last: Shi

first: Yifan

last: Kong

first: Dongmin

last: Son

first: Yulian

last: Tan

first: Ting

last: Hu


title: Transposable element activity captures human pluripotent cell states [RNA-seq]

overall_design: hESC were differentiated into each of the conditions below, and the GFP+, RFP+ and/or GFP+/RFP+ cells were sorted and sequenced. Two replicates were sequenced for each sample.  For the differentiated conditions a control sample of undifferentiated cells was sequenced alongside.


title: Bisulfite Incorporation Hindered Ligation-based Method for Locus-Specific Detection and Quantification of Pseudouridine

city: Chicago

first: Chuan

last: He

state: IL

address: 929 E 57th St

department: Chemistry Department

institute: University of Chicago

summary: We applied BIHIND-seq to 15 sites of Hela 18S rRNA to probe Pseudouridine modification at the target sites and checked how pseudouridine level changed after DKC1 knockdown.

overall_design: DNA probes were annealed to Hela 18S rRNA with or without DKC1 KD. The annealed product was treated with Bst 2.0 and SplintR ligase. The final ligated product was subjected to PCR amplification for library preparation.

first: Yutao

last: Zhao


title: Isolation of psychedelic responsive neurons underlying anxiolytic behavioral states

city: Davis

first: Jessie

last: Muir

address: 1544 Newton Court

department: Center for Neuroscience

institute: UC Davis

summary: Psychedelics hold promising potential as alternate treatments for neuropsychiatric disorders. However, the neural mechanisms by which they drive adaptive behavioral effects remain unclear. We isolated the specific neurons modulated by a psychedelic to determine their role in driving behavior. Using a light- and calcium-dependent activity integrator, we genetically tagged psychedelic-responsive neurons in the medial prefrontal cortex (mPFC) of mice. Single-nucleus RNA sequencing revealed that the psychedelic drove network-level activation of multiple cell-types, beyond those expressing 5-hydroxytryptamine 2A receptors. We labeled psychedelic-responsive mPFC neurons with an excitatory channelrhodopsin to enable their targeted manipulation. We found that reactivation of these cells recapitulated the anxiolytic effects of the psychedelic, without driving its hallucinogenic-like effects. These findings reveal essential insight into the cell-type specific mechanisms underlying psychedelic-induced behavioral states.

overall_design: Prefrontal cortex of C57BL/6J male and female mice expressing scFLARE2 was miscodissected following24 hours after a psychedelic (DOI) or saline injection.

first: Christina

last: Kim


title: SPRR1A is a potential therapeutic target for osteosarcoma:  in vitro and in vivo evaluations using novel generated artificial osteosarcoma cancer stem cell-like cells

city: Kobe

first: Tomohiro

last: Miyamoto

state: Hyogo-ken

address: Kusunokicho Chuo-ku 7-5-2

institute: Kobe University

summary: Osteosarcoma (OS) is the most prevalent primary malignant bone tumor.We have recently succeeded in generating CSC-like cells (MG-OKS) from the OS cell line  MG-63 by transducing defined factors. A significant increase was observed in  small proline-rich protein 1A (SPRR1A) expression, a cross-linked envelope protein  in keratinocytes, in MG-OKS cells. This study aimed to evaluate the role of SPRR1A in OS using novel MG-OKS cells. MG-OKS cells were transfected with SPRR1A siRNA (siMG-OKS) to suppress SPRR1A expression selectively; MG-OKS and scrambled siRNA-transfected MG-OKS (scMG-OKS) cells served as controls.  Our date that genes involved in cell adhesion were suppressed in the siMG-OKS group, suggesting that suppression of cell adhesion may cause phenotypic changes.

overall_design:
To evaluate the role of SPRR1A in OS using novel MG-OKS cells, we transfected SPRR1A siRNA in MG-OKS to suppress SPRR1A expression selectively. MG-OKS and scrambled siRNA-transfected MG-OKS (scMG-OKS) cells served as controls.
we then performed gene expression profiling analysis using data obtained from RNA-seq of 3 group cells.

first: Shuichi

last: Fujiwara

first: Naomasa

last: Fukase


title: Common and Specific Gene Regulatory Programs in Zebrafish Caudal Fin Regeneration at Single-Cell Resolution

city: St. Louis

last: Wang

state: Missouri

address: 4515 McKinley Avenue, Room 5121

department: Genetics

institute: Washington University in St. Louis School of Medicine

summary: Following amputation, zebrafish regenerate their injured caudal fin through lineage-restricted reprogramming. Although previous studies have charted various genetic and epigenetic dimensions of this process, the intricate gene regulatory programs shared by or unique to different regenerating cell types remain under-investigated. Here, we mapped the regulatory landscape of fin regeneration by applying paired snRNA-seq and snATAC-seq on uninjured and regenerating fin. This map delineates the regulatory dynamics of predominant cell populations at multiple stages of regeneration. We observed a marked increase in the accessibility of chromatin regions associated with regenerative and developmental processes at 1 dpa, followed by a gradual closure across major cell types at later stages. This pattern is distinct from that of transcriptomic dynamics which is characterized by several waves of gene upregulation and downregulation. We identified and in vivo validated cell type-specific and position-specific regeneration-responsive enhancers, and constructed regulatory networks by cell type and stage. Our single-cell resolution transcriptomic and chromatin accessibility map across regenerative stages provides new insights into regeneration regulatory mechanisms and serves as a valuable resource for the community.

overall_design: Fin tissue regenerating from adult zebrafish was collected at 1, 2, 4, and 6 days post-amputation (dpa). For the pre-injury (0 dpa) sample, we collected two segments of fin rays from each fish after removing the distal half of their caudal fin. Each time point had two biological replicates. Nuclei were isolated from each fin tissue sample. Libraries were then constructed from these nuclei following the instructions of the Chromium Single Cell Multiome ATAC + Gene Expression (10x Genomics) kit and were subsequently sequenced on NovaSeq 6000 (Illumina). Raw reads were processed by Cell Ranger ARC v2.0.0 (10x Genomics) with default parameters for read tagging, alignment to the zebrafish reference genome (GRCz11), and feature counting based on Ensembl release 104 (cellranger-arc count).

