title: VCAM-1 mediates proximal tubule-immune cell cross talk in failed tubule recovery during AKI-to-CKD transition

city: New Haven

first: Leyuan

last: Xu

state: CT

address: 300 Cedar Street, Ste S255B

department: Internal Medicine/Section of Nephrology

institute: Yale University

summary: Studies in animal models have suggested a linkage between the inflammatory response to injury and subsequent nephron loss during the acute kidney injury (AKI) to chronic kidney disease (CKD) transition. Failure of normal repair during the CKD transition correlates with de novo expression of vascular cell adhesion protein-1 (VCAM-1) by a subset of injured proximal tubule cells. This study identified the role of VCAM-1 expression in promoting the failed repair state. Single-cell transcriptome analysis of patients with AKI and CKD and whole kidney RNA and protein analyses of mouse models of CKD confirmed a marked increase of VCAM-1 expression in the proximal tubules of injured kidneys. In immortalized mouse proximal tubular cells and primary cultured renal cells (PCRCs), VCAM-1 expression was induced by proinflammatory cytokines including tumor necrosis factor (TNF)-α and interleukin (IL)-1β. Analyses of bulk RNA sequencing of TNF-α-treated primary cultured renal cells or pseudo-bulk RNA sequencing of biopsies from Kidney Precision Medicine Project datasets indicated activation of NF-κB and an enrichment of inflammatory response and cell adhesion pathways in VCAM-1-positive cells. Pharmacological inhibition of NF-κB signaling or genetic deletion of myeloid differentiation factor 88 and TIR domain-containing adapter-inducing interferon-β suppressed TNF-α- and IL-1β-induced VCAM-1 expression in vitro. TNF-α stimulation or overexpression of VCAM-1 significantly increased splenocyte adhesion to the mouse proximal tubular monolayer in culture. These results demonstrate that persistence of proinflammatory cytokines after AKI can induce NF-κB-dependent VCAM-1 expression by proximal tubule cells, mediating increased immune cell adhesion to the tubule and thus promoting further tubule injury and greater risk of progression from AKI to CKD.

overall_design:
To study the mechanism(s) of VCAM-1 induction and its role in promoting immune cell adhesion, we used primary cultured renal cells (PCRCs) that were treated with TNFα ± NF-κB-specific inhibitor, ACHP.
We then performed gene expression profiling analysis using data obtained from RNA-seq of PCRCs treated with PBS, TNFα, and TNFα+ACHP.


title: Single cell sequencing unravels the cellular diversity that shapes neuro- and gliogenesis in the fast aging killifish (N. furzeri) brain

status: Public on Sep 03 2024

city: Leuven

first: RAJAGOPAL

last: AYANA

state: Flemish brabant

address: Naamsestraat 59

department: Department of Biology

country: Belgium

institute: KU Leuven

type: Expression profiling by high throughput sequencing

summary: The African turquoise killifish combines a short lifespan with age-dependent loss of neuroregenerative capacity, making it a well-suited model for studying brain repair mechanisms in the context of aging. To investigate the extent of cellular diversity that shapes neuro- and gliogenesis, we performed single cell sequencing of the adult telencephalon. Our analysis identifies seventeen cell types including neuronal cells, and progenitors of glial and non-glial nature. Further subclustering unveils four radial glia (RG) types, one atypical non-glial progenitor (NGP) and two intermediate subtypes. Validation of our data in situ reveals a distinct spatial setting for defined RG subtypes, reflecting the distribution of morphologically and physiologically distinct populations. Lineage inference analysis suggests neuroepithelial-like radial glia and NGP to be the start point and intercessor of neural development, respectively. Neuronal sub-clustering uncovers immature and mature excitatory or inhibitory sub-clusters. This complete catalogue of killifish telencephalon cell types is accessible via an online tool, providing a resource to understand neurogenesis in healthy brains and upon injury or disease.

overall_design: Telencephalon region (homologous to mammalian SGZ/SVZ) of the 6-week-old adult female killifish was harvested for single-cell RNA sequencing.

first: Rajagopal

last: Ayana

first: Caroline

last: Zandecki

first: Jolien

last: Van Houcke

first: Valerie

last: Mariën

first: Eve

last: Seuntjens

first: Lut

last: Arckens


title: Interaction between subventricular zone microglia and neural stem cells impacts the neurogenic response in a mouse model of cortical ischemic stroke

status: Public on Sep 02 2024

city: Miami

first: James

middle: S

last: Choi

state: FL

address: 1095 NW 14th Terrace

country: USA

institute: University of Miami Miller School of Medicine

summary: After a stroke, the neurogenic response from the subventricular zone (SVZ) to repair the brain is limited. Microglia, as an integral part of the distinctive SVZ microenvironment, control neural stem / precursor cell (NSPC) behavior. Here, we show that discrete stroke-associated SVZ microglial clusters negatively impact the innate neurogenic response, and we propose a repository of relevant microglia–NSPC ligand–receptor pairs. After photothrombosis, a mouse model of ischemic stroke, the altered SVZ niche environment leads to immediate activation of microglia in the niche and an abnormal neurogenic response, with cell-cycle arrest of neural stem cells and neuroblast cell death. Pharmacological restoration of the niche environment increases the SVZ-derived neurogenic repair and microglial depletion increases the formation and survival of newborn neuroblasts in the SVZ. Therefore, we propose that altered cross-communication between microglial subclusters and NSPCs regulates the extent of the innate neurogenic repair response in the SVZ after stroke.

overall_design: Using a mouse model of cortical ischemic stroke (photothrombotic ischemia, PT), we performed single-cell RNA sequencing (scRNAseq) on neural stem progenitor cells (NSPCs) and microglia. Tamoxifen-treated Nestin-CreERT2:R26-yfp transgenic mice underwent PT after injection of photosensitive dye and light exposure. Subventricular zone (SVZ) compartments were microdissected from brains of control (Ctrl, no PT), 1 day post-PT (S1d), and 7 days post-PT (S7d) animals. SVZ tissue was digested enzymatically and dissociated mechanically, and GFP+ SVZ NSPCs and CD11b+CD45low microglia were FACS isolated before scRNAseq and library prep using the plate-based mCEL-Seq2 protocol. cDNA libraries were sequenced with Illumina HiSeq 3000. Reads were demultiplexed with bcl2fastq and aligned to the GRCm39 mouse reference genome.

first: Suvra

last: Nath

first: Jose

last: Carlos Martinez Santamaria

first: Yu-Hsuan

last: Chu

first: Pasquale

last: Conforti

first: Jia-Di

last: Lin

first: Roman

last: Sankowski

first: Lukas

last: Amann

first: Christos

last: Galanis

first: Kexin

last: Wu

first: Sachin

last: Deshpande

first: Andreas

last: Vlachos

first: Marco

last: Prinz

first: Jae

middle: K

last: Lee

first: Christian

last: Schachtrup


title: Pseudomonas aeruginosa post-translational responses to elevated c-di-GMP levels

city: Braunschweig

first: Matthias

last: Preusse

address: Inhoffenstr. 7

country: Germany

institute: Helmholtz Centre for Infection Research

summary: C-di-GMP signaling can directly influence bacterial behavior by affecting the functionality of c-di-GMP-binding proteins. In addition, c-di-GMP can exert an indirect and more global effect on gene transcription or translation, e.g. via riboswitches or by binding to transcription factors. In this study, we investigated the effects of changes in intracellular c-di-GMP levels on gene expression and protein production in the opportunistic pathogen Pseudomonas aeruginosa. We induced c-di-GMP production via an ectopically introduced diguanylate cyclase and recorded the transcriptional, translational as well as proteomic profile of the cells. We demonstrate that rising levels of c-di-GMP in P. aeruginosa under growth conditions otherwise characterized by low c-di-GMP levels immediately cause a switch to a non-motile, auto-aggregative phenotype. This switch became apparent before any c-di-GMP-dependent role on transcription, translation, or protein abundance could be observed. Our results indicate that sudden rises in global c-di-GMP pools affect the P. aeruginosa phenotype via an alteration of protein functionality, rather than an impact on global gene transcription or translation.

overall_design: Profiling of cellular consequences of an elevated c-di-GMP level in exponential phase – RNA-seq, Ribo-seq

first: Sarina

last: Bense

first: Julius

last: Witte

first: Michal

last: Koska

first: Lorena

last: Pezoldt

first: Astrid

last: Dröge

first: Oliver

last: Hartmann

first: Mathias

last: Müsken

first: Julia

last: Schulze

first: Timm

last: Fiebig

first: Heike

last: Bähre

first: Sebastian

last: felgner

last: Pich

first: Susanne

last: Häussler


title: Mepolizumab-induced Changes in Nasal Methylome and Transcriptome to Predict Response in Asthma

city: Nottingham

first: Ian

last: Sayers

state: Nottinghamshire

address: Biodiscovery Institute, Science Road, University Park

country: United Kingdom

institute: University of Nottingham

summary: We aimed to detect a predictive or early nasal gene expression and/or DNA methylation signature that could identify responders or non-responders to Mepolizumab at one year.

overall_design: Data for 27 asthma patients enrolled in The PROCLAIM study (Poor Response tO monoClonaL therApy In asthMa) who matched the UK NHS criteria for receiving Mepolizumab was used (ethics REC18/EM/0268). Nasal brushes were taken at baseline (pre-drug) and after 3-months of treatment. Responders were defined as patients who had a 50% reduction in oral corticoid steroid use and/or 50% reduction in exacerbation frequency after one year (NICE guidelines, TA431, 2017). Gene expression was compared between responders and non-responders at baseline and at 3 months post mepolizumab treatment.

first: Kamini

last: Rakkar

first: Yik

middle: L

last: Pang

first: Poojitha

last: Rajasekar

first: Michael

middle: A

last: Portelli

first: Robert

middle: J

last: Hall

first: Rachel

last: Clifford

first: Dominick

last: Shaw


title: STK19 accelerates the clearance of lesion-stalled RNAPII to facilitate transcription-coupled DNA repair

city: Leiden

first: Martijn

middle: S.

last: Luijsterburg

state: zuid-holland

address: Einthovenweg 20

department: Human Genetics

country: Netherlands

institute: LUMC

summary: Transcription-coupled DNA repair (TCR) removes bulky DNA lesions impeding RNA polymerase II (RNAPII) transcription. Recent studies have outlined the stepwise assembly of TCR factors CSB, CSA, UVSSA, and TFIIH around lesion-stalled RNAPII. However, the mechanism and factors required for the transition to downstream repair steps, including RNAPII removal to provide repair proteins access to the DNA lesion, remain unclear. Here, we identify STK19 as a new TCR factor facilitating this transition. Loss of STK19 does not impact initial TCR complex assembly or RNAPII ubiquitylation but delays lesion-stalled RNAPII clearance, thereby interfering with the downstream repair reaction. Cryo-EM and mutational analysis reveal that STK19 associates with the TCR complex, positioning itself between RNAPII, UVSSA, and CSA. The structural insights and molecular modeling suggest that STK19 positions the ATPase subunits of TFIIH onto DNA in front of RNAPII. Together, these findings provide new insights into the factors and mechanisms required for TCR.

overall_design: To study whether STK19 is involved in the removal of stalled RNAPII from chromatin after UV-C irradiation, we performed TCR-seq experiments (stranded RNAPII-S2 ChIP-seq) in WT, STK19-KO or CSB-KO cell lines after mock treatment or at 1 hour or 8 hours after 12J/m2 UV-C.

