title: Differential expression and localization of expansins in Arabidopsis shoots: Implications for cell wall dynamics and drought tolerance

city: Brno

first: Core Facility

last: Bioinformatics

address: Kamenice 5

country: Czech Republic

institute: CEITEC - Masaryk University

summary: Expansins are cell wall-modifying proteins implicated in plant growth and stress responses. In this study, we explored the differential localization of expansins in Arabidopsis thaliana shoots, with a focus on EXPA1, EXPA10, EXPA14, and EXPA15. Utilizing pEXPA::EXPA translational fusion lines, we observed distinct expression patterns of expansins, with EXPA1 primarily localized in stomatal guard cells, while EXPA10 and EXPA15 showed strong cell wall (CW) localization in epidermal and other tissues. Overexpression of EXPA1 resulted in pronounced changes in CW-related gene expression, particularly during early stages of induction, including the upregulation of other expansins and CW-modifying enzymes. The induced EXPA1 line also displayed significant morphological changes in shoots, including smaller plant size, delayed senescence, and structural alterations in vascular tissues. Additionally, EXPA1 overexpression conferred drought tolerance, as evidenced by enhanced photosynthetic efficiency (Fᵥ/FM), and low steady-state non-photochemical quenching (NPQ) values under drought stress. These findings highlight the critical role of EXPA1 in regulating plant growth, development, and stress response, with potential applications in improving drought tolerance in crops.

overall_design: Comparative gene expression analysis of RNA-seq data for wild type and Dex-inducible line under three conditions (control, 3h Dex treatment, 7 days Dex treatment).

first: Balkova

last: Darina

first: Mala

last: Katerina

first: Hejatko

last: Jan

first: Panzarova

last: Klara

first: Abdelhakim

last: Lamis

first: Pleskacova

last: Barbora

first: Samalova

last: Marketa


title: Bulk RNA-seq of CD248+/- PDGFR-α+ cardiac fibroblasts

status: Public on Jan 24 2025

city: Hangzhou

first: Mo

last: Li

address: Jiefang road 88

department: Department of Cardiology of The Second Affiliated Hospital, School of Medicine

country: China

institute: Zhejiang University

type: Expression profiling by high throughput sequencing

summary: Post-injury remodeling is a complex process involving temporal specific cellular interactions in the injured tissue where the resident fibroblasts play multiple roles, including inflammation induction and tissue remodeling.   To dissect the molecular basis of these interactions, we performed single-cell and spatial transcriptome analysis in human and mouse hearts after myocardial infarction.  A subset of fibroblasts identified by unique high expression of CD248 was strongly correlated with collagen synthesis and remodeling, and genetic deletion of CD248 in fibroblast ameliorated cardiac fibrosis and dysfunction following ischemia/reperfusion, highlighting the functional importance of CD248 positive fibroblasts in post-injury pathological remodeling. CD248 stabilizes TGF-βR Ⅰ protein and activates ACKR3 expression in fibroblast, leading to enhanced T cell adhesion and retention. This CD248 mediated fibroblast-T cell interaction is required to sustain fibroblast activation and scar expansion in injured heart. Disruption of this interaction using monoclonal antibody or Chimeric antigen receptor T cell reduces T cell infiltration, ameliorates ischemia/reperfusion-induced cardiac fibrosis and improves heart function.  Therefore, single cell and spatial molecular profiling in post-injury heart has uncovered a CD248 specific subclass of fibroblast with a critical role in fibroblast-T cell interaction and a new potential therapy to treat tissue fibrosis.

overall_design: CD248+ PDGFR-α+ and CD248- PDGFR-α+ cardiac fibroblasts from mouse hearts 14 days post MI were FACS-sorted followed by bulk RNA-seq.