first: Yujie

last: Chen

first: Yiran

last: Hou

first: Qinglin

last: Zeng

first: Irene

first: Mary

last: Shang

first: Kwangdeok

last: Shin

first: Christopher

last: Hemauer

first: Xiaoyun

last: Xing

first: Junsu

last: Kang

first: Guoyan


title: Loss of TREM2 diminishes CAA despite an overall increase of amyloid load in Tg-SwDI mice

city: Minneapolis

first: Ling

last: Li

email: lil@umn.edu, linglilab@umn.edu

state: MN

address: 2001 6th Street SE Rm 4-208

department: Department of Experimental and Clinical Pharmacology

institute: University of Minnesota

summary: Our study aimed to investigate the impact of germlineTrem2knockout in Tg-SwDI mice, a transgenic mouse model of cerebral amyloid angiopathy and Alzheimer's disease.

overall_design: snRNA-seq was performed to explore the transcriptomic changes followingTrem2knockoutin Tg-SwDI. Six mice from two genotype groups(n=3 SwDI/TWT, n=3 SwDI/TKO) were used for snRNA-seq. Sequencing was performedusing Illumina NovaSeq 6000 with a sequencing depth of ~40,000 reads/nucleus. The estimated number of nuclei sequenced from individual samples ranges from 5700 to 8100.

first: Rui

last: Zhong

first: Yingzheng

last: Xu

first: Jesse

last: Williams


title: CRISPR-screening identifies mechanisms of resistance to KRASG12C and SHP2 inhibitor combinations in non-small cell lung cancer

city: Basel

first: Grainne

last: Kerr

address: Fabrikstrasse 1

country: Switzerland

institute: Novartis

summary: Although KRASG12C inhibitors show clinical activity in patients with KRAS G12C mutated NSCLC and other solid tumor malignancies, response is limited by multiple mechanisms of resistance. The KRASG12C inhibitor JDQ443 shows enhanced preclinical antitumor activity combined with the SHP2 inhibitor TNO155, and the combination is currently under clinical evaluation. To identify rational combination strategies that could help overcome or prevent some types of resistance, we evaluated the duration of tumor responses to JDQ443 ± TNO155, alone or combined with the PI3Kα inhibitor alpelisib and/or the CDK4/6 inhibitor ribociclib, in xenograft models derived from a KRASG12C-mutant NSCLC line and investigated the genetic mechanisms associated with loss of response to combined KRASG12C/SHP2 inhibition.

overall_design: CRISPR sequencing after treatment with KRAS inhibitor, SHP2 inhibitor or combination.

first: Anirudh

last: Prahallad

first: Andreas

last: Weiss

first: Saskia

middle: M


title: Gene expression profile at single cell level of blood CD4 T and B cells from patients after kidney transplantation

first: Kevin

last: Louis

institute: Inserm U976, Paris Cité University

summary: CD4 T and B cells are considered pivotal in driving immune responses in the context of allograft rejection. However, CD4 T and B cell transcriptional profiles according to type of allograft rejection is poorly characterized. We used scRNAseq to profile blood CD4 T and B cells in antibody-mediated rejection (ABMR), T cell mediated rejection (TCMR) and patients without rejection (STABLE).

overall_design: We used scRNAseq to profile blood Fluorescence-activated cell sorting (FACS) sorted CD4 T and B cells from 4 patients with antibody mediated rejection (ABMR), 4 patients with T cell mediated rejection (TCMR) and 4 STABLE patients.

first: Diana

last: Metes


title: Cardiopharyngeal Mesoderm specification into cardiac and skeletal muscle lineages in gastruloids

city: Marseille

first: fabienne

last: lescroart

address: 27 bvd Jean Moulin

department: MMG

institute: Aix-Marseille Univ

summary: Cardiopharyngeal mesoderm contributes to the formation of the heart and head muscles. However, the mechanisms governing cardiopharyngeal mesoderm specification remain unclear. Indeed, there is a lack of an in vitro model replicating the differentiation of both heart and head muscles to study these mechanisms. Such models are required to allow live-imaging and high throughput genetic and drug screening. Here, we show that the formation of self-organizing or pseudo-embryos from mouse embryonic stem cells (mESCs), also called gastruloids, reproduces cardiopharyngeal mesoderm specification towards cardiac and skeletal muscle lineages. By conducting a comprehensive temporal analysis of cardiopharyngeal mesoderm establishment and differentiation in gastruloids and comparing it to mouse embryos, we present the first evidence for skeletal myogenesis in gastruloids. By inferring lineage trajectories from the gastruloids single-cell transcriptomic data, we further suggest that heart and head muscles formed in gastruloids derive from cardiopharyngeal mesoderm progenitors. We identify different subpopulations of cardiomyocytes and skeletal muscles, which most likely correspond to different states of myogenesis with “head-like” and “trunk-like” skeletal myoblasts. These findings unveil the potential of mESC-derived gastruloids to undergo specification into both cardiac and skeletal muscle lineages, allowing the investigation of the mechanisms of cardiopharyngeal mesoderm differentiation in development and how this could be affected in congenital diseases.

overall_design: Gastruloids's cells were dissociated and processed on the 10XGenomics controller and analyzed using scRNAseq.

first: Laurent

last: Argiro

first: Céline

last: Chevalier

first: Caroline

last: Choquet

first: Nitya

last: Nandkishore

first: Adeline

last: Ghata

first: Anaïs

last: Baudot

first: Stéphane

last: Zaffran

first: Fabienne

last: Lescroart


title: Whole genome methylome and transcriptome study of tree shrew brains [RNA-seq]

first: Soojin

last: Yi

first: Lei

last: Shi

first: Yifan

last: Kong

first: Dongmin

last: Son

first: Yulian

last: Tan

first: Ting

last: Hu


title: Rhizopus stolonifer displays necrotrophic behavior when infecting fruit in postharvest

first: Barbara

last: Blanco-Ulate

address: One Shields Ave

department: Plant Sciences

institute: University of California Davis

summary: Soft rot or Rhizopus rot, caused by the fungal pathogen Rhizopus stolonifer, is an aggressive postharvest disease that affects many fruit and vegetables. We proposed that R. stolonifer displays a necrotrophic behavior when infecting fruit, actively killing the host tissues to complete its life cycle. We tested this hypothesis by identifying R. stolonifer infection strategies when interacting with four fruit hosts (tomato, grape, strawberry, and plum). First, we generated a complete and highly contiguous genome assembly for R. stolonifer using PacBio sequencing, of 45.02 Mb in size, an N50 of 2.87Mb, and 12,644 predicted loci with protein-coding genes. We then performed a transcriptomic analysis to identify genes preferentially used by R. stolonifer when growing in fruit versus culture media, and then classified these host-related genes into clusters according to their expression patterns across four time points. Based on the expression data, we determined that R. stolonifer deploys infection mechanisms characteristic of necrotrophs, including a suite of oxidases, proteases, and cell wall degrading enzymes, when it is actively breaking down tissues of all four fruit hosts. Better understanding R. stolonifer – fruit host interactions can support better diagnostic tools and efficient management strategies in postharvest.