first: Diana

last: van den Heuvel

first: Marta

last: Rodríguez-Martínez

first: Paula

last: van der Meer

first: Nicolas

last: Nieto Moreno

first: Jiyoung

last: Park

first: Hyun-Suk

last: Kim

first: Janne

last: van Schie

first: Annelotte

middle: P

last: Wondergem

first: Areetha

last: D'Souza

first: George

last: Yakoub

first: Anna

middle: E

last: Herlihy

first: Krushanka

last: Kashyap

first: Thierry

last: Boissière

first: Jane

last: Walker

first: Richard

last: Mitter

first: Katja

last: Apelt

first: Klaas

last: de Lint

first: Idil

last: Kirdök

first: Mats

last: Ljungman

first: Rob

middle: M

last: Wolthuis

first: Patrick

last: Cramer

first: Orlando

middle: D

last: Schärer

first: Goran

last: Kokic

first: Jesper

middle: Q

last: Svejstrup


title: STK19 accelerates the clearance of lesion-stalled RNAPII to facilitate transcription-coupled DNA repair [BrU-seq]

first: Martijn

last: Luijsterburg

overall_design: To study the role of STK19 in transcription recovery after UV-C irradiation, we performed nascent RNA-seq experiments in WT, STK19-KO or STK19-rescue cell lines after mock treatment or at 3 hours or 16 hours after 12J/m2 UV-C

first: Diana

last: van den Heuvel

first: Marta

last: Rodríguez-Martínez

first: Paula

last: van der Meer

first: Nicolas

last: Nieto Moreno

first: Jiyoung

last: Park

first: Hyun-Suk

last: Kim

first: Janne

last: van Schie

first: Annelotte

last: Wondergem


title: G-quadruplexes within the SVA retrotransposon modulate TAF1 gene expression in X-linked Dystonia Parkinsonism

city: Padova

first: Giulia

last: Nicoletto

address: Via Gabelli 63

department: Molecular Medicine

country: Italy

institute: UNIPD

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Epigenome reprogramming through H3K27 and H3K4 trimethylation as a resistance mechanism to DNA methylation inhibition in BRAFV600E-mutated CRC. [ChIP-Seq]

city: HOUSTON

first: Ajay

middle: Kumar

last: Saw

state: TX

address: 7900 Cambdrige St

department: Department of Genomic Medicine

institute: University Of Texas MD Anderson Cancer Center, Houston, Texas, USA.

summary: BRAFV600E-mutated colorectal cancer (CRC) exhibits a strong correlation with DNA hypermethylation suggesting this subgroup of tumors is uniquely presenting epigenomic phenotypes. Nonetheless, the traditional epigenomic therapeutic agent, 5-azacitidine, which inhibits DNA methyltransferase activity, did not yield sufficient improvements in the efficacies of BRAFV600E CRC in vivo. We utilized a patient-derived xenograft model and confirmed an effective reduction of DNA methylation levels upon 5-azacitidine treatment yet failed to restore gene expression patterns. This study unbiasedly explored the adaptive engagement of other epigenomic modifications upon the profound decrease in DNA methylation by azacitidine treatment. A loss of histone acetylation and a gain of histone methylations, including H3K27 and H3K4 trimethylation, were observed around hypomethylated regions. Our findings suggested a compensatory increase in repressive histone mark, H3K27 trimethylation, around treatment-induced hypomethylated regions, which suggests the involvement of polycomb repressive complex (PRC) activity around the genome with lost DNA methylation, therefore maintaining the suppression of key genes. Combined inhibition of PRC activity through EZH2 inhibitor with azacitidine treatment additively improved efficacies in BRAFV600E CRC cells. In conclusion, DNA hypermethylation exhibits a close association with H3K27me3 and PRC activity in BRAFV600E CRC, and simultaneous blockade of DNMT and EZH2 holds promise as a potential therapeutic strategy for patients with BRAFV600E-mutated CRC.

overall_design: Chromatin Immunoprecipitation DNA sequencing (ChIP-seq) on histone marks heH3K4me3, H3K27ac, H3K27me3, and H3K9me3 in BRAFV600E-mutated CRC patient-derived xenograft (PDX).

first: Hey

last: Lee

first: Scott

last: Kopetz

first: Kunal

last: Rai


title: CBP/P300 BRD Inhibition Reduces Neutrophil Accumulation and Activates Antitumor Immunity in TNBC

city: Houston

first: Xiang

last: Zhang

address: One Baylor Plaza

institute: Baylor College of Medicine

summary: In neutrophil-enriched triple-negative breast cancer (TNBC) models, CREB binding protein (CBP)/P300 bromodomain (BRD) inhibition reduces tumor growth and systemic neutrophil accumulation while stimulating an antitumor immune response. This improves standard-of-care therapies, suggesting a potential therapeutic benefit of CBP/P300 BRD inhibitors for neutrophil-enriched TNBC.

overall_design: For single-cell RNA sequencing of 2208L tumors, the tumors were dissociated and sorted to select live cells. For single-cell RNA-sequencing of bone marrow cells, CD45+/Ter119-/DAPI- cells and lineage-/c-Kit+/CD45+/DAPI- cell were selected and sequenced separately. For both experiments, cells from 3 mice were pooled for each treatment group (vehicle or IACS-70654). The mice were treated for 7 days on a 3-on/2-off schedule.

first: Xueying

last: Yuan

first: Jeffrey

last: Rosen

first: Fengshuo

last: Liu


title: Retinal ganglion cell-derived semaphorin 6A segregates starburst amacrine cell dendritic scaffolds to organize the mouse inner retina

city: Baltimore

first: Rebecca

last: James

state: MD

address: 725 N. Wolfe Street

department: Neuroscience

institute: Johns Hopkins University

summary: To form functional circuits, neurons must settle in their appropriate cellular locations, and then project and elaborate neurites to contact their target synaptic neuropils. Laminar organization within the vertebrate retinal inner plexiform layer (IPL) facilitates pre- and postsynaptic neurite targeting, yet the precise mechanisms underlying establishment of functional IPL subdomains are not well understood. Here, we explore mechanisms defining the compartmentalization of OFF and ON neurites generally, and OFF and ON direction-selective neurites specifically, within the developing mouse IPL. We show that semaphorin 6A (Sema6A), a repulsive axon guidance cue, is required for delineation of OFF versus ON circuits within the IPL: in the Sema6a null IPL, the boundary between OFF and ON domains is blurred. Furthermore, Sema6A expressed by retinal ganglion cells (RGCs) directs laminar segregation of OFF and ON starburst amacrine cell dendritic scaffolds, which themselves serve as a substrate upon which other retinal neurites elaborate. These results demonstrate that RGCs, the first type of neuron born within the retina, play an active role in functional specialization of the IPL.

overall_design: mRNA from each sample corresponds to an isolated single TdTomato-labeled starburst amacrine cell isolated by FACS across early postnatal development (P1, P4, P8, P11, and P16). scRNA-seq libraries were generated using a modified SmartSeq2 protocol (Chevee et al., 2018, doi: 10.1016/j.celrep.2017.12.046) and profiled by NextSeq mid output 2x75 paired-end sequencing.

first: Natalie

middle: R

last: Hamilton

first: Lola

last: Nicole Huffman

first: Matthew

last: Brown

first: Victoria

middle: N

last: Neckles

first: R. Jeroen

last: Pasterkamp

first: Loyal

last: Goff

first: Alex

last: Kolodkin


title: Mouse enteric neurons control intestinal plasmacytoid dendritic cell function via serotonin-HTR7 signaling [DC_5HT RNA-seq]

status: Public on Sep 04 2024

city: boston

first: Hailong

state: MA

address: 181 longwood ave

institute: Brigham and Women's Hospital, Harvard Medical School

summary: Serotonergic neurons have multiple roles, including regulating mood and behavior1,2, yet it remains unclear which of these phenotypes are caused by serotonin produced by the central versus the peripheral nervous system. Neuronal serotonin is produced by the enzyme tryptophan hydroxylase 2 (Tph2). Mice that lack Tph2 exhibit decreased gut motility4 and several behavioral disorders, including decreased anxiety-like behavior and increased aggression5. To clarify the specific role of peripheral serotonergic circuits, here we engineered mice with intact Tph2 in central neurons but lacking Tph2 in peripheral neurons (Tph2fl/fl; Hand2-Cre). We discovered that PSN contribute to gut motility and are important in the control of anxiety-like behavior but do not contribute to impulsive aggression. Thus, central and peripheral serotonergic neurons have different roles in behavioral regulation. Intriguingly, animals lacking peripheral neuronal Tph2 had deficiencies in enteric immune cells, including reduced abundance of dendritic cells (DC) and IgA+ B cells in the small intestine and elevated susceptibility to oral Salmonella infection. Mechanistic studies suggest that serotonin produced by PSN promotes activation of DC through the 5-HT7 receptor, thereby facilitating DC-mediated differentiation of IgA+ B cells. Our findings highlight the importance of serotonin produced by peripheral neurons for control of behavior and gut defense and suggest that these circuits could be valuable targets for therapeutics.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for LPDC with serotonin or Htr7 antagonist treatment

last: Waldor


title: Targeting transcriptional factor YY1 is synthetic lethal with loss of the histone demethylase KDM5C [CUT&Tag]

city: Wuhan

first: Jie

last: Xiong

address: 185, Donghu road

country: China

institute: Wuhan University

summary: In this study, we screened KDM5C-binding proteins and found that Yin Yang 1 (YY1) interacts with KDM5C. Interestingly, a synergistic antitumor effect was exerted when both KDM5C and YY1 were depleted, and targeting YY1 appeared to be a vulnerability in KDM5C-deficient cancer cells. Mechanistically, KDM5C is essential for global YY1 chromatin recruitment, especially at promoters. Moreover, an intact KDM5C JmjC domain but not KDM5C histone demethylase activity is required for KDM5C-mediated YY1 chromatin binding.Transcriptional profiling revealed that dual inhibition of KDM5C and YY1 led to significantly increased transcriptional repression of many cell cycle- and apoptosis-related genes. In summary, our work demonstrates a synthetic lethal interaction between YY1 and KDM5C and suggests a therapeutic vulnerability that can be targeted using combination therapies.

overall_design: Cleavage Under Targets and Tagmentation (CUT&Tag) for KDM5C and YY1 in ACHN cells.

first: Qian

last: Zheng

first: Feng

last: Li


title: Rosmarinic acid alleviates radiation-induced pulmonary fibrosis by downregulating tRNA N7-Methylguanosine modification-regulated fibroblast to myofibroblast transition through the exosomes pathway [Ribo-Seq]

city: Nanning

first: jinglin

last: mi

address: shuangyong road

institute: the first affiliated hospital of guangxi medical university

summary: Radiation-induced pulmonary fibrosis (RIPF) is a common complication after radiotherapy in thoracic cancer patients, and there is a lack of effective treatment methods. The aim of this study was to explore the protective effect of rosmarinic acid (RA) on RIPF in mice as well as the underlying mechanism.We found that RA exerted an antifibrotic effect on lung tissues of RIPF mouse models and inhibited the progression of FMT through exosomes derived from lung epithelial cells. Mechanistically, RA reduced the transcription and translation efficiency of SPHK1 in lung fibroblasts by decreasing the tRNA N7-methylguanosine modification and downregulating the expression of tRNAs in lung epithelial cell-derived exosomes after irradiation, as well as inhibiting the interaction of SPHK1 with the NAT10 protein in fibroblasts. Furthermore, exosomes derived from irradiated lung epithelial cells after RA intervention decreased the acetylation and cytoplasmic translocation of PFKFB3, suppressing the FMT process triggered by glycolysis, and ultimately decelerating the progression of RIPF.