first: Guohua

first: Xinyang

last: Hu


title: Single-Cell Sequencing Unveils the Cellular Characteristics Underlying the Onset and Progression of Hematogenous Disseminated Tuberculosis

city: Chongqing

first: Zhen

last: Gong

address: Tian Sheng road

institute: Southwest University

summary: The etiology of hematogenous disseminated tuberculosis (DTB) likely involves multiple factors, though it remains incompletely understood. In this study, we performed single-cell RNA transcriptome and T cell receptor (TCR) sequencing on samples from 7 DTB patients. We observed a significantly elevated proportion of inflammatory immune cells (e.g., monocytes and macrophages) and a markedly decreased abundance of various lymphocytes (e.g., T cells, B cells, and plasma cells) in DTB. This suggests that lymphopenia might be a prominent feature of the disease. T cell pseudotime analysis revealed that most hypervariable gene expression decreased over time in DTB, suggesting that T cell functional exhaustion may be an immunological characteristic of DTB patients. Another important manifestation of T cell exhaustion in DTB is the significant absence of mucosal-associated invariant T (MAIT) cells, a crucial cytological feature in DTB peripheral blood. Specific polymorphisms were identified in the TCR repertoire of DTB patients, including TRAV9-2, TRAV13-1, TRBV20-1, and TRBV5-1, with dominant clones TRAJ49, TRBJ2-7, and TRBJ2-1. Analysis of complementarity determining region 3 (CDR3) revealed that the most commonly used combination was TRAV1-2/TRAJ33. Notably, a motif containing "CAAMD" was significantly reduced in DTB patients, which may indicate unique characteristics of the disease. In summary, our study offers new insights into the complex dynamic changes of TCR clonotypes, CDR3 dominant motifs, and various cell types in the pathogenesis of DTB.

overall_design: Whole blood samples were collected from patients diagnosed with 7 disseminated tuberculosis (DTB) at Shenyang Chest Hospital.

first: Hongxaing

last: Xu

first: Qiao

last: Zhang

first: Yu

last: Chen

first: Jianping

last: Xie


title: CTCF is selectively required for maintaining chromatin accessibility and gene expression in human erythropoiesis

status: Public on Jan 26 2025

city: Suzhou

first: Peng

state: Jiangsu

address: Office3625 Building703, RENAI ROAD No.199, INDUSTRIAL PARK DISTRICT

department: Suzhou Medical College

institute: Soochow University

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: Deficiency of the palmitoyl acyltransferase ZDHHC7 modulates gene expression in mice after a mild chronic stress paradigm

city: Münster

first: Christa

last: Hohoff

address: Albert-Schweitzer-Campus 1, A9

department: Department of Psychiatry

country: Germany

institute: University of Muenster

summary: Chronic stress (CS) is a debilitating condition that negatively affects body and brain. In mice, CS effects range from changes in behaviour down to the level of gene expression. These effects are partly mediated by sex and sex steroid hormones, which in turn are affected by the palmitoyl acyltransferase ZDHHC7. ZDHHC7 might modulate also the response to CS via palmitoylation of sex steroid hormone receptors and other proteins critical for neuronal structure and functions. Therefore, we aimed to investigate the role of ZDHHC7 in response to CS on different system levels in a mouse model of Zdhhc7-deficiency. Female and male Zdhhc7-knockout (KO) and -wildtype (WT) mice underwent a four-week-mild CS paradigm or non-stress control (C) condition. After C or CS, amongst other analyses, brain gene expression profiles (via mRNA-seq transcriptomics) were investigated. Analyses focused on effects of genotype (KO vs. WT) or condition (C vs. CS) separately in both sexes. Our results revealed significant effects particularly in females. Female KOs showed upregulated cortistatin expression after CS (C vs. CS: p<0.05). Zdhhc7-deficiency seemed to be related to a sex-specific stress response and may reveal genetic factors of CS-resilience in female mice.

overall_design:
Female and male Zdhhc7-KO and -WT mice underwent a four-week-mild CS paradigm or non-stress C condition. After C or CS, mRNA-seq of brain tissues (hippocampus, medial prefrontal cortex) was performed to analyse effects of genotype (KO vs. WT) or condition (C vs. CS) separately in both sexes.