sample_organism:
Fragaria x ananassa
Prunus domestica
Solanum lycopersicum
Rhizopus stolonifer
Vitis vinifera

overall_design: Comparative transcriptomic analysis of R. stolonifer infecting tomato, grape, strawberry, and plum at 6, 12, 24, and 48 hours post inoculation.

first: Saskia

middle: D

last: Mesquida-Pesci

first: Abraham

last: Morales-Cruz

first: Silvia

last: Rodríguez-Pires

first: Rosa

last: Figueroa-Balderas

first: Christian

middle: J

last: Silva

first: Adrián

last: Sbodio

first: Elia

last: Gutiérrez-Baeza

first: Petros

last: Raygoza

first: Dario

last: Cantù


title: Impact of hyaluronic acid on cutaneous T-cell lymphoma microenvironment: A novel anti-tumor mechanism of bexarotene

city: Sendai

first: Tetsuya

last: Ikawa

state: Miyagi

address: 1-1 Seiryo, Aoba-ku

institute: Tohoku University

summary: Cutaneous T-cell lymphoma (CTCL) is a type of non-Hodgkin lymphoma that primarily affects the skin rich in hyaluronic acid（HA）. HA composes extracellular matrix in the dermis, which likely affects the development of CTCL but is poorly understood. Here, we show that low molecular weight HA (LMWHA) possibly exacerbates CTCL in mouse and human specimens, and bexarotene already being used in CTCL treatment can decrease HA production. Patients’ serum and skin sections showed higher HA expression than healthy controls. HA extracted from the skin of mice inoculated with tumors showed an increase in LMWHA. LMWHA increased lymphoma cell proliferation in vitro and accelerated tumor formation in mice in vivo. LMWHA also created a favorable environment for tumor cells by influencing fibroblasts, vascular endothelial cells, and tumor-associated macrophages. Together, increased HA, mainly LMW aids in CLCL progression by affecting tumor cells and microenvironment. Next, bexarotene treatment reduced the amount of total HA in murine tumor-inoculated skin and the supernatant of cultured normal human dermal fibroblasts (NHDFs) and HuT78 cells. Detailed in vitro assessments showed that bexarotene treatment decreased hyaluronic acid synthase (HAS) 1 and HAS2 expression in NHDFs and HAS1, HAS3, and CEMIP in HuT78. Chromatin immunoprecipitation assay revealed bexarotene reduced retinoid X receptor-alpha binding to the HAS1 and HAS2 promoters in NHDFs. Bexarotene potentially exerts its anti-tumor effect by reducing HA through decreased expression of HA synthase. These findings provide us with new insights into the developmental process of CLCT and additional insight into bexarotene treatment.

overall_design: To assess the effects of bexarotene on hyaluronic acid-related molecules in vivo, we created lymphoma cell-inoculated mice. Bexarotene was administrated intraperitoneally after tumor formation. The back skin including tumor tissue teated with bexarotene was collected and performed bulk RNA-seq. Then, the expression of  hyaluronic acid generating and degrading molecules were examined.

first: Taku

last: Fujimura


title: The glucocorticoid receptor potentiates aldosterone-induced transcription by the mineralocorticoid receptor

city: Bethesda

first: Thomas

last: Johnson

state: Maryland

address: 41 Medlars Dr. Room B307

department: CCR

institute: National Cancer Institute

summary: We show that GR co-expression alters MR genome-wide binding and transcriptional activity in a locus and ligand specific way.

overall_design: Performed ChIP Sequencing, RNA Sequencing, and Single Molecule Tracking Data.

first: Gregory

last: Fettweis

first: Kaustubh

last: Wagh

first: Brian

last: Almeida-Prieto

first: Manan

last: Krishnamurthy

first: Arpita

last: Upadhyaya

first: Gordon

middle: L

last: Hager

first: Diego

last: de la Rosa

first: Fermin

last: Sánchez de Medina

first: Olga

last: Martinez-Augustin


title: Non-canonical Telomerase Reverse Transcriptase Controls Osteogenic Reprogramming of Aortic Valve Cells Through STAT5 [CUT&Run]

status: Public on Oct 31 2024

city: Pittsburgh

first: Rolando

last: Cuevas

state: PA

address: 200 LOTHROP ST

institute: University of Pittsburgh

summary: Background: Calcific aortic valve disease (CAVD) is the pathological remodeling of valve leaflets. The initial steps in the osteogenic reprogramming of the leaflet are not fully understood. Studies have shown that TERT overexpression primes mesenchymal stem cells to differentiate into osteoblasts, and we investigated whether TERT contributes to the osteogenic reprogramming of valve interstitial cells (VICs). Methods: Human control and CAVD aortic valve leaflets and patient-specific hVICs were used in in vivo and in vitro calcification assays. Loss of function experiments in hVICs and cells isolated from Tert-/- and Terc-/- mice were used for mechanistic studies. In silico modeling, proximity ligation and co-immunoprecipitation assays defined novel TERT interacting partners. Chromatin immunoprecipitation and CUT&TAG sequencing defined protein-DNA interactions. Results: TERT protein was highly expressed in calcified valve leaflets without changes in telomere length, DNA damage, or senescence markers, and these features were retained in isolated primary hVICs. TERT expression increased with osteogenic stimuli, while knock-down or genetic deletion of TERT prevented calcification. Mechanistically, TERT was required to initiate osteogenic reprogramming independent of its canonical telomere-extending activity and the lncRNA TERC. TERT’s osteogenic functions via binding with Signal Transducer and Activator of Transcription 5 (STAT5). Depletion and inhibition of STAT5 prevented calcification. STAT5 was found to bind the promoter region of Runt-Related Transcription Factor 2 (RUNX2), the master regulator of osteogenic reprogramming. Finally, we demonstrate that TERT and STAT5 are upregulated and colocalized in CAVD tissue. Conclusions: TERT's non-canonical activity is required to initiate VIC calcification. TERT partners with STAT5 to bind the RUNX2 gene promoter. These data identify a novel mechanism and potential therapeutic target to decrease vascular calcification.