overall_design: In this study, m7G-tRNA-seq and tRNA-seq analysis were conducted to identify the m7G modified tRNAs. Western blotting, immunohistochemistry, northwestern blotting, northern blotting, immunofluorescence, wound-healing assays and Edu experiments were performed to explore the molecular mechanism by which RA regulated fibroblast to myofibroblast transformation (FMT) by affecting the exosomes of lung epithelial cells. Ribo-seq and mRNA-seq analysis were used to explore the underlying target mRNA. Seahorse assays and immunoprecipitation were carried out to elucidate the effects of RA on glycolysis and FMT processes via the regulation of PFKFB3 acetylation.

first: Tingting

first: Jinglin

last: Mi

first: Zhechen

last: Ouyang

first: Xinling

last: Qin


title: scTCR-seq sequencing and analysis of fresh resected lung tissues from tuberculosis patients

city: Shanghai

first: Zilu

last: Wen

address: No. 2901, Caolang Road, Jinshan District, Shanghai

institute: SHANGHAI PUBLIC HEALTH CLINICAL CENTER

summary: Tuberculosis-affected lungs with chronic inflammation harbor abundant immunosuppressive immune cells but the nature of such inflammation is unclear. Dysfunction in T cell exhaustion, while implicated in chronic inflammatory diseases, remains unexplored in tuberculosis. Given that immunotherapy targeting exhaustion checkpoints exacerbates tuberculosis, we speculate that T cell exhaustion is dysfunctional in tuberculosis. Using integrated single-cell RNA sequencing and T cell receptor profiling we reported defects in exhaustion responses within inflamed tuberculosis-affected lungs. Tuberculosis lungs demonstrated significantly reduced levels of exhausted CD8+ T cells and exhibited diminished expression of exhaustion-related transcripts among clonally expanded CD4+ and CD8+ T cells. Additionally, clonal expansion of CD4+ and CD8+ T cells bearing T cell receptors specific for CMV was observed. Expanded CD8+ T cells expressed the cytolytic marker GZMK. Hence, inflamed tuberculosis-affected lungs displayed dysfunction in T cell exhaustion. Our findings likely hold implications for understanding the reactivation of tuberculosis observed in patients undergoing immunotherapy targeting the exhaustion checkpoint.

overall_design: Fresh resected human lung tissues with high 18F-FDG avidity were profiled with single-cell RNA sequencing.Using integrated single-cell RNA sequencing and T cell receptor profiling we reported defects in exhaustion responses within inflamed tuberculosis-affected lungs.

first: Lin

last: Wang

first: Hui

last: Ma

first: Ka-Wing

last: Wong

first: Yanzheng

last: Song


title: RNA sequencing of 31 pre-treatment, primary cervical cancer samples

city: St. Louis

first: Jin

last: Zhang

address: 4511 Forest Park Parkway

department: Radiation Oncology

summary: Investigate the transcriptomic differences between HPV-positive and HPV-negative primary cervical tumors.

overall_design: Whole transcriptome sequencing was performed on 31 pre-treatment, primary cervical cancer samples from cervical cancer patients treated at the Washington University School of Medicine. A minimum of 0.5µg of RNA with RIN > 7 was extracted for library preparation. Sequencing was performed using Illumina HiSeq 3000, 1x50nt, ~40 million reads per sample), and aligned to GRCh38 using STAR 2.7.0f.

first: Fiona

last: Ruiz

first: Julie

last: Schwarz

first: Matthew

last: Inkman

first: Kay

last: Jayachandran

first: Perry

middle: W

last: Grigsby

first: Christopher

last: Miller

first: Michael

last: McLellan


title: Rosmarinic acid alleviates radiation-induced pulmonary fibrosis by downregulating tRNA N7-Methylguanosine modification-regulated fibroblast to myofibroblast transition through the exosomes pathway [tRNA-Seq]

first: jinglin

last: mi

first: Tingting


title: The impact of  chemical modulator of cytoskeleton in murine CD8+ T cell.

city: China

first: 达瀚

last: 温

address: Dongdan Santiao No. 5

institute: Medicine Basic sciences, Peking Union Medical College

summary: T cells cytoskeleton is invovlved in the recognition of the antigen-presenting cells and target cells. So we used  chemical modulators of cytoskeleton (Blebbistatin) to treat murine CD8+ T cells. Then RNA-seq was performed. We found that Ble, compared with the control group, made  significantly changes in the mRNA levels of murine CD8+ T cells.

overall_design: Total RNA was isolated from murine T cells treated with Veh., Ble (2 µM) for 24 hr by using TRIzol-chloroform method. Then RNA-seq was performed by using those RNA.

first: Yuying

last: Liu


title: Inhibition of SUMOylation Induces Adaptive Anti-Tumor Immunity Against Pancreatic Cancer through Multiple Effects on the Tumor Microenvironment

city: New York

first: Hyojae

middle: James

last: Lee

state: NY

address: 101 6th Ave

institute: New York Genome Center

summary: Improvement of pancreatic ductal adenocarcinoma (PDAC) patients’ outcomes require exploration of novel therapeutic mechanisms. Thus far, most studies of PDAC therapies, including those inhibiting small ubiquitin-like modifications (SUMOylation), have focused on PDAC epithelial cell biology. The mechanisms by which SUMOylation impacts PDAC in the context of its tumor microenvironment (TME) are poorly understood. We used clinically relevant orthotopic PDAC mouse models to investigate the effect of SUMOylation inhibition using a specific, clinical stage compound, TAK-981. TAK-981 monotherapy improved survival in immunocompetent mice. However, in contrast to its inhibition of PDAC cell proliferation in vitro, the survival benefit of TAK-981 in vivo is dependent on the presence of T cells.  Induction of adaptive anti-tumor immunity is the dominant anti-tumor effect of SUMOylation inhibition in vivo.  To elucidate the mechanism of adaptive immunity, we performed comparative single cell transcriptomics analyses of murine and human PDAC tumors, including cancer cells, T cells, tumor associated macrophages (TAM) and cancer associated fibroblasts (CAF), in combination with immunofluorescence, immunohistochemistry, western blots and qPCR. Mechanistically, SUMOylation regulates T cell activation and differentiation.  In addition, TAK-981 changed gene expression in TAM, CAF and cancer cells that are likely to enhance the anti-tumor T cell response. These findings provide rationale for combination strategies to augment adaptive immune responses to PDAC that are necessary for durable disease control in PDAC patients.

overall_design: KPC46 tumors were harvested from mice that are either treated with vehicle (Cremophor) or TAK-981. The processed tumors were then analyzed using scRNA-seq

first: Suna

last: Erdem

first: Jayanth

last: Narayanan

first: Neranjan

last: Tharuka

first: Jorge

last: De La Torre Medina

first: Tianchen

last: Ren

first: Yixuan

last: Kuang

first: Tharindumala

last: Abeywardana

first: Kevin

last: Li

first: Allison

last: Berger

first: Andrew

last: Lowy

first: Rebekah

last: White

first: Yuan

last: Chen


title: Circadian Dysfunction in Skeletal Muscle Impairs Limb Perfusion and Muscle Regeneration in Peripheral Artery Disease [Myotube-RNAseq]

status: Public on Sep 01 2024

city: Chicago

first: Pei

last: Zhu

state: IL

address: 420 E. Superior St

department: Biochemistry and Molecular Genetics

institute: northwestern university

summary: Peripheral arterial disease (PAD), caused by atherosclerosis, leads to limb ischemia, muscle damage, and impaired mobility in the lower extremities. Recent studies suggest that circadian rhythm disruptions can hinder vascular repair during ischemia, but the specific tissues involved and the impact on muscle health remain unclear. This study investigates the role of the skeletal muscle circadian clock in muscle adaptation to ischemic stress using a surgical mouse model of hindlimb ischemia. We used mice with specific genetic loss of the circadian clock activator, BMAL1, in adult skeletal muscle tissues (Bmal1muscle). Bmal1muscle mice and controls underwent femoral artery ligation surgery to induce hindlimb ischemia. Laser doppler imaging was used to assess limb perfusion at various time points after the surgery. Muscle tissues were analyzed with RNA sequencing and histological examination to investigate PAD-related muscle pathologies. Additionally, we studied the role of BMAL1 in muscle fiber adaptation to hypoxia using RNA and ATAC sequencing analyses in primary myotube culture model. Disrupted expression of circadian rhythm-related genes was observed in existing RNA-seq datasets from PAD patient-derived endothelial cells and ischemic limb skeletal muscles. Genetic loss of Bmal1 specifically in adult mouse skeletal muscle tissues delayed reperfusion recovery following induction of hindlimb ischemia. Histological examination of muscle tissues showed reduced regenerated myofiber number and a decreased proportion of type IIB fast-twitch myofibers in Bmal1musc mouse muscles in the ischemic limbs, but not in their contralateral non-ischemic limbs. Transcriptomic analysis revealed abrogated metabolic, angiogenic, and myogenic pathways relevant to hypoxia-adaptation in Bmal1musc mouse muscles. These changes were corroborated in Bmal1-deficient cultured primary myotubes cultured under hypoxic conditions.

overall_design: We isolated primary myoblasts from Bmal1-floxed mice and induced the cells into myotubes. The myotubes were then infected with Cre-expressing adenovirus for 48 hours to induce deletion of the circadian clock activator, BMAL1. Myotubes infected with an empty vector-expressing virus were served as control. The viral infected myotubes were then exposed to 21% O2 or 1% O2 for 6 hours to simulate post-surgery intramuscular hypoxia.

first: Calvin

last: Chao

first: Adam

last: Steffeck

first: Caitlyn

last: Dang

first: Noah

middle: X

last: Hamlish

first: Eric

last: Pfrender

first: Bin

last: Jiang

first: Clara

middle: B

last: Peek


title: Polyomavirus ALTOs, but not MTs, downregulate viral early gene expression by activating the NF-κB pathway

city: Seattle

first: Denise

last: Galloway

state: WA

address: 1100 Fairview Ave N

department: Human Biology/Pathogen-Associated Malignancies IRC

institute: Fred Hutch Cancer Center

summary: Polyomaviruses are small, circular dsDNA viruses that can cause cancer. Alternative splicing of polyomavirus early transcripts generates large and small tumor antigens (LT, ST) that play essential roles in viral replication and tumorigenesis. Some polyomaviruses also express middle tumor antigens (MTs) or alternate LT open reading frames (ALTOs), which are evolutionarily related but have distinct gene structures. MTs are a splice variant of the early transcript whereas ALTOs are overprinted on the second exon of the LT transcript in an alternate reading frame and are translated via an alternative start codon. Merkel cell polyomavirus (MCPyV), the only human polyomavirus that causes cancer, encodes an ALTO but its role in the viral lifecycle and tumorigenesis has remained elusive. Here, we performed bulk RNA sequencing in MKL-2 cells, a Merkel cell carcinoma cell line, expressing MCPyV ALTO or RFP as a negative control via a doxycycline inducible lentiviral vector. The goal of the study was to identify Differentially Expressed Genes and pathways activated or inhibited via Gene Set Enrichment Analysis.

overall_design: MKL-2 cells were transduced with doxycycline inducible lentiviral vectors encoding Merkel cell polyomavirus ALTO or RFP as a negative control. Cells were selected with puromycin and then treated with doxycycline for 48 hours prior to harvest.