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

last: Kerkenberg

first: Mingyue

first: Weronika

last: Palkowska

first: Lydia

last: Wachsmuth

first: Maja

last: Peng

first: Lena

last: Stiehl

first: Christiane

last: Schettler

first: Johannes

last: Zang

first: Andreas

last: Huge

first: Evgeni

last: Ponimaskin

first: Cornelius

last: Faber

first: Bernhard

middle: T

last: Baune

first: Weiqi


title: Effect of HDAC1 deficiency on in vitro Th2 and pathogenic Th2 differentiated murine CD4+ T cells

status: Public on Jan 23 2025

city: Vienna

last: Boucheron

address: Lazarettgasse 19

country: Austria

institute: Institute of Immunology

summary: The differentiation of naïve CD4+ T cells into distinct effector T helper (Th) subsets is controlled by the chromatin state of the cells which in turn is regulated by various epigenetic mechanisms such as reversible lysine acetylation mediated by histone acetyltransferases and histone deacetylases (HDACs). In addition, Th2 cells can aquire pathogenic features, secreting pro-inflammatory factors and high levels of the cytokine IL-5. By using HDAC1 deficient naïve CD4+ T cells and establishing a protocol for differentiation of pathogenic Th2 cells, we could establish a critical role for HDAC1 in regulating the differentiation and pathogenicity of in vitro generated pathogenic Th2 cells

overall_design: To study the function of HDAC1 in the differentiation of pathogenic Th2, naïve CD4+ T cells were isolated by magnetic cell sorting from CD4cre-HDAC1f/f (WT) and CD4cre+HDAC1f/f (KO) and differentiated into Th2 and pathogenic Th2 in vitro. Briefly, cells were activated with immobilized anti-CD3/CD28 in a polarizing cytokine milieu. Cells wer collected after 3 days in culture and RNA processed for RNA sequencing. We performed gene expression profiling by comparing WT Th2 to WT pathogenic Th2 conditions as well as KO Th2 to KO pathogenic Th2 and WT Th2.

first: Matarr

last: Khan

first: Lina

last: Dobnikar

first: Michael

last: Schuster

first: Moritz

last: Madern


title: Suppression of endothelial ceramide de novo biosynthesis contributes to cardiometabolic diseases

city: New York

first: Olivier

last: Elemento

state: NY

address: 1305 York Avenue

country: USA

institute: WEILL MEDICAL COLLEGE OF CORNELL UNIV

summary: Metabolic disorders and obesity increase the risk for cardiovascular (CV) disease, including hypertension, atherosclerosis, and myocardial infarction. Systemic endothelial dysfunction, a well-established response to CV risk factors, precedes the development of CV diseases. However, growing evidence suggest that endothelial dysregulation also contributes to metabolic disorders, underlying a centric role of the endothelium in cardiometabolic diseases. The accrual of the sphingolipid ceramide, bioactive molecule, has been causally implicated in endothelial dysfunction in vitro. However, direct in vivo evidence supporting the accrual of ceramide in the endothelium, underlying mechanisms, and pathological implications are lacking. Here, we showed that the suppression rather than the accrual of ceramides is causally linked to endothelial dysfunction, and imparts a pro-inflammatory and pro-atherosclerotic phenotype, contributing to both vascular and metabolic disorders.  Mechanistically, the upregulation of Nogo-B and ORMDLs proteins, inhibitors of the first and rate limiting enzyme of the de novo biosynthesis, in the endothelium of obese and diabetic mice suppresses sphingolipid signaling, particularly ceramides and sphingosine-1-phosphate, resulting in vascular and metabolic dysfunctions. Systemic and endothelial specific deletion of Nogo-B restores sphingolipid signaling and functions of the endothelium, improves hypertension and lowers hepatic glucose production. Our results demonstrating that Nogo-B-mediated suppression of sphingolipid metabolism and signaling in the endothelium has pathological implication in cardiometabolic disorders and set a framework for the development of therapeutic strategies to treat these conditions.

overall_design: FACS sorted ECs were isolated from hearts of ApoE-/- and Nogo-A/B(ECKO)ApoE-/- mice after 8 weeks post-TAC and were then processed for bulk RNAseq.