overall_design: Cleavage Under Targets & Release Using Nuclease (CUT&RUN) coupled with DNA sequencing for the Histone marker H3K4me3 in human valve interstitial cells after seven days of osteogenic treatment in vitro.

first: Cynthia

last: St. Hilaire


title: miRNA expression profiles in the dorsal striatum of a mouse model of nicotine withdrawal

city: Seoul

first: BAEKSUN

last: KIM

state: Seoul

address: 75-10, Ohyeon-ro 6-gil, Gangbuk-gu

institute: Korea Institute of Science and Technology

type: Non-coding RNA profiling by high throughput sequencing

summary: Nicotine withdrawal can diversely affect brain gene expression patterns. In the brain, the dorsal striatum is a hub of nicotine dependence. Previous studies have shown that nicotine dependence leads to functional alterations in the dorsal striatum. miRNAs are small non-coding RNAs that regulates cellular functions and dysfunctions. Here, we performed small RNA sequencing from the dorsal striatum of mice at 7 days after nicotine withdrawal. Our results show that nicotine withdrawal does not notably impact miRNA expression profile in the dorsal striatum, but a few miRNAs were significantly altered in response to nicotine withdrawal. These results suggest that nicotine withdrawal has a small but significant impact on the miRNA expression profile in the mouse dorsal striatum.

overall_design: 7-weeks-old C57BL/6N mice were used for experimentation. For modeling nicotine withdrawal, three mice were injected with the nicotine solution (0.5 mg/kg, 10 ml/kg, i.p.) once daily for three days, and then were injected with the mecamylamine solution (3.0 mg/kg, 10 ml/kg, i.p.) 24 hours after the last nicotine injection to precipitate nicotine withdrawal. Control mice received saline injection. One week after the last injection, mice were sacrificed and the dorsal striatum was isolated via cryosection. Total RNA was isolated by Trizol extraction.

first: Heh-In

last: Im

first: Baeksun

last: Kim


title: Transcriptome in naturally-occuring self-reactive thymocytes

city: San Francisco

first: Julie

last: Zikherman

state: CA

address: 513 Parnassus Ave

department: Medicine, Rheumatology

institute: University of California, San Francisco

summary: In the Nur77/Nr4a1-eGFP BAC Tg reporter mice, GFP expression positively correlates with TCR signaling strength. We postulate that among post-selection thymocytes, GFP expression reflects self-antigen recognition across the TCR repertoire. Here we identify differentially expressed transcripts in semimature SP4 thymocytes (CD25- CD69+ CD24+) from Nur77-eGFP BAC Tg reporter mice sorted on 10% highest, intermediate, and lowest GFP expression.

overall_design: Examination of semi-mature SP4 thymocytes with low, medium, and high self-reactivity as marked by Nur77-eGFP reporter expression. Thymi were harvested from Nur77-eGFP reporter mice, and cells were stained for live/dead fixable near-IR, CD25, CD24, CD69, CD4, and CD8a. Cells were gated on the basis of live CD25- CD24+ CD69+ CD4+ CD8a-, and then sorted on the basis of GFP expression. ~10% highest, ~10% medium, and ~10% lowest GFP-expressing cells were sorted directly into RLT buffer for generation of RNAsequencing data sets to identify differentially expressed genes. Biological triplicates of each GFP subset was obtained.

first: Letitia

last: Yang

first: James

last: Mueller

first: Alex

last: Ritter

first: Christopher

last: Scharer


title: AICAR-cardioprotection in doxorubicin-induced heart failure in rats

city: Oxford

first: Kerstin

middle: N

last: Timm

address: Mansfield Road

department: Pharmacology

country: United Kingdom

institute: University of Oxford

summary: Doxorubicin (DOX) is a widely used chemotherapeutic agent that can cause serious cardiotoxic side effects, leading to functional cardiac decline and ultimately, congestive heart failure (HF). Impaired mitochondrial function and energetics are thought to be key factors driving progression into HF. We have previously shown in a rat model of chronic intravenous DOX-administration that heart failure with reduced ejection fraction correlates with mitochondrial loss and dysfunction. Adenosine monophosphate-dependent kinase (AMPK) is a cellular energy sensor, regulating mitochondrial biogenesis and oxidative metabolism, including fatty acid oxidation. We hypothesized that AMPK activation could restore mitochondrial number and function and therefore be a novel cardioprotective strategy for the prevention of DOX-HF. Consequently, we set out to assess whether 5-aminoimidazole-4-carboxamide 1-β-D-ribofuranoside (AICAR), an activator of AMPK, could prevent cardiac functional decline in this clinically relevant rat model of DOX-HF. In line with our hypothesis, AICAR improved cardiac systolic function. This was associated with normalisation of substrate supply to the heart, as AICAR prevented DOX-induced dyslipidaemia. AICAR furthermore improved cardiac mitochondrial fatty acid oxidation, independent of mitochondrial number. In addition, we found that AICAR reduced the degree of myocardial atrophy, and RNAseq analysis showed that this was driven by normalisation of protein synthesis pathways, which are impaired in DOX-treated rat hearts. Taken together, these results show promise for the use of AICAR as a cardioprotective agent in DOX-HF to both preserve cardiac function and mass.