first: Nicholas

last: Salisbury


title: Autoantigen-specific CD4+ T cells acquire an exhausted phenotype and persist in human antigen-specific autoimmune diseases

city: Kiel

first: Seyed Mohammd Mahdi

last: Rasa

state: Schleswig-Holstein

address: Michaelisstrasse 5

department: Institut for Immunology

institute: Universitätsklinikum Schleswig-Holstein

summary: Pro-inflammatory autoantigen-specific CD4+ T helper (auto-Th) cells are central orchestrators of autoimmune diseases (AIDs). We aimed to characterize these cells in human AIDs with defined autoantigens by combining human leukocyte antigen (HLA)-tetramer-based and activation-based multidimensional ex vivo analyses. In aquaporin4-antibody-positive neuromyelitis optica spectrum disorder (AQP4-NMOSD) patients, auto-Th cells expressed CD154, but proliferative capacity and pro-inflammatory cytokines were strongly reduced. Instead, exhaustion-associated co-inhibitory receptors were expressed together with FOXP3, the canonical regulatory T cell (Treg) transcription factor. Auto-Th cells responded in vitro to checkpoint inhibition and provided potent B cell help. Cells with the same exhaustion-like (ThEx) phenotype were identified in soluble liver antigen (SLA)-antibody-autoimmune hepatitis and BP180-antibody-positive bullous pemphigoid, AIDs of the liver and skin, respectively. While originally described in cancer and chronic infection, our data point to T cell exhaustion as a common mechanism of adaptation to chronic (self-)stimulation across AID types and link exhausted CD4+ T cells to humoral autoimmune responses, with implications for therapeutic targeting.

overall_design:
For single-cell RNA-seq, PBMCs were stimulated for 6 h (with AQP4, MP65, and Influenza HA peptide pools or PMA/Ionomycin), CD154+ memory cells were isolated by MACS and further purified by FACS sorting on a FACS Aria Fusion (BD Bioscience, San Jose, CA, USA) based on dual expression of CD154 and CD69 and expression of CD45RO. Cells were directly sorted into skirted 96-well PCR plates which were equipped with 2 l lysis buffer (nuclease-free water with 0.2% Triton-X 100 solution (Roche Diagnostics GmbH, Mannheim, Germany) with 4 U/µL RNase Inhibitor (New England Biolabs)) in advance. After FACS sorting plates were shortly centrifuged (30 sec, 200xg) and immediately frozen at -80°C until shipment. Library preparation and sequencing was performed at the DRESDEN-concept Genome Center (DcGC, TU Dresden, Germany). The workflow was based on the previously described SMARTseq2 protocol 81. After thawing the samples, 2 ul of a primer mix was added (5 mM dNTP (Invitrogen), 0.5 uM dT-primer*, 4 U RNase Inhibitor (NEB)). RNA was denatured for 3 minutes at 72°C and the reverse transcription was performed at 42°C for 90 min after filling up to 10 ul with RT buffer mix for a final concentration of 1X superscript II buffer (Invitrogen), 1 M betaine, 5 mM DTT, 6 mM MgCl2, 1 uM TSO-primer*, 9 U RNase Inhibitor and 90 U Superscript II. After synthesis, the reverse transcriptase was inactivated at 70°C for 15 min. The cDNA was amplified using 2X Kapa HiFi HotStart Readymix (Roche) at a final 1X concentration and 0.1 uM UP-primer* under following cycling conditions: initial denaturation at 98°C for 3 min, 22 cycles [98°C 20 sec, 67°C 15 sec, 72°C 6 min] and final elongation at 72°C for 5 min. The amplified cDNA is purified using 0.6X volume of hydrophobic Sera-Mag Beads (Cytvia: GE24152105050250,) and DNA was eluted in 12 ul nuclease free water. The cDNA quality and concentration of a few representative samples was determined with the Fragment Analyzer (Agilent). For library preparation, 2 µl amplified cDNA was tagmented in 1X Tagmentation Buffer using 0.8 µl bead-linked transposome (Illumina DNA Prep, (M) Tagmentation, Illumina) at 55°C for 15 min in a total volume of 4 µL. The reaction was stopped by adding 1 µl of 0.1 % SDS (37°C, 15 min). Magnetic beads were bound to a magnet, the supernatant was removed, beads were resuspended in 4 µl indexing PCR Mix containing 1x KAPA Hifi HotStart Ready Mix (Roche). After adding 700 nM unique dual indexing primers (i5 and i7) using Echo Acoustic Liquid Handler (Echo 525, Beckman Coulter), the reaction mixture was subjected to a PCR (72°C 3 min, 98°C 30 sec, 12 cycles [98°C 10 sec, 63°C 20 sec, 72°C 1 min], 72°C 5 min). After PCR, libraries are quantified with a Tecan plate reader Infinite 200 pro in 384 well black flat bottom low volume plates (Corning) using AccuBlue Broad range chemistry (Biotium) and equimolarly pooled using Echo Acoustic Liquid Handler (Echo 525, Beckman Coulter GmbH).  The pooled libraries were then purified with 0.9x volume Sera-Mag SpeedBeads, followed by a double size selection with 0.6x and 0.9x volume of beads. Re-double size selection with 0.6x and 0.9x volume of beads was performed if needed. Sequencing was performed after quantification using a Fragment Analyzer on an Illumina Novaseq 6000 aiming at an average sequencing depth of 0.5 mio 100 bp paired-end fragments per cell.

The method used for this single cell RNA seq is SMARTseq2 which is the full length transcriptome sequencing, thus the TCR sequences come from the same library as the expression data. However, always in some random cells, the full length of TCR alpha/beta or both are not sequenced, so it is not possible to recognize the TCR sequence, but still the expression data is available for these cells. This is the reason why fewer cells in the TCR file compare to the expression matrix (i.e. 928 samples vs. 924 columns in RawExpressionCountTable.csv, 829 data rows in TCR_data.csv)

first: Carina

last: Saggau

first: Petra

last: Bacher

first: Daniela

last: Esser

first: Mahdi

first: Silja

last: Meise

first: Sinje

last: Leitzke

first: Andreas

last: Hutloff

first: Sarah-Sophie

last: Schacht

first: Justina

last: Dargvainiene

first: Klarissa

middle: H

last: Stürner

first: Ina

last: Schröder

first: Robert

last: Markewitz

first: Maria

last: Hastermann

first: Ankelien

last: Duchow

first: Patrick

last: Schindler

first: Jörn

last: Walter

first: Judith

last: Gottfreund

first: Julia

last: KPolansky

first: Mingxing

last: Yang

first: Reza

last: Naghavian

first: Mareike

last: Wendorff

first: Ev-Marie

last: Schuster

last: Dahl

last: Petzold

first: Susanne

last: Reinhardt

first: Andre

last: Franke

first: Marek

last: Wieczorek

first: Lea

last: Henschel

first: Daniel

first: Simon

last: Fillatreau

first: Dirk

last: Busch

first: Klaus-Peter

last: Wandinger

first: Kilian

last: Schober

first: Roland

last: Martin

first: Friedemann

last: Paul

first: Frank

last: Leypoldt

first: Alexander

last: Scheffold


title: RNA Expression Profiling in Lymphoblastoid Cell Lines from Mutated and Non-Mutated Amyotrophic Lateral Sclerosis Patients

city: Pavia

first: Jessica

last: Garau

address: Via Mondino 2

institute: IRCCS Mondino Foundation

summary: Amyotrophic Lateral Sclerosis (ALS) is a neurodegenerative disease characterized by the death of upper and lower motor neurons with unknown etiology. The difficulty to recover biological material from patients led to employ lymphoblastoid cell lines (LCLs) as a model for ALS, since many pathways, typically located in neurons, are also activated in these cells. To investigate the expression of coding and long noncoding RNAs in LCLs, a transcriptomic profiling of Sporadic ALS (SALS) and mutated patients (FUS, TARDBP, C9ORF72 and SOD1), and matched controls was realized. Thus, Differentially Expressed Genes (DEGs) were investigated among the different subgroups of patients. Peripheral Blood Mononuclear Cells (PBMCs) were isolated and immortalized into LCLs via Epstein-Barr Virus (EBV) infection, RNA was extracted and RNA-sequencing analysis was performed. Gene expression profiles of LCLs were genetic-background specific, indeed only 12 genes were commonly deregulated in all groups. Nonetheless, pathways enriched by DEGs in each group were also compared and a total of 89 KEGG terms were shared among all patients. Eventually, the similarity of affected pathways was also assessed when our data were matched with a transcriptomic profile realized in PBMCs of the same patients. Thus, we concluded that LCLs are a good model for the study of RNA deregulation in ALS.

overall_design: PBMCs were isolated from peripheral venous blood samples. Lymphocytes B were isolated from an aliquot of PBMCs and were immortalized into LCLs via Epstein-Barr Virus (EBV) infection. Samples were homogenized and total RNA was isolated by Trizol® reagent. Sequencing libraries were prepared by the Illumina TruSeq RNA Library Prep, version 2, Protocol D, using 500-ng total RNA (Illumina, California, USA). FastQ files were generated using the llumina bcl2fastq2 software (Version 2.17.1.14 -http://support.illumina.com/downloads/bcl2fastq-conversion-software-v217.html) from raw sequencing reads gener-ated by the Illumina NextSeq 500 sequencer (Illumina, California, USA). STAR/RSEM software was used to cal-culate gene and transcript intensities using Gencode Release 19 (GRCh37.p13) as a reference and the "stranded" op-tion. The mRNA differential expression analysis was carried out using the R package EBSeq. The R package DESeq2 was used to perform differential expression analysis on lncRNAs. With |log2(disease sample/healthy con-trol)|1 and an FDR of 0.1, genes were considered differentially expressed and retained for further analysis.

first: Maria

last: Garofalo

first: Francesca

last: Dragoni

first: Eveljn

last: Scarian

first: Rosalinda

last: Di Gerlando

first: Luca

last: Diamanti

first: Susanna

last: Zucca

first: Stella

last: Gagliardi

first: Matteo

last: Bordoni

first: Orietta

last: Pansarasa


title: Phospholipase D2 downregulates interleukin-1β secretion from tumor-associated macrophages to suppress bladder cancer progression

city: Tsukuba

first: Kazuki

last: Hamada

state: Ibaraki

address: 1-1-1 Tennodai

country: Japan

institute: University of Tsukuba

summary: The tumor microenvironment (TME) modulates therapeutic response and prognosis in patients with bladder cancer (BC). The roles of two phospholipase D (PLD) isoforms, PLD1 and PLD2 (hydrolysis of phosphatidylcholine to phosphatidic acid) in cancer cells have been well-studied in numerous cancer types, but their roles in the TME remain unclear. We used a mouse BC Pld2-knockout carcinogenesis model and global transcriptomic analysis to reveal that PLD2 was significantly involved in BC progression through immunosuppressive pathways in the TME. We therefore focused on PLD2 and tumor-associated macrophages (TAMs), which were increased in Pld2-knockout mice and further associated with poor prognoses in BC patients. In vitro, we found that Pld2- knockout mouse TAMs had significantly enhanced proliferation, correlating closely with increased IL-1β production. These results indicate that PLD2 suppresses BC progression by regulation of IL-1β secretion from TAMs in the TME, suggesting that PLD2 could serve as a potential therapeutic target for BC.

overall_design: To induce bladder cancer in mice, female 6-8-week-old WT, Pld1-/- and Pld2-/- mice were given drinking water with 0.025% N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) (Tokyo Kasei Kogyo).Total RNA from extracted whole bladders was isolated using a NucleoSpin RNA kit (TaKaRa).Total RNA was extracted from sorted tumor-associated macrophages using TRIzol reagent (Invitrogen).For RNA sequencing, 500 ng total RNA were rRNA-depleted using the NEBNext rRNA Depletion Kit (New England Biolabs) according to the manufacturer’s instructions. Library preparation was performed using an NEBNext Ultra Directional RNA Library Prep Kit (New England Biolabs). Libraries were validated via Bioanalyzer (Agilent Technologies) to determine size distribution and concentration. Validated libraries were then sequenced at Tsukuba i-Laboratory LLP (Tsukuba, Japan) on a NextSeq500 (Illumina) with the paired-end 36-base read option.Sequencing reads were mapped to the mm10 mouse reference genome, quantified, and then normalized with the quantile method using CLC Genomics Workbench version 10.1.1 (Qiagen).