first: Annarita

last: Di Lorenzo

first: Rohan

last: Bareja

first: Luisa

last: Rubinelli


title: Urolithin A Provides Cardioprotection and Mitochondrial Quality Enhancement Preclinically and Improves Cardiovascular Health Biomarkers in Humans

city: Ecublens

first: Davide

last: D'Amico

address: EPFL Innovation Park, Building C

country: Switzerland

institute: Amazentis SA

summary: Cardiovascular diseases (CVDs) remain the primary cause of global mortality. Nutritional interventions hold promise to reduce CVD risks in an increasingly aging population. However, few nutritional interventions are proven to support heart health and act mostly on blood lipid homeostasis rather than at cardiac cell level.  Here, we show that mitochondrial quality and dysfunction are a common hallmark in human cardiomyocytes upon heart aging and in chronic conditions. Preclinically, the post-biotic and mitophagy activator, Urolithin A (UA), reduced both systolic and diastolic cardiac dysfunction in models of natural aging and heart failure. At a cellular level, this was associated with a recovery of mitochondrial ultrastructural defects and mitophagy. In humans, UA supplementation for 4 months in healthy older adults significantly reduced plasma ceramides clinically validated to predict CVD risks. These findings extend and translate UA’s benefits to heart health, making UA a promising nutritional intervention to support cardiovascular function as we age.

overall_design: Fifty Wistar male rats were purchased from Janvier Labs (France). Animals were collectively housed in cages at SYNCOROSOME’s facilities and had free access to food (RM1, SDS Dietex) and drinking water ad libitium.  Surgery was initiated in 7-week-old rats that were separated into three groups. MI was induced in animals from Surgery Vehicle (n=20) and Surgery UA (n=20), by chronic left anterior descending coronary artery (LAD) ligation performed at day 0. Rats were anesthetized by an intraperitoneal injection (IP) of Medetomidine (0.5 mg/kg) and Ketamine (50mg/kg). The animals were then intubated and ventilated at 10 ml/kg tidal volume and 70-80 cycles/minutes. The body temperature was maintained around 37°C using a thermoregulated heating pad connected to a rectal probe. Rats were placed in supine position, chest-shaved and prepared for standard surgical aseptic conditions. A left lateral thoracotomy was carefully performed to expose the heart. Suture was placed around the LAD (4/0 Silk, Ethicon), around 2 mm below the left atrium and close to the interventricular junction, to obtain infarct size (IS) near 40% of the total left ventricular (LV) area. The chest was closed, air was expelled from the rib cage to avoid pneumothorax and quick reanimation was performed using atipamezole hydrochloride (IM, 0,5 mg/ml, Zoetis). The sham vehicle animals (n=10) were subjected to the same protocol. However, after the left lateral thoracotomy exposing the heart, the rib cage was closed without passing the suture thread around LAD.  A peri-operative care of the animals was performed during the surgery. For 8 weeks, starting 24 hours after the surgery UA animals were daily treated with Urolithin A at 50 mg/ml by oral gavage. The rats from sham vehicle and surgery vehicle group received the same dose of a vehicle solution (0.5% carboxymethylcellulose) at the same time. The volume of administration was adjusted every week based on the mean body weight.  One day after the last dosing, overnight-fasted rats were anesthetized with 2.5% isoflurane in 50% O2  50% air mixture followed by an injection of buprenorphine (0.03mg/kg). Hearts were stopped in diastole (KCL 2M, 1ml/kg) by jugular injections. The atria were removed, the hearts were washed in cold PBS and quickly sponged to remove the liquid from the ventricles. LV was separated from the remaining part of the heart, weighed and incubated in NBF 10% for 48h. Hearts were transferred in ethanol 70%, cut in three transverse sections and embedded in paraffin in three different blocks, deriving from apical, middle and basal LV regions (Group 1, N=10, group2 and N=20).

first: Julie

last: Faitg

first: Charlotte

last: Tissot


title: Single-cell assay for transposase-accessible chromatin sequencing of human clear cell renal cell carcinoma

city: Nanning

first: Zhenyuan

last: Yu

state: Guangxi

address: Shuangyong road 22

department: Center for Genomic and Personalized Medicine

institute: Guangxi Medical University

summary: In recent years, with the development of single-cell sequencing technology, it has become possible to perform single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq), especially the high-throughput single-cell analysis. Here, we provide a high-quality chromatin accessibility data for ccRCC at single-cell level. Performing high-throughput scATAC-seq on 3 human ccRCC samples (RCC30, RCC61 and RCC76), we obtained a total of 18,703 high quality nuclei and 104,818 unique peaks, including coding, non-coding regions, promoters and enhancers.we demonstrate a complete analysis process for scATAC-seq data, and makes the application more feasible and convenient.