overall_design: We conducted RNASeq using mRNA extracted from rat hearts to elucidate the effect of doxorubicin on the heart and the cardioprotective effect of AICAR in this setting. Rats were 250g at the start of the treatment protocol.  Tissues were collected post five weekly intavenous injections of either sterile saline (n=6) or 3mg/kg/week doxorubicin and subcutanous saline (n=6) or subcutaneous 0.5g/kg AICAR (n=6). Hearts were excised under deep isoflurane anaesthesia and rapidly frozen with liquid-nitrogen cooled Wollenberger tongs. Total mRNA was extracted with a Qiagen RNeasy fibrous tissue mini kit.

first: Eshita

last: Sharma


title: Effect of conditional deletion of Bmal1 in pancreatic β-cells in modulating transcriptional islet response to STZ-mediated pro-inflammatory injury

city: Rochester

first: Kuntol

last: Rakshit

address: 200 1st St. SW

department: Physiology & Biomedical Engineering

institute: Mayo Clinic

summary: The circadian clock plays a vital role in modulating the cellular immune response. However, its role in mediating pro-inflammatory diabetogenic β-cell injury remains largely unexplored. Our studies demonstrate that conditional deletion of Bmal1 in mouse β-cells was shown to impair the ability of β-cells to recover from streptozotocin-mediated pro-inflammatory injury in vivo, leading to β-cell failure and the development of diabetes. Our study suggests that the β-cell circadian clock mediates β-cell response and recovery from pro-inflammatory injury common to the pathogenesis of diabetes mellitus.

overall_design: Control & β-Bmal1-/- mice were given submaximal dose of streptozotocin (STZ) or citrate buffer vehicle (VEH) and pancreatic islets were isolated at 24 h and 4 wk post STZ/ VEH injection.

first: Aleksey

last: Matveyenko


title: Structure Activity Relationship Read Across and Transcriptomics for Branched Carboxylic Acids

city: Mason

first: Yuqing

last: Shan

state: Ohio

address: 8700 mason montgomery road

institute: Procter & Gamble

summary: The purpose of this study was to use chemical similarity evaluations, transcriptional profiling, in vitro toxicokinetic data and physiologically based pharmacokinetic (PBPK) models to support read across for a series of branched carboxylic acids using valproic acid (VPA), a known developmental toxicant, as a comparator. The chemicals included 2-propylpentanoic acid (VPA), 2-ethylbutanoic acid (EBA), 2-ethylhexanoic acid (EHA), 2-methylnonanoic acid (MNA), 2-hexyldecanoic acid (HDA), 2-propylnonanoic acid (PNA), dipentyl acetic acid (DPA) or 2-pentylheptanoic acid (PHA), octanoic acid (OA, a straight chain alkyl acid) and 2-ethylhexanol. Transcriptomics was evaluated in four cell types (A549, HepG2, MCF7 and iCell cardiomyocytes) 6 hours after exposure to 3 concentrations of the compounds, using the L1000 platform. The transcriptional profiling data indicate that two- or three-carbon alkyl substituents at the alpha position of the carboxylic acid (EHA and PNA) elicit a transcriptional profile similar to the one elicited by VPA. The transcriptional profile is different for the other chemicals tested, which provides support for limiting read across from VPA to much shorter and longer acids. Molecular docking models for histone deacetylases, the putative target of VPA, provides a possible mechanistic explanation for the activity cliff elucidated by transcriptomics. In vitro toxicokinetic data was utilized in a PBPK model to estimate internal dosimetry. The PBPK modeling data show that as the branched chain increases, predicted plasma Cmax decreases. This work demonstrates how transcriptomics and other mode of action-based methods can improve read across.

overall_design: Four cell types were used for the transcriptomic experiments: MCF-7 (breast epithelial adenocarcinoma), A549 (lung epithelial carcinoma), HepG2 (hepatocellular carcinoma) and iCell cardiomyocytes (derived from induced pluripotent stem cells, FujiFilm Cellular Dynamics, Madison, WI). MCF-7, A549 and HepG2 cells were purchased from American Type Culture Collection (Manassas, VA) and grown in phenol red-free DMEM media containing 10% serum (Invitrogen, Carlsbad, CA). The iCell cardiomyocytes were grown in a proprietary maintenance medium (FujiFilm Cellular Dynamics, Inc.) for 168 h before chemical treatment. All cell cultures were performed in 96-well plates with 12 DMSO controls on each plate. Cells were seeded on 96 well plates and treated with 3 concentrations of each chemical DMSO for 6 h. Chemical samples were randomized across the plate and DMSO were placed on fixed well locations for each plate. Following 6 h treatment, Cells were lysed with 50 μl of Genometry Lysis Buffer to each well, sealed and stored at -80⁰C. Cell lysate plates were shipped frozen to Genometry for L1000 assasy.

first: George

middle: P

last: Daston


title: Development and evaluation of an autism pig model

city: 上海市

first: Shuai

last: Qiu

address: 上海市华东师范大学普陀校区理科大楼B327

institute: ECNU

summary: Developing cost-effective and disease-relevant animal models is essential for advancing biomedical research into human disorders. This study investigates the feasibility of a pig model for autism spectrum disorder (ASD) using embryonic exposure to valproic acid (VPA), an antiepileptic drug known to increase ASD risk. We established experimental paradigms to assess the behavioral characteristics of these pig models. Administration of VPA to Bama miniature pigs (Sus scrofa domestica) during critical embryonic stages resulted in abnormal gait, increased anxiety levels, reduced learning capabilities, and altered social patterns, while largely preserving social preference of treated piglets. Notably, we detected significant neuroanatomical changes in cortical regions associated with ASD in the VPA-treated pigs, including cortical malformation, increased neuronal soma size, decreased dendritic complexity, and reduced dendritic spine maturation. Transcriptome analysis of the prefrontal cortex of VPA-treated pigs further revealed substantial alterations in the expression of genes linked to ASD, especially genes of the dopamine signaling pathway, highlighting the model’s relevance and potential for shedding light on ASD’s underlying neuropathological and molecular mechanisms. These findings suggest that pig models could serve as a promising alternative to traditional rodent models and provide an ethical substitute for using primates in the translational research of neurodevelopmental disorders.

overall_design: To assess differential gene expression in the PFC of 3-month-old piglets treated with VPA during early gestation, we conducted RNA sequencing on six samples (three VPA-treated and three controls).