title: Endolysosomal dysfunction in radial glia progenitor cells leads to defective cerebral angiogenesis and compromised blood-brain barrier integrity

city: Rehovot

first: Karina

last: Yaniv

state: Israel

address: Hertzel 234

department: Immunology and Regenerative Biology

country: Israel

institute: Weizmann Institute of Science

summary: The neurovascular unit (NVU) is a complex multicellular structure that helps maintain cerebral homeostasis and blood-brain barrier (BBB) integrity.  While extensive evidence links NVU alterations to cerebrovascular diseases and neurodegeneration, the underlying molecular mechanisms remain unclear.  Here, we use zebrafish embryos carrying a mutation in Scavenger Receptor B2, a highly conserved endolysosomal protein expressed predominantly in Radial Glia Cells (RGCs), to investigate the interplay among different NVU components. Through live imaging and genetic manipulations, we demonstrate that compromised acidification of the endolysosomal compartment in mutant RGCs leads to impaired Notch3 signaling, thereby inducing excessive neurogenesis and reduced glial differentiation. We further demonstrate that alterations to the neuron/glia balance result in impaired VEGF and Wnt signaling, leading to severe vascular defects, hemorrhages, and a leaky BBB. Altogether, our findings provide novel insights into NVU formation and function and offer new avenues for investigating diseases involving white matter defects and vascular abnormalities. The neurovascular unit (NVU) is a complex multicellular structure that helps maintain cerebral homeostasis and blood-brain barrier (BBB) integrity.  While extensive evidence links NVU alterations to cerebrovascular diseases and neurodegeneration, the underlying molecular mechanisms remain unclear.  Here, we use zebrafish embryos carrying a mutation in Scavenger Receptor B2, a highly conserved endolysosomal protein expressed predominantly in Radial Glia Cells (RGCs), to investigate the interplay among different NVU components. Through live imaging and genetic manipulations, we demonstrate that compromised acidification of the endolysosomal compartment in mutant RGCs leads to impaired Notch3 signaling, thereby inducing excessive neurogenesis and reduced glial differentiation. We further demonstrate that alterations to the neuron/glia balance result in impaired VEGF and Wnt signaling, leading to severe vascular defects, hemorrhages, and a leaky BBB. Altogether, our findings provide novel insights into NVU formation and function and offer new avenues for investigating diseases involving white matter defects and vascular abnormalities. The neurovascular unit (NVU) is a complex multicellular structure that helps maintain cerebral homeostasis and blood-brain barrier (BBB) integrity.  While extensive evidence links NVU alterations to cerebrovascular diseases and neurodegeneration, the underlying molecular mechanisms remain unclear.  Here, we use zebrafish embryos carrying a mutation in Scavenger Receptor B2, a highly conserved endolysosomal protein expressed predominantly in Radial Glia Cells (RGCs), to investigate the interplay among different NVU components. Through live imaging and genetic manipulations, we demonstrate that compromised acidification of the endolysosomal compartment in mutant RGCs leads to impaired Notch3 signaling, thereby inducing excessive neurogenesis and reduced glial differentiation. We further demonstrate that alterations to the neuron/glia balance result in impaired VEGF and Wnt signaling, leading to severe vascular defects, hemorrhages, and a leaky BBB. Altogether, our findings provide novel insights into NVU formation and function and offer new avenues for investigating diseases involving white matter defects and vascular abnormalities.

overall_design: 50 heads (3 biological replicates) was dissected from euthanized 48 hpf scarb2a mutants and wt siblings and dissociated into a single-cell suspension . Sorting and RNA extraction was performed on TgBAC(scar2ba:KalTA4; UAS-mKate2)

first: Ivan

last: Bassi

first: Moshe

last: Grunspan

first: Gideon

last: Hen

first: Kishore

last: Ravichandran

first: Noga

last: Moshe

first: Laura

last: Gutierrez-Miranda

first: Stav

last: Safriel

first: Daria

last: Kostina

first: Amitay

last: Shen

first: Carmen

last: Ruiz de Almodovar


title: KAS-ATAC reveals the genome-wide single-stranded accessible chromatin landscape of the human genome

city: Stanford

first: Georgi

middle: Kolev

last: Marinov

state: California

address: 279 Campus Drive West, Beckman Center, B-257A/259

department: Genetics

institute: STANFORD UNIVERSITY

summary: Gene regulation in most eukaryotes involves two fundamental physical processes -- alterations in the packaging of the genome by nucleosomes, with active cis-regulatory elements (CREs) generally characterized by an open-chromatin configuration, and the activation of transcription. Mapping these physical properties and biochemical activities genome-wide -- through profiling chromatin accessibility and active transcription -- have now long been key tools used to understand the logic and mechanisms of transcription and its regulation. However, the direct relationship between the two has not been accessible to measurements, leaving the exact chromatin state of actively transcribed DNA molecules unresolved. To address this question, we developed KAS-ATAC, a combination of the KAS-seq (Kethoxal-Assisted SsDNA sequencing and ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) methods for mapping single-stranded DNA (and thus active transcription) and chromatin accessibility, respectively, which maps DNA fragments that are simultaneously accessible and containing ssDNA genome-wide. We use KAS-ATAC to evaluate levels of active transcription over different classes of regulatory elements in the human genome, to estimate the absolute levels of transcribed accessible DNA over CREs, to map the nucleosomal configurations associated with RNA polymerase activities, and to assess transcription factor association with transcribed DNA through transcription factor binding site (TFBS) footprinting.

overall_design: KAS-ATAC in human cell lines

middle: K

first: Samuel

last: Kim

first: Anshul

last: Kundaje

first: William

last: Greenleaf


title: B cell differentiation into germinal center and antibody-secreting cells depends on SMARCA5-mediated chromatin remodeling[Ribotag]

first: Ziv

last: Shulman

address: 234 Herzl st.

department: Department of Immunology

summary: The establishment of long-lasting immunity against pathogens is facilitated by the germinal center (GC) reaction, during which B cells increase their antibody affinity, and differentiate into antibody-secreting cells (ASC) and memory cells. These events involve modifications in chromatin packaging that orchestrate the profound restructuring of gene expression networks that determine cell fate. While several chromatin remodelers were implicated in lymphocyte functions, less is known about SMARCA5. Here, by using ribosomal pull-down for analyzing translated genes in GC B cells, coupled with functional experiments in mice, we identified SMARACA5 as a key chromatin remodeler in B cells. While the naive B cell compartment and early B cell activation remained unaffected following conditional depletion of Smarca5, immunoglobulin class switching, GC formation and ASC differentiation were impaired. Single-cell multi-omic sequencing analyses revealed that SMARCA5 is crucial for facilitating the transcriptional modifications and genomic accessibility of genes that support B cell differentiation. These findings offer novel insights into SMARCA5, which can be targeted in lymphoma patients.

overall_design: Bulk RNA analysis of Ribotag dark and light zone B cells 7 days after NP-KLH immunization

first: Liat

last: Stoler-Barak

first: Dominik

last: Schmiedel

first: Avital

last: Sarusi-Portuguez

first: Ronnie

last: Blecher-Gonen

first: Zhana

last: Haimon

first: Tomas

last: Stopka


title: Gene expression profile of intestinal organoids from people with cystic fibrosis upon exposure to elexacaftor/tezacaftor/ivacaftor

city: Prague 5

first: Ondrej

last: Cinek

address: V Uvalu 84

department: Dept of Medical Microbiology

country: Czech Republic

institute: Charles University

summary: Background: The forskolin-induced swelling assay (FIS) in intestinal organoids (IOs), used to determine in vitro responsiveness to elexacaftor/tezacaftor/ivacaftor (ETI), showed variability in swelling among IOs obtained from people with CF (pwCF) carrying the same genotype. We aimed to characterise the effect of ETI on the transcriptional activity of IOs-derived cells to understand the intracellular processes triggered by ETI and the differences in treatment response. Methods: Six high- and six low-responding IOs to ETI, derived from F508del/F508del pwCF, were incubated with or without ETI for 2 to 6 hours. Gene expression was modelled using negative binomial models adjusted for subjects, having time and incubation with ETI as predictors.  Results: Incubation with ETI resulted in a significant upregulation of several biological processes: mostly related to chemokines and signalling, chemotaxis, and tissue development processes. No changes were observed in abundance of the CFTR transcripts or in CFTR-related gene sets and pathways. The genes and pathways associated with ETI did not overlap with those whose expression changed with time only. IOs with a high FIS response did not significantly differ in any interpretable gene from the FIS-low organoids. Conclusions: The changes in the IOs gene expression upon the exposure to ETI cannot explain differences in the magnitude of IOs FIS-measured response to ETI. Genes of the CFTR-related pathways do not change their transcriptional activity; instead, overexpression was observed in genes of inflammatory-like cytokine response and receptor activation pathways, seemingly unrelated to the mechanisms of FIS, but possibly related to the effect of ETI itself.

overall_design: The present study aimed to investigate the effect of ETI on the overall gene expression profile, in particular, on the transcriptional activity of CFTR and CF-modifying genes. The inclusion of organoids at opposite ends of the FIS response spectrum also allowed us to assess whether gene expression changes are related to the magnitude of the organoid response to ETI. Briefly, each IOs culture was split into seven culture wells, then three wells were incubated with ETI medium, while 4 control wells were incubated in ETI-free medium. One of the control wells was immediately harvested at 0 hours (baseline). Three pairs of ETI-free and ETI-containing cultures were harvested at 2, 4 and 6 hours. In total, 12 IOs from 12 pwCF (F508del homozygotes) were included, making 12 biological replicates. Of note, six IOs had the best and six IOs had the worst response to ETI when tested by the FIS.

first: Eva

last: Furstova

first: Stepanka

last: Novotna

first: Klara

last: Hubackova

first: Tereza

last: Dousova

first: Lucie

last: Borek-Dohalska

first: Pavel

last: Drevinek


title: DIISCO: A Bayesian framework for inferring dynamic intercellular interactions from time-series single-cell data

city: Boston

first: Alexandria

last: Maurer

address: 450 Brookline Ave

department: Department of Medical Oncology

institute: Dana-Farber Cancer Institute

summary: Characterizing cell-cell communication and tracking its variability over time is essential for understanding the coordination of biological processes mediating normal development, progression of disease, or responses to perturbations such as therapies. Existing tools lack the ability to capture time-dependent intercellular interactions, such as those influenced by therapy, and primarily rely on existing databases compiled from limited contexts. We present DIISCO, a Bayesian framework for characterizing the temporal dynamics of cellular interactions using single-cell RNA-sequencing data from multiple time points. Our method uses structured Gaussian process regression to unveil time-resolved interactions among diverse cell types according to their co-evolution and incorporates prior knowledge of receptor-ligand complexes. We show the interpretability of DIISCO in simulated data and new data collected from CAR-T cells co-cultured with lymphoma cells, demonstrating its potential to uncover dynamic cell-cell crosstalk.