overall_design: ccRCC samples from 3 patients were used for single-cell ATAC sequencing by 10X Genomics.

first: Wenhao

last: Lu

first: Yixuan

last: Liu

first: Jiaping

last: Li

first: Shengzhu

last: Huang

first: Zheng

last: Wen

first: Cheng

last: Su

first: Zengnan

last: Mo


title: Urolithin A Provides Cardioprotection and Mitochondrial Quality Enhancement Preclinically and Improves Cardiovascular Health Biomarkers in Humans [mouse_cardiac_aging]

overall_design: .Twenty-four male C57BL/6Rj mice aged 21 months old and eighteen male C57BL/6Rj mice aged 8 weeks old were purchased from Janvier Labs (France). Mice were collectively housed in cages in controlled room at 22°C and 12 h light/ dark cycle at Biomeostasis’ facilities. All animals were allowed ad libitium access to an irradiated normal chow diet (NC, irradiated pellet A04; SAFE, Villemoisson-sur-Orge, France) and to ultrapure and laboratory-grade acidified water (Aquavive®, INNOVIVE, France).  For 8 weeks, 21-months-old and 8-week-old mice were treated by oral gavage, either with a vehicle solution (0.5% carboxymethylcellulose) (21 months vehicle, n=12 or 8 weeks vehicle n=9) or with UA at 50mg/ml (21 months UA, n=12 or 8 weeks UA, n=9). One day after the last dosing, overnight-fasted mice were anaesthetized by Isoflurane.  The hearts were collected, weighed and the LV and RV were separated.  LV was embedded in OCT.  Excess OCT was trimmed away from the frozen tissues block with a sterile scalpel blade.


title: Single-cell transcriptomic profiling of cardiac CD45+ immune cells 24 hours after ischemia-reperfusion injury in mice with or without splenectomy

status: Public on Jan 22 2025

city: shanghai

first: qiang

last: long

address: OE

institute: OE

summary: This study investigates the impact of splenectomy on the cardiac immune cell landscape post-I/R injury. We performed single-cell RNA sequencing (scRNA-seq) on CD45+ immune cells isolated from mouse hearts 24 hours after I/R injury in both splenectomized and sham-operated (control) mice.

overall_design: Male C57BL/6 mice aged 8 weeks were randomly assigned to two groups: splenectomy and sham surgery.  both groups underwent myocardial ischemia-reperfusion (I/R) injury induced by occluding the left anterior descending (LAD) coronary artery for 60 minutes, followed by 24 hours of reperfusion. Hearts were then harvested, and CD45⁺ immune cells were isolated using magnetic-activated cell sorting (MACS).

first: Qiang

last: Long

first: Kristina

last: Rabi

first: Yu

last: Cai

first: Lihui

first: Shixing

first: Bei

last: Qian

first: Yiming

last: Zhong

first: Zhaoxi

last: Qi

first: Yecen

last: Zhang

first: Kaichen

first: Xinming

last: Wang

first: Lan

last: Chang

first: Weichang

last: Xie

first: Huaiyu

last: Jiang

first: Haonan

first: Junjie

first: Ting

last: Ren

first: Zichen

first: Tambet

last: Teesalu

first: Caisheng

last: Wu

first: Lin

first: Zhengbin

last: Zhu

first: Hélder

last: Santos

first: Zehua

last: Zhao

first: Xiaofeng

last: Ye


title: Molecular differences between neonatal and adult stria vascularis from organotypic explants and transcriptomics

status: Public on Jan 27 2025

city: Bethesda

first: Robert

last: Morell

state: MD

address: 35A Convent Drive, Room 1F-103

institute: National Institute on Deafness and Other Communication Disorders

summary: The stria vascularis (SV) generates the endocochlear potential that is required for hearing and maintains the blood-labyrinth barrier which protects the inner ear from pathogenic infiltration. SV degeneration disrupts cochlear homeostasis and results in progressive and irreversible hearing loss, yet little is known about this tissue. We elucidate the molecular differences between the neonatal (post-natal day 1, P1) and adult (P30) SV through the analysis of single-cell RNA sequencing on SV from both stages. We revealed key genes, transcription factors, and pathways unique to each stage that may play a role in SV proliferation, development, and maintenance.