first: Jingyan

last: Jia


title: m6A-seq in the human vanetromedial prefrontal cortex in major depressive disorder

city: Montreal

first: Gustavo

last: Turecki

state: Quebec

address: 6875 Boulevard LaSalle

department: Psychiatry

country: Canada

institute: McGill University

summary: The role of N6-Methyladenosine (m6A) in the context of Major Depressive Disorder (MDD) were investigated in the ventromedial prefrontal cortex of males and females.

overall_design: m6A sequencing was performed on post-mortem ventromedial prefrontal cortex of 19 control subjects and 20 MDD cases who died by suicide, a total of 39 subjects.

first: Haruka

last: Mitsuhashi

first: Corina

last: Nagy


title: The antitumor function of Th17-derived Th1 cell is better than traditional Th1 and Th17 cells

city: Guangzhou

first: Xiao

last: Ruipei

address: Guangzhou North Stress

institute: Southern Medical University

summary: Th17 cells have better stemness and differentiation potential, and under certain circumstances, they can be stimulated to differentiate into other subcelltypes including Th1 and Treg. Here, we report that compared to traditional Th1 and Th17 cells, Th17-derived Th1 cells had greater anti-tumor activity. Both C57 mice ACT therapy and CAR-T therapy models were used to validate it. Mechanism: Th17-derived Th1 cells are able to express Th1 effector molecules as well as retain a lower level of exhaustion than Th1 cells. Additionally, it has a higher amount of type 1 interferon response and a stronger capacity for migration. Our research offers a fresh perspective on how ACT therapy and CAR-T therapy might develop in the future.

overall_design: Naïve CD4 T cell was iolated from spleen of 6 weeks old C57, then cells was activated and differentiated into Th1/Th17 hybrid cells, Th17 derive Th1 cells and pTh17 derive Th1 cells. Ultimately, these three kinds of cells (Th1/Th17, Th17 derived Th1 and pTh17 derived Th1) were used to compare differences at the transcriptome level.

first: Xiaoyi

last: Lei

first: Ruipei

last: Xiao

first: Zhe

last: Chen

first: Enguang

last: Bi


title: Epstein-Barr virus reactivation induces divergent abortive, reprogrammed, and host shutoff states by lytic progression

city: Ann Arbor

first: Elliott

middle: Daniel

last: SoRelle

state: MI

address: 1137 Catherine St.

department: Microbiology & Immunology

institute: The University of Michigan

summary: Viral infection leads to heterogeneous cellular outcomes ranging from resistance to virion amplification and cell death. In this study, we apply single-cell techniques to analyze diverse fate trajectories during Epstein-Barr virus (EBV) lytic reactivation in three B cell models. We expand upon the previous finding that c-Myc regulates lytic reactivation and determine that MYC downregulation during the lytic phase is correlated with a loss of germinal center (GC) B cell expression signatures. Consistent with prior work, lytic induction yields both abortive and complete reactivation. Abortive lytic cells, which express high STAT3 and initiate but do not complete the lytic cycle, also upregulate NF-kB and IRF3 pathway target genes. Cells that proceeded through the full lytic cycle exhibited unexpected and striking expression of genes associated with cellular reprogramming, developmental progenitor phenotypes, and cytokine upregulation. Distinct subpopulations of lytic cells further displayed variable profiles for transcripts known to escape virus-mediated host shutoff. These data reveal previously unknown and promiscuous outcomes of lytic reactivation with broad implications for viral replication and EBV-associated oncogenesis.

sample_organism:
Saguinus oedipus
Homo sapiens

platform_organism:
Saguinus oedipus
Homo sapiens

overall_design: Induction of EBV lytic reactivation in three B cell model systems was examined using scRNA-seq. The models used in this study included two Burkitt Lymphoma cell lines (Akata, P3HR1-ZHT), one of which (P3HR1-ZHT) is engineered for chemically-inducible viral reactivation, and one lymphoblastoid cell line (B958-ZHT) also engineered for inducible reactivation with 4-hydroxytamoxifen (HT). Sequence libraries were prepared from P3HR1-ZHT cells under the following conditions: unstimuated (UT), methanol control treatment for 24 h (MeOH), 24 h after treatment with HT (HT_24h; 2 replicates), 48 h after HT (HT_48h), and 72 h after HT (HT_72h). Sequence libraries from B958-ZHT cells were prepared from the following conditions: unstimulated (UT) and 24 h after HT (HT_24h). Sequence libraries from Akata cells were prepared from the following conditions: unstrimulated (UT) and 24 h after lytic induction with anti-Ig antibody (Ig).

first: Lauren

last: Haynes

first: Katherine

last: Willard

first: James

last: Ch'ng

first: Heather

last: Christofk

first: Micah

last: Luftig


title: Transcriptomic analysis of the effects of mTOR inhibitor everolimus, CDK8/19 inhibitor SNX631 and their combination in MDA-MB-468 xenografts

city: COLUMBIA

first: HAO

last: JI

state: SC

address: 715 SUMTER ST CLS RM 109

department: College of Pharmacy

institute: University of South Carolina

summary: RNA-Seq analysis was carried out to investigate the transcriptomic changes associated with the development of in vivo everolimus resistance and the effects of CDK8/19 inhibition on the development of resistance in MDA-MB-468 TNBC xenografts.  In one study (short-term), tumor samples were collected after 30-40 days of treatment with vehicle or everolimus, when the tumor growth was still suppressed by the mTOR inhibitor.   In another study (long-term),  tumor samples were collected after 60-153 days of treatment with vehicle, everolimus, CDK8/19 inhibitor SNX631 or their combination,  when tumors treated with everolimus but not with the drug combination developed resistance.

overall_design: MDA-MB-468 cells (2x10^6 cells in 0.1 mL 50% Matrigel per animal) were inoculated in female NSG mice orthotopically under fat pad. Once tumors reached ~80 mm^3, mice were randomized into groups with similar-size tumors for further treatments. For the short-term study, mice received vehicle (30 days) or 2 mg/kg everolimus (38 days) by daily oral gavage.  For the long-term study, mice received vehicle (p.o., q.d., up to 78 days) or SNX631 (350 ppm in medicated food, up to 96 days) or  2 mg/kg everolimus (p.o., q.d., up to 148 days) or in everolimus-SNX631 combination (148 days) .   At the endpoint for each tumor, tumors were dissected from euthanized animals using sterilized surgical tools. Pieces of non-necrotic tumor tissues 50~100mm^3 were excised at the margin near midline section for RNA extraction.  At least seven biological replicates (xenograft tumors from different animals) per study group were analyzed.