overall_design: single-cell RNA sequencing from 4 replicates of co-cultured CAR-T cells with MEC1 cancer cells, a CLL cell line

first: Cameron

middle: Y

last: Park

first: Shouvik

last: Mani

first: Nicolas

last: Beltran-Velez

first: Katie

first: Teddy

last: Huang

first: Shuqiang

last: Li

first: Satyen

last: Gohil

first: Ken

last: Livak

first: David

middle: A

last: Knowles

first: Catherine

middle: J

last: Wu

first: Elham

last: Azizi


title: Strain-specific quorum-sensing responses determine virulence properties in Vibrio anguillarum

city: Helsingør

first: Mathias

last: Middelboe

address: Strandpromenaden 5

country: Denmark

institute: University of Copenhagen

summary: Here, we define the VanT-QS regulon and explore the diversity and trajectory of traits under QS regulation in the fish pathogen Vibrio anguillarum through comparative transcriptomics of two wildtype strains and their corresponding mutants artificially locked in QS-on (ΔvanO) or QS-off (ΔvanT) states. We reveal that populations of one strain primarily employ a QS-off response, while the other strain mainly maneuvers within the QS-on spectrum. Further examination of our V. anguillarum collection revealed that ~15 % of the strains were QS-negative and measurements of a total of 9 QS-positive strains suggested diverse QS responses, which underlines an extensive diversity of QS delegations. We show that QS control a plethora of genes involved in processes such as central metabolism, biofilm, competence, T6SS and virulence in V. anguillarum. Interestingly, the low QS response strain had an enlarged QS regulon compared to the high QS response strain, suggesting a potential trade-off between engagement in QS and output of the commitment. Moreover, the combination of present and previous virulence trials coupled with QS response measurements indicate that the QS state is an important driver of virulence toward fish larvae in V. anguillarum. We propose that infections by mixed-strain communities spanning diverse QS strategies might utilize the animal host more efficiently. Furthermore, we emphasize the importance of applying whole-cell spike-in normalization to study gene expression of cell populations with differential total RNA content per cell.

overall_design: mRNA profiles of 2 different wildtype Vibrio anguillarum strains and their corresponding ΔvanT and ΔvanO mutants sampled at LCD (OD~0.1) and HCD (OD~1.8) in biological triplicates (total n = 36). All samples were spiked-in with 1 % strain 90-11-287 carrying plasmid pEVS143 with IPTG-inducible gfp and cmr

first: Jesper

last: Mauritzen

first: Emilie

last: Søndberg

first: Panos

middle: G

last: Kalatzis

first: Line

last: Roager

first: Lone

last: Gram

first: Sine

middle: L

last: Svenningsen


title: B cell differentiation into germinal center and antibody-secreting cells depends on SMARCA5-mediated chromatin remodeling

first: Ziv

last: Shulman

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

overall_design: Refer to individual Series


title: B cell differentiation into germinal center and antibody-secreting cells depends on SMARCA5-mediated chromatin remodeling[scMultiome - RNA + ATAC]

overall_design: Multiome analysis of control or CD23.Cre.Smarca5fl/fl B1-8hi GFP+ B cells adoptively transferred to WT mice, 5 days after NP-KLH immunization

first: Liat

last: Stoler-Barak

first: Dominik

last: Schmiedel

first: Avital

last: Sarusi-Portuguez

first: Ronnie

last: Blecher-Gonen

first: Zhana

last: Haimon

first: Tomas

last: Stopka


title: Transcriptomics profiling of white adipose tissue of wildtype (WT) and miR-10a knockout (KO) mice fed a normal chow diet (NCD) or high-fat diet (HFD)

city: Seoul, Republic of Korea

first: Sumin

last: Lee

address: 1, Gwanak-ro, Gwanak-gu

country: South Korea

institute: Seoul National University

summary: Adipose tissue remodeling is a dynamic process that involves adipocyte death and turnover. This study investigates the influence of miR-10a-5p on the remodeling of adipose tissue and the metabolic dysfunction prompted by a high-fat diet. Our objective involves examining the impact of a 16-week HFD feeding on both WT mice and miR-10a KO mice, comparing the genes that are differentially regulated based on the dietary intake.

overall_design: White adipose tissue mRNA profiles of WT and miR-10a KO mice fed a NCD or HFD were generated by deep sequencing. Three biological replicates were analyzed for each of conditions.

first: Yun-Hee

first: YoonKeun

last: Cho


title: B cell differentiation into germinal center and antibody-secreting cells depends on SMARCA5-mediated chromatin remodeling[RNA-Seq]

overall_design: Bulk RNA analysis of control or CD23.Cre.Smarca5fl/fl B1-8hi GFP+ B cells adoptively transferred to WT mice, 5 days after NP-KLH immunization


title: Extracellular vesicles are involved in the paracrine communication between epithelial cells in different regions of the domestic cat epididymis

city: Washington

first: Danielle

last: Sosnicki

state: DC

address: 3001 Connecticut Ave NW

institute: Smithsonian's National Zoo and Conservation Biology Institute

summary: Sperm maturation depends on exposure to specific microenvironments within the different segments of the epididymis, but mechanisms underlying how these microenvironments are produced or maintained are not well understood. We hypothesize that epididymal extracellular vesicles (EVs) could play a role in this process, and therefore set out to test whether the EVs from different regions of the epididymis can serve as a form of paracrine communication between epithelial cells. Domestic cat tissues were used to develop a reproducible in vitro culture system for corpus epididymis explants that were then exposed to EVs collected from upstream (i.e., caput) segments. The impacts of different culture or exposure conditions were compared by analyzing the morphology, apoptosis, transcriptional activity, and gene expression in the explants. Here, we report the development of the first in vitro culture system for epididymal tissue explants in the domestic cat model. Using this system, we found that EVs from the caput segment have a significant effect on the transcriptional profile of tissue from the corpus segment (1,233 differentially expressed genes due to EV supplementation). Of note, expression of genes associated with regulation of epithelial cell differentiation and cytokine signaling in the epididymis were regulated by the presence of EVs. Together, our findings comprise the first report of paracrine control of segmental gene regulation by epididymal EVs in any species. These results contribute to a better understanding of epididymis biology and could lead to techniques to enhance or suppress male fertility.

overall_design: To determine the effect of caput EVs on the transcriptome of corpus epididymis explants, we cultured corpus explants with and without EV supplementation and also included fresh tissue controls. RNA was extracted, and sequenced.

middle: M

first: Alexander

last: Travis

first: Pierre

last: Comizzoli


title: B cell differentiation into germinal center and antibody-secreting cells depends on SMARCA5-mediated chromatin remodeling[CUT & RUN]

overall_design: CUT&RUN analysis of WT GC B cells from inguinal lymph nodes 7 days after immunization with KLH+CFA


title: Epigenome reprogramming through H3K27 and H3K4 trimethylation as a resistance mechanism to DNA methylation inhibition in BRAFV600E-mutated CRC. [RNA-Seq]

city: HOUSTON

first: Ajay

last: Saw

address: 7900 Cambdrige St

institute: University Of Texas MD Anderson Cancer Center, Houston, Texas, USA.

summary: BRAFV600E-mutated colorectal cancer (CRC) exhibits a strong correlation with DNA hypermethylation suggesting this subgroup of tumors is uniquely presenting epigenomic phenotypes. Nonetheless, the traditional epigenomic therapeutic agent, 5-azacitidine, which inhibits DNA methyltransferase activity, did not yield sufficient improvements in the efficacies of BRAFV600E CRC in vivo. We utilized a patient-derived xenograft model and confirmed an effective reduction of DNA methylation levels upon 5-azacitidine treatment yet failed to restore gene expression patterns. This study unbiasedly explored the adaptive engagement of other epigenomic modifications upon the profound decrease in DNA methylation by azacitidine treatment. A loss of histone acetylation and a gain of histone methylations, including H3K27 and H3K4 trimethylation, were observed around hypomethylated regions. Our findings suggested a compensatory increase in repressive histone mark, H3K27 trimethylation, around treatment-induced hypomethylated regions, which suggests the involvement of polycomb repressive complex (PRC) activity around the genome with lost DNA methylation, therefore maintaining the suppression of key genes. Combined inhibition of PRC activity through EZH2 inhibitor with azacitidine treatment additively improved efficacies in BRAFV600E CRC cells. In conclusion, DNA hypermethylation exhibits a close association with H3K27me3 and PRC activity in BRAFV600E CRC, and simultaneous blockade of DNMT and EZH2 holds promise as a potential therapeutic strategy for patients with BRAFV600E-mutated CRC.

overall_design: To investigate the gene expression changes by 5-azacitidine treatment in BRAFV600E mutant CRC PDX model, mice were treated with either vehicle or 5-azacitidine.

first: Hey

first: Scott

last: Kopetz

first: Kunal

last: Rai


title: Rosmarinic acid alleviates radiation-induced pulmonary fibrosis by downregulating tRNA N7-Methylguanosine modification-regulated fibroblast to myofibroblast transition through the exosomes pathway [mRNA-Seq]

city: Nanning

first: jinglin

last: mi

address: shuangyong road

institute: the first affiliated hospital of guangxi medical university

summary: Radiation-induced pulmonary fibrosis (RIPF) is a common complication after radiotherapy in thoracic cancer patients, and there is a lack of effective treatment methods. The aim of this study was to explore the protective effect of rosmarinic acid (RA) on RIPF in mice as well as the underlying mechanism.We found that RA exerted an antifibrotic effect on lung tissues of RIPF mouse models and inhibited the progression of FMT through exosomes derived from lung epithelial cells. Mechanistically, RA reduced the transcription and translation efficiency of SPHK1 in lung fibroblasts by decreasing the tRNA N7-methylguanosine modification and downregulating the expression of tRNAs in lung epithelial cell-derived exosomes after irradiation, as well as inhibiting the interaction of SPHK1 with the NAT10 protein in fibroblasts. Furthermore, exosomes derived from irradiated lung epithelial cells after RA intervention decreased the acetylation and cytoplasmic translocation of PFKFB3, suppressing the FMT process triggered by glycolysis, and ultimately decelerating the progression of RIPF.

overall_design: In this study, m7G-tRNA-seq and tRNA-seq analysis were conducted to identify the m7G modified tRNAs. Western blotting, immunohistochemistry, northwestern blotting, northern blotting, immunofluorescence, wound-healing assays and Edu experiments were performed to explore the molecular mechanism by which RA regulated fibroblast to myofibroblast transformation (FMT) by affecting the exosomes of lung epithelial cells. Ribo-seq and mRNA-seq analysis were used to explore the underlying target mRNA. Seahorse assays and immunoprecipitation were carried out to elucidate the effects of RA on glycolysis and FMT processes via the regulation of PFKFB3 acetylation.

first: Tingting

last: Zhang

first: Jinglin

last: Mi

first: Zhechen

last: Ouyang

first: Xinling

last: Qin


title: Activating mutations remodeled the chromatin accessibility landscape to drive distinct regulatory networks in KMT2A-rearranged acute leukemia

city: Lund

first: Anna

last: Hagström-Andersson

address: BMC C13

country: Sweden

institute: Lund University

summary: Activating FLT3 and RAS mutations are common in KMT2A-rearrangement (KMT2A-r) leukemia, but their functions in remodeling the chromatin accessibility is unknow. Using a retroviral acute myeloid leukemia (AML) mouse model driven by KMT2A::MLLT3, we show that FLT3/ITD, FLT3/N676K, and NRAS/G12D rewired the chromatin accessibility landscape and associated transcriptional networks. FLT3/N676K and NRAS/G12D mutations resulted in similar networks which were distinct from those mediated by FLT3/ITD. Motif analysis revealed that the AP-1 family transcription factors had a role in KMT2A::MLLT3 leukemia with FLT3/N676K and NRAS/G12D, whereas RUNX1/2 and Stat5a/Stat5b were active in the presence of FLT3/ITD. Further, transcriptional programs related with stemness, activation of T-cells and negative regulation of NK-cells were activated in the presence of NRAS/G12D and FLT3/N676K.  Taken together, activating mutations established mutation-specific changes in the epigenetic landscape that led to changes in the transcriptional programs driven by specific transcription factors.