overall_design: Neonatal (post-natal day 1, P1) and adult (P30) SV tissue was dissociated and run on a 10X Controller for single cell isolation and library production. Libraries were sequenced on an Illumina NextSeq500 and raw gene expression matrices analyzed in Seurat, followed by CellChat for intercellular communication analyses and Slingshot for trajectory analyses.

first: Matsya

last: Thulasiram

first: Ryosuke

last: Yamamoto

first: Rafal

last: Olszewski

first: Shoujun

last: Gu

middle: J

first: Michael

last: Hoa

first: Alain

last: Dabdoub


first: Peng

last: Xu

summary: Backgrounds：The three-dimensional chromatin niche provides a precise gene expression control of cell identity, differentiation, and disease development. CTCF is considered as the most essential transcription factor regulating chromatin architecture and gene expression. However, the genome-wide impact of CTCF on erythropoiesis has not yet been extensively investigated. Results：Here, using a state-of-the-art human erythroid progenitor cell model (HUDEP-2 and HEL cell lines), we systematically investigated the effects of acute CTCF loss by an auxin-inducible degron system on transcriptional programs, chromatin accessibility, CTCF genome occupancy, and the 3D genome architecture. By integrating multi-omics datasets, we revealed that acute CTCF loss notably disrupted genome-wide chromatin accessibility and transcription network. Importantly, we identified that over thousands of chromatin accessibility regions were decreased while only a few hundred increased regions after depletion of CTCF in both HUDEP-2 and HEL lines, suggesting the role of CTCF in maintaining the proper chromatin openness in erythroid lineage. Moreover, CTCF depletion in the erythroid context significantly disrupted TAD boundary integrity and chromatin loops while not affecting the nuclear compartmentalization. Our study also reported that hundreds of erythroid lineage-specific genes were suppressed in both immature and mature stages, including metabolism-related target genes such as GOT2 and FXN. Notably, we identified a subset of genes increased their transcriptional level after the CTCF depletion, accompanied by the decreased chromatin accessibility regions that were enriched with the GATA motif. Specifically, we identified that CTCF repressed the master transcription factor GATA2 with distal decreased chromatin regions via multiple functional studies, including CTCF occupancy profiling and CRISPR/Cas9 editing of the CTCF binding site. These results suggest a suppressive role of CTCF in gene expression in erythroid lineage specification. Conclusions：In summary, our results revealed a novel role of CTCF in regulating erythroid differentiation by maintaining its proper chromatin openness, which will undoubtedly extend our understanding of CTCF biology.

overall_design: The HEL cells used in the experiment were knock in cell lines of GFP-miniAID-CTCF constructed by ourselves. We treated the cell lines with drugs for 24 hours, The drugs used included DMSO and IAA. IAA wa used for quick depletion CTCF  protein.

first: Xue

last: Yang

first: Li

last: Cheng

first: Ye

last: Xin

first: Jianxiang

last: Zhang

first: Beisi

first: Chunliang

last: Li


title: CRISPR editing of candidate host factors that impact influenza A virus infection

city: Worcester

first: Evan

middle: Stuart

last: Bradley

state: MA

address: 55 N Lake Avenue

department: Microbiology

institute: UMass Chan Medical School

summary: Influenza A virus (IAV) is a respiratory pathogen with a segmented negative-sense RNA genome that is capable of causing epidemics and pandemics. The error-prone nature of the IAV polymerase results in antigenic drift and antigenic shift which contribute to low vaccine efficacy and escape from antivirals. The host factors required for the complete IAV infectious cycle have not been fully identified. Here, we examined select host factors that were identified by independent CRISPR screens as candidate contributors to IAV infectivity. We performed CRISPR-mediated knockout of cytidine monophosphate N-acetylneuraminic acid synthetase (CMAS) as well as CRISPR-mediated overexpression of beta-1,4 N-acetylgalactosaminyltransferase 2 (B4GALNT2) and adenosine deaminase acting on RNA 1 (ADAR1) in the human bronchial epithelial A549 cell line and evaluated for reduction of IAV infection. We confirmed that the knockout of CMAS or overexpression of B4GALNT2 restricts IAV infection by diminishing binding to the cell surface. While ADAR1 overexpression did not significantly inhibit IAV replication, it had a pro-viral effect with coxsackie B virus infection.  Taken together, these data provide insight into how host factors identified in CRISR screens can modulate IAV infection and be utilized to further understand the IAV life cycle and for developing antiviral strategies.