first: Xiaokai

last: Ding

first: Hao

last: Ji

first: Mengqian

first: Igor

last: Roninson

first: Eugenia

last: Broude


title: Absence of SMARCB1 in rhabdoid tumor cells increases sensitivity to translation inhibition and alters translation efficiency of specific mRNAsn

city: Stanford

first: Anastasia

last: Hains

email: hainsa@uchicago.edu, hains@stanford.edu

address: 269 Campus Drive

institute: Stanford University

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

overall_design: Refer to individual Series


title: RAN directly binds and stabilizes G3BP1 mRNA in the nucleus to facilitate proliferation and metastasis of nasopharyngeal carcinoma (si-RAN RNA-seq)

first: Yang

last: Pan-Yang

address: 651 Dongfeng East Road

department: Collaborative Innovation Center of Cancer Medicine

institute: Sun Yat-sen University Cancer Center

summary: RNA binding proteins (RBPs) play a critical role in tumor progression by participating in the post-transcriptional regulation of RNA. However, the expression and function of RBPs in nasopharyngeal carcinoma (NPC) remain elusive. This study identified 5 RBPs (RAN, EZH2, RDM1, HRSP12, and ALYREF) that were up-regulated in NPC and could promote NPC cells migration or proliferation. RAN was first investigated because of its most significant effect on NPC cells. Functionally, RAN facilitated NPC proliferation, migration, and invasion in vitro and in vivo. High expression of RAN was associated with poor prognosis of NPC patients and could be performed as a prognostic biomarker. Mechanistically, RAN mediated the nucleus import of TDP43 and enhanced TDP43 nuclear distribution. On the other hand, RAN was directly bound to the coding sequence of G3BP1 mRNA and served as an adapter to facilitate TDP43 interacting with G3BP1 mRNA 3’untranslated region. Thereby increased G3BP1 mRNA stability in the nucleus and led to up-regulation of G3BP1, which further enhanced ATK/ERK phosphorylation and ultimately promoted NPC proliferation and metastasis.

overall_design: RNA-seq was performed to search downstream RNAs regulated by RAN, RAN was knocked down by siRNA in HONE-1 cells and total RNA from RAN silenced or control  HONE-1 cells were extracted. Gene expression profiling analysis was performed using data from RNA-seq of si-RAN and control HONE-1 cells.

first: Pan-Yang

last: Yang

first: Zhen-Yu

first: Denghui

last: Wei

first: Ying

last: Sun


title: RAN directly binds and stabilizes G3BP1 mRNA in the nucleus to facilitate proliferation and metastasis of nasopharyngeal carcinoma (si-G3BP1 RNA-seq)

overall_design: RNA-seq was performed to search downstream pathway regulated by G3BP1, G3BP1 was knocked down by siRNA in HONE-1 cells and total RNA from G3BP1 silenced or control  HONE-1 cells were extracted. Gene expression profiling analysis was performed using data from RNA-seq of si-G3BP1 and control HONE-1 cells.


title: RAN directly binds and stabilizes G3BP1 mRNA in the nucleus to facilitate proliferation and metastasis of nasopharyngeal carcinoma


title: RAN directly binds and stabilizes G3BP1 mRNA in the nucleus to facilitate proliferation and metastasis of nasopharyngeal carcinoma (anti-RAN RIP-seq)

overall_design: RIP-seq was performed to identify downstream RNAs interacting with RAN in two NPC cell lines HONE-1 and SUNE-1 respectively. Cell lysates were incubated with protein A/G magnetic beads coated with 5 μg of RAN antibody overnight at 4 °C. The immunoprecipitated RNA was separated with elution buffer and purified for next-generation sequencing.


title: Gene expression profile of human monocyte-derived macrophage stimulated with AGP, anti-CD74 antibody, and mycobacterium avium infaction

city: Kumamoto

first: Yoshihiro

last: Komohara

address: Honjo 1‐1‐1, Chuouku

department: Department of Cell Pathology

institute: Graduate School of Medical Sciences, Kumamoto University

summary:
Macrophages are immune cells inctributing to several kinds of diseases, and activation status of macrophages are considered to be regulated by several stimulations.
To test the gene expression change in activated macrophages, human monocyte-derived macrophages were stimulated with alpha1-acid glycoprotein (AGP), anti-CD74 blocking antobody, anti-CD74-agonistic antibody. Macrophages infected with Micobacterium avium were also tested. The mRNA expression in control, stimulated, infected macrophages were analysed.

overall_design: Macrophages (200000 cells/well in 6-well plate) were stimulated with AGP(ORM1) (1mg/ml) and anti-CD74 antibodies (10mg/mL). Macrophages were infected with mycobacterium avium for 3 days and 7days. Based culture madium was DMEM/HamF12 medium supplemented with 10% FBS.


title: High Density Domain-Focused CRISPR Screens Reveal Novel Epigenetic Regulators of HOX/MEIS Activation in Acute Myeloid Leukemia [RNA-seq, SGF29 KO vs. NTC in MOLM13 and U937]

city: La Jolla

first: Karina

middle: Ofelia

last: Barbosa Guerra

address: 10901 North Torrey Pines Rd, Deshpande Lab RM6210B

department: Tumor Initiation & Maintenance Program

institute: Sanford Burnham Prebys Medical Discovery Institute

summary: To investigate the transcriptomic effects upon loss of SGF29 in the AML cell lines U937 and MOLM13, we performed CRISPR knockouts, as well as transduction with non-targeting controls, and harvested samples for bulk RNA-Seq.

overall_design: We performed gene expression profiling analysis using data obtained from RNA-seq of SGF29 CRISPRko perturbation compared to non-targeting controls.

last: Barbosa

first: Anagha

last: Deshpande

first: Rabi

last: Murad.