overall_design: 15 mice with acute myeloid leukemia (AML) driven by KMT2A::MLLT3-mCherry and either of FLT3/ITD-GFP (n=4), FLT3/N676K-GFP (n=4), NRAS/G12D-GFP (n=3), or control vector (n=4).

first: Qirui

first: Ton

last: Falqués-Costa

first: Mattias

last: Pilheden

last: Hagström


title: m6a modifications following morphine exposure or Alkbh5 siRNA in primary cortical neuron cultures

city: Philadelphia

first: Stephanie

last: Daws

state: PA

address: 3500 N Broad St

institute: Temple University

summary: m6a modified RNA and unmodified RNA were quantified in primary neuronal cultures following either siRNA knockdown of Alkbh5 or chronic morphine exposure for 3 days. The goal was to identify mRNA that have m6A modifications following one or more treatment conditions.

overall_design:
4 biological replicates were analyzed per treatment group in a two-color experiment. Cy3 labeling represents unmodified RNA and C5 represents RNA with m6A modification.
Please note that two channel arrays were used for these samples but the data were analyzed as single-channel so that each sample represents either IP or supernatent and the sample table includes the normalized signal values. Each raw data file is linked to the corresponding IP sample records.


title: Epigenome reprogramming through H3K27 and H3K4 trimethylation as a resistance mechanism to DNA methylation inhibition in BRAFV600E-mutated CRC [ATAC-seq]

overall_design: The assay for transposase-accessible chromatin with sequencing (ATAC-seq) was performed on BRAFV600E-mutated CRC patient-derived xenograft models.

last: Lee


title: Sdc4 deletion perturbs intervertebral disc matrix homeostasis and promotes early osteopenia in the aging mouse spine

first: Makarand

middle: V

last: Risbud

address: 1025 Walnut St., Suite 511 College Building

department: Orthopedic Surgery

institute: Thomas Jefferson University

summary:
Transcriptional analysis of 6-month-old primary nucleus pulposus (NP) mouse disc tissues from wild-type (WT) and SDC4 KO mice, which harbor deletions of exon 2 to part of exon 5 that endcodes the N-terminal coding region. Breeder mice were provided by Dr. Michael Simons.
SDC4's regulation of cytokine-dependent activation of the aggrecanse ADAMTS-5 is linked with inflammation-associated intervertebral disc degeneration. In this study, we aim to establish the role of SDC4 in matrix homeostasis under the context of aging.

overall_design: Tissues from the nucleus pulposus (NP)  were harvested from the intervertebral discs of four WT and four SDC4 KO mice at 6 months of age. NP and AF tissues were collected from lumbar discs in each animal for RNA extraction and hybridization using Affymetrix microarray.

first: Kimheak

last: Sao


title: Transcriptomics of impaired hypoxic recovery of L2HGDH mutants

city: Bloomington

first: Jason

last: Tennessen

email: tennessenlab@gmail.com, tennlab@iu.edu, jtenness@iu.edu, tourigny@iu.edu

state: IN

address: 1001 E 3rd St

department: Biology

institute: Indiana University

summary: We found that L2HGDH mutants, in addition to accumulating L-2HG, are profoundly sensitive to hypoxic conditions (1% O2), in contrast to normal or heterozygote control flies which can survive and recover from hypoxia even after 12 hours of exposure.  Here, we sought to investigate the trascriptomic changes associated with the response to hypoxia and recovery in both control and mutant backgrounds, with emphasis on the contrast between the normal and defective response at each time point.

overall_design: Control (heterozygous: L2hgdh[12]/[+]) and mutant (L2hgdh[12]/[14])  adult flies were subjected to hypoxia (1% O2, 12h), then collected during a timecourse of normoxic recovery (0h, 1h, 12h) for RNA extraction and RNA-seq analysis.

first: Nader

last: Mahmoudzadeh

middle: P

last: Toiurigny


title: Epigenome reprogramming through H3K27 and H3K4 trimethylation as a resistance mechanism to DNA methylation inhibition in BRAFV600E-mutated CRC.

first: Ajay

middle: Kumar

last: Saw


title: Gene expression differences associated with intrinsic hindfoot muscle loss in the jerboa, Jaculus jaculus

city: Salt Lake City

first: Mai

last: Tran

state: UT

address: 201 President Circle

institute: University of Utah

summary: Muscle is a highly adaptable tissue capable of undergoing environmentally-induced changes and remarkable evolutionary adaptations. Muscle atrophy, a common feature in various myopathies as well as other conditions like cancer and diabetes, has been studied at the transcriptomic level, uncovering shared molecular factors. The lesser Egyptian jerboa, adapted for bipedal locomotion in a desert environment, experiences natural muscle loss in its foot. Cellular characterization of this process revealed similarities with human skeletal muscle atrophy, such as the loss of Desmin and upregulation of the ubiquitin proteasome pathway. By using RNA-sequencing, gene candidates associated with muscle loss in jerboa were identified, and genome-wide comparisons with established mouse models of muscle dystrophy/atrophy elucidated shared molecular signatures. These findings provide valuable insights into the genes and pathways involved in natural muscle loss and contribute to understanding the evolutionary relationship between natural and pathological muscle loss.

sample_organism:
Mus musculus
Jaculus jaculus

platform_organism:
Mus musculus
Jaculus jaculus

overall_design: To investiate muscle loss transcriptome, we isolate muscles from sections of foot and dissect FDS muscles of new born and P3 jerboa and mouse


title: Comparison of the pathogenicity of CA10 with CA16 and EV71 that cause hand, foot and mouth disease in a mouse model

city: Beijing

first: wanjun

last: Peng

address: No.5, Panjiayuan Nanli,Chaoyang District, Beijing

institute: Institute of Laboratory Animal Science

summary: In this study, models of CA10, CA16 and EV71 infection were established in 7-day-old Institute of Cancer Research (ICR) mice by intraperitoneal injection to analyze the pathogenicity of these viruses. RNA sequencing analysis was used to screen the differentially expressed genes (DEGs) after CA10 infection, and innate immune-related pathways were activated in muscle. Compared with CA16 and EV71 infection, CA10 may have an inhibitory effect on Th cell differentiation and cell growth. Additionally, the common DEGs in the three viruses were most enriched in the immune system response, including the Toll-like receptor pathway and the nucleotide-binding and oligomerization domain (NOD)-like pathway. Our findings revealed a group of genes that coordinate in response to CA10 infection and are shared by the three viruses, which increases our understanding of the pathological mechanism of HFMD.

overall_design: The differentially expressed genes between CA10,CA16,EV71 infected mice and WT mice were compared and analyzed


title: Age-related STAT3 signaling regulates severity of respiratory syncytial viral infection in human bronchial epithelium cells

first: Xingbin

last: Ai

address: 55 Fruit St

department: Pediatrics

institute: MASSACHUSETTS GENERAL HOSPITAL

summary: Respiratory syncytial virus (RSV) causes severe disease mostly in infants; however, mechanisms of age association remain elusive. Here, employing human bronchial epithelium models generated from tracheal aspirate-derived basal stem cells of neonates and adults, we investigate whether age regulates RSV-epithelium interaction to determine disease severity. We show that following RSV infection, only neonatal epithelium model exhibits cytopathy and mucus hyperplasia, and neonatal epithelium has more robust viral spread and inflammatory responses than adult epithelium. Mechanistically, RSV-infected neonatal ciliated cells display age-related impairment of STAT3 activation, rendering susceptibility to apoptosis, which facilitates viral spread. In contrast, SARS-CoV-2 infection of ciliated cells has no effect on STAT3 activation and is not affected by age. Taken together, our findings identify an age-related and RSV-specific interaction with neonatal bronchial epithelium that critically contributes to severity of infection, and STAT3 activation offers a potential strategy to battle severe RSV disease in infants.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for basal stem cells from adult and term human tracheal aspirate.

first: Caiqi

last: Zhao

first: Wei

last: Wang


title: miR196b-/- DNMT3A-mutant AML [RNA-seq]

first: Sara

middle: E

last: Meyer

department: Cancer Biology

summary: microRNA (miRNA or miR) are aberrantly expressed in acute myeloid leukemia (AML), and clinically may have diagnostic, prognostic, and therapeutic value. We identify miR-196b, is overexpressed in high-risk subset of DNMT3A-mutant AML and its activity is important to maintain a differentiation block and limit innate immune signaling pathways in AML. We interrogated previously published experimentally defined and computationally predicated miR-196b targets compared with genes differentially expressed in miR-196b-/-Dnmt3a/Flt3ITD AML versus Dnmt3a/Flt3ITD AML. This identified that miR-196b regulates Pou2f1 and Pou2f2 (Oct1 and Oct2) in Dnmt3a/Flt3-mutant AML.

overall_design: To identify the gene expression and pathway changes associated with murine Dnmt3a/Flt3ITD-mutant leukemia cells with and without miR-196b. n=3 independent AML harvested from different miR-196b-/-Dnmt3a/Flt3ITD mice. AML cells were enriched for cKit prior to RNA isolation. Dnmt3a/Flt3ITD AML controls (n=3) were previously published PMID:27016502 and can be found under GSE77849.

first: Michael

last: Lawler

first: Melanie

last: Goetz

first: Jennifer

middle: S

last: Romer-Seibert

first: Holly

last: Gamlen

first: Edwina

last: McGlinn


title: Rosmarinic acid alleviates radiation-induced pulmonary fibrosis by downregulating tRNA N7-Methylguanosine modification-regulated fibroblast to myofibroblast transition through the exosomes pathway [m7G-tRNA MeRIP-Seq]

first: jinglin

last: mi

first: Tingting

last: Zhang

first: Jinglin

last: Mi

first: Zhechen

last: Ouyang

first: Xinling

last: Qin


title: BATF is a major driver of NK cell epigenetic reprogramming and dysfunction in AML[RNA]

status: Public on Aug 31 2024

city: Houston

first: Anand

last: Singh

address: 1901 East Road

institute: MD Anderson Cancer Center

summary: We identified BATF as a core transcription factor and the main mediator of this NK cell dysfunction in AML. our findings reveal a previously unidentified mechanism of NK immune evasion in AML manifested by epigenetic rewiring and inactivation of NK cells by myeloid blasts.  This work highlights the importance of using healthy allogeneic  NK cells as adoptive cell therapy to treat patients with myeloid malignancies combined with strategies aimed at preventing the dysfunction by targeting the TGF-b pathway or BATF

overall_design: Libraries were sequenced Illumina NovaSeq6000. Samples: 1-NKcellsfromPB1withCas9only 2-NKcellsfromPB2withCas9only 3-NKcellsknockoutforTGFbR2fromPB1 4-NKcellsknockoutforTGFbR2fromPB2 5-NKcellsfromPB1withCas9only cocultured directly with AML1cells 6-NKcellsfromPB2withCas9only cocultured directly with AML1cells 7- NKcellsfromPB2withCas9only cocultured directly with AML2cells 8-NKcellsfromPB1withCas9only cocultured through transwell with AML1cells 9-NKcellsfromPB2withCas9only cocultured through transwell with AML1cells 10-NKcellsfromPB1withCas9only cocultured through transwell with AML2cells 11-NKcellsknockoutforTGFbR2fromPB1 cocultured directly with AML1cells 12-NKcellsknockoutforTGFbR2fromPB2 cocultured directly with AML1cells 13-NKcellsknockoutforTGFbR2fromPB1 cocultured directly with AML2cells 14-NKcellsknockoutforTGFbR2fromPB1 cocultured directly with AML2cells

first: Vakul

last: Mohanty

first: Bijender

last: Kumar

first: May

last: Daher

first: Kunal

last: Rai


title: Effect of abemaiclicb and deletion of TP53 on gene expression in MCF7 cells at different timepoints

city: New York

first: Rei

last: Kudo

address: 417 E 68th Street

department: HOPP

institute: Memorial Sloan Kettering Cancer Center

summary: CDK4/6 inhibitor, abemaciclib induced cell cycle arrest and senescence in breast cancer. We reported that deletion of TP53 prevented the senescence features. In this study, we knockout TP53 by CRISPR Cas9 system and treated MCF7 cells and p53 knockout cells with abemaciclib for up to 29 days to identify the difference of senescence associated secretary phenotype genes and cell cycle related genes. features.