overall_design: A549 Cas9 and A549-dCas9-VP64 cells were generated via lentiviral transduction of SpCas9 or the VP64 activator domain (dCas9-VP64) into A549 cells. Clonal cell lines with high expression of Cas9 or dCas9-VP64 proteins were established by limiting dilution after selection with 10 ug/mL blasticidin S HCl (ThermoFisher #A1113903) for 14 d. For A549 CalB4GALNT2 and CalADAR1 overexpressing lines, sgRNA sequences for the CRISPRa candidate genes were cloned into expression vector pXPR-502 as described previously (28, 37). sgRNA targeting the B4GALNT2 promoter region or ADAR1 and complementary oligos with appropriate nucleotide overhangs were ordered from ThermoFisher and annealed with a thermocycler to form dsDNA duplex following established protocol. Briefly, 1 uM of oligos were mixed with T4 Ligase Buffer (NEB) and T4 PNK enzyme. Annealing of the oligos was performed with the thermocycler following the program: 37 °C for 30 min, 95 °C for 5 min, then decreasing the temperature by 5 °C/min until 25 °C was reached. 90 uL of water was added to each reaction and 2 uL of duplexed oligos were ligated to 25 ng of linearized pXPR-502 lentiviral expression vector in 20 uL ligation reaction at 16 °C overnight. 7 uL of the ligated material was used to transform Stbl3 E. coli, which was plated on LB plates containing 100 ug/mL ampicillin. At least three bacterial colonies were sent out for Sanger sequencing and plasmids with the correct sgRNA sequences were expanded for lentiviral transduction (37). Confluent HEK293T cells in a 10 cm dish were transfected with plasmids expressing VSV-G envelope protein (pMD2.G), HIV structural proteins (psPAX2), and pXPR-502 expressing the specific sgRNA from a U6 promoter and a puromycin resistance gene from an EF-1a promoter at the ratio of 1:1.5:3 (4, 6, and 12 ug, respectively) using TransIt-293T. Supernatants were collected every 24 h for 3 d and pooled. The supernatant was concentrated with LentiX-concentrator and added onto A549-dCas9-VP64 cells. Lentivirus-transformed cells were then selected with 2 ug/mL puromycin for 7 d. The survivor cells were then expanded for maintained in complete DMEM supplemented with 2 ug/mL puromycin. Supernatants were collected for 3 d and passed through a 0.45 uM filter to remove debris before adding onto target cell lines. Media was replaced with DMEM + 10 ug/mL blasticidin, 2 ug/mL puromycin (A1113803, Invitrogen) at 24 h post-transduction and cells were cultured for 7 d. Clonal knockdown and overexpression cell lines were established by limiting dilution and with passage in selective media. RNA was purified from cell lines by TRIzol Reagent. Strand-specific total RNA with ribosomal RNA depletion was used for libraries generated with 1 µg of input RNA using the TruSeq Stranded mRNA Sample Prep Kit (Illumina) and sequenced on an Illumina NextSeq 2000 machine. Paired-end sequence reads were aligned to the human reference genome (hg38_v44) using STAR (38) and transcript quantification  was performed with RSEM (39) through the UMass Chan Medical School DolphinNext (40). Downstream analysis was performed in R version 4.4.0 using the RStudio interface. Relative transcript abundance was estimated using DESeq2 based on normalized counts of alignments, three replicates of parental A549-dCas9-VP64 were compared to clones containing the ADAR and B4GALNT2 constructs. Cutoff for significance was p < 0.1 adjusted for multiple comparisons.  Volcano plots were constructed using the EnhancedVolcano package.