first: Aniruddha


title: Monocytes Give Rise To Langerhans Cells That Preferentially Migrate To Lymph Nodes At Steady State

city: Palo Alto

first: Rebeca

last: Arroyo Hornero

state: California

address: Campus Drive

institute: Stanford

summary: It is still debated whether the homeostatic function of tissue macrophages is dependent on their origin, and evidence suggests this may depend on the tissue of macrophage residence. Here, we answered this question for Langerhans cells (LCs), an embryonic-derived mononuclear phagocyte that has features of both tissue macrophages and dendritic cells. Using time-course analyses, bone marrow chimeras and fate tracing models, we found that the complete elimination of embryonic-derived LCs at steady state results in their repopulation from circulating monocytes. However, monocyte-derived LCs inefficiently replenished the epidermal niche. Instead, these cells preferentially migrated to skin draining lymph nodes. Mechanistically, we show that the superior migratory capability of monocyte-derived LCs is associated with higher expression of CD207/Langerin, a C-type lectin involved in the capture of skin microbes. Our data shows that ontogeny plays a role in the migratory behavior of epidermal LCs.

overall_design: Epidermal LCs from a Cd207-DTR BMC were FACs sorted using a gating strategy and origin was determined using index-sorting for monocyte-derived LC (mLC) and embryonic LC (eLC)

first: Hayley

last: Raquer-McKay

first: Juliana

last: Idoyaga


title: m6Am sequesters PCF11 to promote full-length transcription [catabolicSLAM]

city: Shenzhen

first: Wee Siong Sho

last: Goh

state: Guangdong

address: Shenzhen Bay Laboratory, No. 5, Kelian Road, Goh Lab, A1819, Guangming District

department: Institute of Molecular Physiology

institute: Shenzhen Bay Laboratory

type: Expression profiling by genome tiling array

summary: RNA modifications regulate how RNAs metabolize and function to impact development and diseases. N6,2’-O-dimethyladenosine (m6Am) is one such modification and despite being abundant, m6Am function remains unclear. Here, we identified cleavage and polyadenylation factor, PCF11 as a m6Am-specific binding protein. Direct quantification of mature versus nascent RNAs revealed that m6Am does not regulate mRNA stability but promotes transcription of nascent RNAs. m6Am caused RNA Polymerase II (Pol II) to be more processive when transcribing m6Am-modified RNAs. Rather than PCF11 regulating m6Am-modified RNA, m6Am sequesters PCF11 away from proximal Pol II, suppressing premature dissociation of elongating Pol II and promoting Pol II full-length transcription of m6Am-modified RNAs. This establishes a mechanism through which an RNA modification regulates transcription.

overall_design: s4U labelling of nascent RNA for 3'end mRNA sequencing

first: Wee Siong


title: Analysis of transcriptomes of germinal center (GC) B cells derived from miR-155-sufficient and -deficient mice

city: London

first: Probir

last: Chakravarty

address: 1 Midland Road

institute: Francis Crick Institute

summary: To investigate the impact caused by miR-155 ablation, positively selected light zone (LZ) GC B cells were sorted by flow cytometry and used to analyse transcriptomic profiles.

overall_design: Splenic B cells derived from miR-155-sufficient or -deficient mice expressing HEL-specific B cell receptor together with endogenous cMyc-GFP reporter cassette were adoptively transferred into congenic mice, followed by immunization with HEL3x mutant conjugated with sheep red blood cells (SRBCs). After five days, cMyc-GFP+ HEL+ LZ GC-B cells were sorted and used for the mRNA sequencing library.

first: Rinako

last: Nakagawa


title: m6Am sequesters PCF11 to promote full-length transcription [RNA-seq]

overall_design: RNA-seq

first: W

last: Sho Goh


title: m6Am sequesters PCF11 to promote full-length transcription [m6ACE]


title: m6Am sequesters PCF11 to promote full-length transcription


title: Evaluating the presence of rare ERα-antagonism induced cells in WT tissues

city: South San Francisco

first: Jackson

last: Liang

address: 1 DNA Way

department: Discovery Oncology

institute: Genentech

summary: To examine the effects of ERα antagonism (giredestrant) on Esr1-wildtype and Esr1-mutant mammary glands, animals were treated with hormone pellets (E2, P4), then dosed daily with vehicle (MCT) or giredestrant (30 mg/kg, p.o.) for four days. Following dosing, individual mammary epithelial cell populations were FACS-isolated and sequenced. Rare giredestrant-induced novel cells (at least 1000 cells per sample) were collected from WT tissues and compared to mutant-induced novel cells.

overall_design: To examine the effects of ERα antagonism (giredestrant) on Esr1-wildtype and Esr1-mutant mammary glands, animals were treated with hormone pellets (E2, P4), then dosed daily with giredestrant for four days. 18 samples are included, 3 biological replicates per condition.

first: Marc

last: Hafner

first: Ciara

last: Metcalfe


title: Evaluating an Esr1-mutant novel cell population emerging upon ERα-antagonism by single RNA-seq

summary: To examine the effects of ERα antagonism (giredestrant) on Esr1-wildtype and Esr1-mutant mammary glands, animals were treated with hormone pellets (E2, P4), then dosed daily with vehicle (MCT) or giredestrant (30 mg/kg, p.o.) for four days. Following dosing, EpCAM+ cells were isolated and used for single cell sequencing. For each condition, cells from 2 animals were pooled for sequencing.

overall_design: Esr1-wildtype and Esr1-mutant mammary glands, animals were treated with hormone pellets (E2, P4), then dosed daily with vehicle (MCT) or giredestrant. 4 samples are included.


title: Evaluating an Esr1-mutant novel cell population emerging upon ERα-antagonism by bulk RNA-seq

summary: To examine the effects of ERα antagonism (giredestrant) on Esr1-wildtype and Esr1-mutant mammary glands, animals were treated with hormone pellets (E2, P4), then dosed daily with vehicle (MCT) or giredestrant (30 mg/kg, p.o.) for four days. Following dosing, individual mammary epithelial cell populations were FACS-isolated and sequenced.

overall_design: To examine the effects of ERα antagonism (giredestrant) on Esr1-wildtype and Esr1-mutant mammary glands, animals were treated with hormone pellets (E2, P4), then dosed daily with vehicle (MCT) or giredestrant for four days. 51 samples are included, 3 biological replicates per condition.