overall_design: We established TP53 knockout (p53KO) cell line by CRISPR Cas9 system in MCF7 breast cancer cell line. Cells were treated with 50 nM abemaciclib. We collected cells on day 0 (no treatment), day1, day 3, day 14 and day 21. We then performed gene expression profiling analysis from RNA sequencing of parental MCF7 and p53KO cell.

first: Sarat

last: Chandarlapaty


title: BATF is a major driver of NK cell epigenetic reprogramming and dysfunction in AML[CUT&RUN]

overall_design: Libraries were sequenced using Illumina NextSeq 500 system to obtain 75bp paired end reads per sample.  Samples- 1 NKCells fromPB1 untreated 2. NKCells fromPB1 TGFb1 treated 1 NKCells  fromPB2 untreated 4. NKCells fromPB2 TGFb1 treated,  these samples were Cut and run for abs - BATF (CST), and IgG( millipore)

first: Abhinav

last: Jain


title: Unraveling the CDK9/PP2A/ERK network in transcriptional pause release and complement activation in KRAS-mutant cancers

city: Shanghai

first: yafang

last: Wang

address: 393 Middle Huaxia Road

institute: Shanghaitech University

summary: A time-course RNA-seq experiment testing the effect of drug treatment using selective CDK9 inhibitor AZD4573. The experiment was done using a pancreatic cancer cell line HPAF-II carrying KRAS G12D mutation.

overall_design: The experiment tests the effect of CDK9 inhibitor AZD4573 at 4 hrs. and 8 hrs. time-points vs. a DMSO untreated control in HPAF-II cell lines.

first: Yafang

first: Lansong

last: Xu

first: Shangxuan

last: Shi

first: Chengying

last: Xie


title: Unraveling the CDK9/PP2A/ERK network in transcriptional pause release and complement activation in KRAS-mutant cancers (MIA PaCa-2 )

summary: KRASG12C inhibitors (KRASG12Ci) AMG510 and MRTX849 have shown promising efficacy in clinical trials and been approved for the treatment of KRASG12C-mutant cancers. However, the emergence of therapy-related drug resistance limits their long-term potential. We measured gene expression using RNA-sequencing for pancreatic cancer cell line MIA PaCa-2 parental and their AMG510-resistant equivalent without treatment or treated with with G12C KRAS inhibitor AMG510.

overall_design: AMG510-resistant MIA PaCa-2 cell line were generated from parental cell line by continuous dose escalation from as low as 1nM to 10μM. Once resistance to AMG510 was confirmed, both parental and resistant cell lines were treated with DMSO or AMG510. Gene expression analyses of these samples were performed.


title: RNAseq analysis of mouse monocytes exposed to IL4 ex vivo

city: Paris

first: Elodie

last: Segura

address: 156 rue de Vaugirard

department: INSERM U1151

country: France

institute: INSERM

summary: Immune cells can rapidly adapt their functional program in response to cytokines. How cytokine-induced transcriptional responses are affected by micro-environmental cues remains poorly understood. The Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor sensing environmental signals including metabolites. Here we addressed the cross-talk between Interleukin-4 and AhR signaling in monocytes.

overall_design: We analysed the transcriptome of mouse monocytes (wild-type or deficient for AhR) to IL-4 after 3h of ex vivo exposure.


title: ChIP-seq analysis of STAT6 in human monocytes

overall_design: We immuno-precipitated STAT6 20 min after inducing its translocation to the nucleus by exposing monocytes to IL-4, in the presence of SR1 (AhR inhibitor) or FICZ (AhR agonist).

first: Darawan

last: Tabtim-On


title: BATF is a major driver of NK cell epigenetic reprogramming and dysfunction in AML[ATAC-seq]

overall_design: Libraries were sequenced 2 x 100 bp on an Illumina NovaSeq6000 to obtain at least 40 million high quality mapping reads per sample. Samples: 1-NKcellsfromPB1withCas9only 2-NKcellsfromPB2withCas9only 3-NKcellsknockoutforTGFbR2fromPB1 4-NKcellsknockoutforTGFbR2fromPB2 5-NKcellsfromPB1withCas9only cocultured directly with AML1cells 6-NKcellsfromPB2withCas9only cocultured directly with AML1cells 7- NKcellsfromPB2withCas9only cocultured directly with AML2cells 8-NKcellsfromPB1withCas9only cocultured through transwell with AML1cells 9-NKcellsfromPB2withCas9only cocultured through transwell with AML1cells 10-NKcellsfromPB1withCas9only cocultured through transwell with AML2cells 11-NKcellsknockoutforTGFbR2fromPB1 cocultured directly with AML1cells 12-NKcellsknockoutforTGFbR2fromPB2 cocultured directly with AML1cells 13-NKcellsknockoutforTGFbR2fromPB1 cocultured directly with AML2cells 14-NKcellsknockoutforTGFbR2fromPB1 cocultured directly with AML2cells

first: Manrong

last: Wu


title: Effect of long-term abemaiclicb and deletion of TP53 on gene expression in MCF7 cells

overall_design: We established TP53 knockout (p53KO) cell line by CRISPR Cas9 system in MCF7 breast cancer cell line. Cells were treated with 50 nM abemaciclib. We collected cells on day 0 (no treatment), day1, day 3, day 5, day 21 and day 29. We then performed gene expression profiling analysis from RNA sequencing of parental MCF7 and p53KO cell.


title: Examinining the myelin transcription factor dependent genes in islet beta cells under euglycemia

city: Nashville

first: Guoqiang

last: Gu

state: TN

address: 2213 Garland Avenue, MRB4, Rm 9415

department: Cell and Developmental Biology

institute: Vanderbilt University

summary: A hallmark of type 2 diabetes (T2D) is endocrine islet beta-cell failure, which can occur via cell dysfunction, loss-of-identity, and/or death. How each is induced remains largely unknown. Here, we use mouse beta-cells that are deficient for Myelin transcription factors (Myt TFs, including Myt1, 2, and 3) to address this question. We have reported that inactivating all three Myt genes in pancreatic progenitor cells (MytPancD) causes beta-cell failure and late onset diabetes in mice. Their lower expression in human beta-cells is correlated with beta-cell dysfunction and SNPs in Myt2 and Myt3 are associated in higher risk of T2D. We now show that these Myt TF-deficient beta-cells also de-differentiate by reactivating several progenitor markers, assayed by both immuno-assays and RNAseq. Intriguingly, mosaic Myt TF inactivation in only a portion of islet beta-cells does not results in overt diabetes, but this creates a condition where Myt TF-deficient beta-cells stay alive while dedifferentiating and trans-differentiating into Ppy-expressing cells.  By transplanting MytPancD islets into the anterior eye chambers of immune-compromised mice, we directly show that glycemic and obesity-related conditions influence cell fate. These findings suggest that the observed beta-cell defects in T2D depend on not only their inherent genetic/epigenetic defects, but also the metabolic load.

overall_design: Control and mutant beta cells from 2-months old mice were purified, RNA prepared, and sequenced. Note that the raw data of the control samples, done at the same time as the mutants, were deposited in Array-Express under the submission number: E-MTAB-2266.

first: Yanwen

first: Chen

last: Huang

first: Ruiying

last: Hu

first: Xin

last: Tong


title: ChromaFold predicts the 3D contact map from single-cell chromatin accessibility

city: New York City

first: Christina

last: Leslie

address: 417 East 68th Street

summary: The identification of cell-type-specific 3D chromatin interactions between regulatory elements can help to decipher gene regulation and to interpret the function of disease-associated non-coding variants. However, current chromosome conformation capture (3C) technologies are unable to resolve interactions at this resolution when only small numbers of cells are available as input. We therefore present ChromaFold, a deep learning model that predicts 3D contact maps and regulatory interactions from single-cell ATAC sequencing (scATAC-seq) data alone. ChromaFold uses pseudobulk chromatin accessibility, co-accessibility profiles across metacells, and predicted CTCF motif tracks as input features and employs a lightweight architecture to enable training on standard GPUs. Once trained on paired scATAC-seq and Hi-C data in human cell lines and tissues, ChromaFold can accurately predict both the 3D contact map and peak-level interactions across diverse human and mouse test cell types. In benchmarking against a recent deep learning method that uses bulk ATAC-seq, DNA sequence, and CTCF ChIP-seq to make cell-type-specific predictions, ChromaFold yields superior prediction performance when including CTCF ChIP-seq data as an input and comparable performance without. Finally, fine-tuning ChromaFold on paired scATAC-seq and Hi-C in a complex tissue enables deconvolution of chromatin interactions across cell subpopulations. ChromaFold thus achieves state-of-the-art prediction of 3D contact maps and regulatory interactions using scATAC-seq alone as input data, enabling accurate inference of celltype- specific interactions in settings where 3C-based assays are infeasible.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: single cell ATAC sequencing data of human embryonic stem cell and mouse germinal center B cells and Hi-C of mouse regulatory T cells


title: NSC95397 is a Novel HIV Latency Reversing Agent

city: Los Angeles

first: Oliver

middle: I.

last: Fregoso

state: CA

address: 615 Charles E Young

department: MIMG

institute: University of California Los Angeles

summary: The latent viral reservoir represents one of the major barriers of curing HIV. Focus on the “kick and kill” approach, in which virus expression is reactivated then cells producing virus are selectively depleted, has led to the discovery of many latency reversing agents (LRAs) that can reactivate latently integrated virus and further our understanding of the mechanisms driving HIV latency and latency reversal. Thus far, individual compounds have yet to be robust enough to work as a therapy, highlighting the importance of identifying new compounds that target novel pathways and synergize with known LRAs. In this study, we identified a promising LRA, NSC95397, from a screen of ~4250 compounds in J-Lat cell lines. We validated that NSC95397 reactivates latent viral transcription and protein expression from cells with unique integration events. Co-treating cells with NSC95397 and known LRAs demonstrated that NSC95397 has the potential to synergize with different drugs under both standard normoxic and physiological hypoxic conditions. We showed that NSC95397 does not globally increase open chromatin, and bulk RNA sequencing revealed that NSC95397 does not greatly change cellular transcription. Instead, NSC95397 downregulates many pathways key to metabolism, cell growth, and DNA repair – highlighting the potential of these pathways in regulating HIV latency. Overall, we identified NSC95397 as a novel LRA that does not alter global transcription, that shows potential for synergy with known LRAs, and that may act through novel pathways not previously recognized for their ability to modulate HIV latency.

overall_design: To characterize differential genes expressed when latent HIV is reactivated by different latency reversing agents, 4x10^6 J-Lat 10.6 cells were cultured in RPMI media and were untreated or treated with 10.0 ng/mL PMA, 25.0 uM SAHA, or 5.0 uM NSC95397 for 24 hrs and then treated with TRIzol for RNA isolation. Experiment was done in quadruplicate.

first: Randilea

last: Nichols Doyle

first: Vivian

last: Yang

middle: I