first: Kyawe

last: Pyae

first: Liu

last: Ping

first: Jiang

last: Zhaozhao

first: Bradley

last: Evan

first: Cicuto

last: Thomas

first: Trombly

last: Melanie

first: Silverman

last: Nate

first: Fitzgerlad

last: Katherine

first: McDougall

last: William

first: Wang

last: Jennifer


title: Microbiota exposure following DSS-induced colitis promotes Mincle-mediated training of myeloid progenitors

city: Madrid

first: Manuel

last: Gomez

state: Madrid

address: Melchor Fernández Almagro, 3

country: Spain

institute: CNIC

summary: Impairment of the intestinal barrier allows the systemic translocation of commensal bacteria, inducing a pro-inflammatory state in the host. To explore this link, we investigated innate immune responses following increased gut permeability upon administration of dextran sulfate sodium (DSS) in mice. We found that microbiota translocation induced trained immunity (TI) in mouse myeloid bone marrow progenitors (BMPs). Enterococcus faecalis was isolated from bone marrow following DSS treatment and induced trained immunity hallmarks in bone marrow-derived mouse macrophages and human monocytes. Notably, DSS treatment or heat-killed E. faecalis reprogrammed BMPs, resulting in enhanced inflammatory responses in vitro and in vivo, and protection against pathogen infections.  The C-type lectin receptor Mincle (Clec4e) was essential for E. faecalis-induced TI in BMPs. Clec4e-/- mice showed impaired TI upon E. faecalis and reduced pathology following DSS treatment. Thus, Mincle sensing of E. faecalis induces TI that may contribute to pathologies associated with increased gut permeability. This SuperSeries is composed of the SubSeries listed below.


title: RNAseq  of GMP from mice treated with DSS, HK-EF or its excipient.

summary: Inflammatory bowel disease induced by treatment with DSS induce destruction of gut barrier inducing massive bacterial translocation to distal tissue. Our findings suggest that gut microbiota translocation induces trained immunity in Granulocyte-monocyte progenitors that can underlie pathologies related to low-grade systemic inflammation. The identification of E.faecalis translocation to the bone marrow as a main driver of this effect and its sensing by Mincle receptor in bone marrow progenitors offers potential targets for intervention in pathophysiological settings where gut permeability is altered. Moreover, intravenously and intranasal administration of HK-EF provided protection against systemic candidiasis in wild-type and Rag1-deficient mice lacking functional lymphocytes, indicative of innate immune-mediated protection.

overall_design: Three replicate samples (obtained pooling 3 biological replicates) of Granulocyte-monocyte progenitors from mice treated with DSS or HK-EF or its excipient were used to identify changes in gene expression using RNAseq

first: Iñaki

last: Robles-Vera

first: David

last: Sancho


title: Cut&Tag of GMP from mice treated with DSS, HK-EF or its excipient.

overall_design: Three replicate samples (obtained pooling 3 biological replicates) of Granulocyte-monocyte progenitors from mice treated with DSS or HK-EF or its excipient were used to identify changes in  H3K4me3 regions using Cut&Tag.


title: Cooperation of TRADD- and RIPK1-dependent Cell Death Pathways in Maintaining Intestinal Homeostasis [3 DPI colon]

city: Shanghai

first: ZIYU

last: SUN

address: HAIKE RD 100

institute: IRCBC

summary: Dysfunctional NF-kB signaling is critically involved in Inflammatory bowel disease (IBD). We investigated the mechanism by which RIPK1 and TRADD, two key mediators of NF-kB signaling, in mediating intestinal pathology using TAK1 IEC deficient model. We show that phosphorylation of TRADD by TAK1 modulates RIPK1-dependent apoptosis. TRADD and RIPK1 act cooperatively to mediate cell death mediated by TNF and TLR signaling. We demonstrate the pathological evolution from RIPK1-dependent ileitis to RIPK1- and TRADD-co-dependent colitis in TAK1 IEC deficient condition. Combined RIPK1 inhibition and TRADD knockout completely protect against intestinal pathology and lethality in TAK1 IEC KO mice. Furthermore, we identify distinctive microbiota dysbiosis biomarkers for RIPK1-dependent ileitis and TRADD-dependent colitis. These findings reveal the cooperation between RIPK1 and TRADD in mediating cell death and inflammation in IBD with NF-kB deficiency and suggest the possibility of combined RIPK1 kinase inhibition and TRADD knockout as a new therapeutic strategy for IBD.

overall_design:
To test the role of RIPK1 kinase in colitis  in TAK1 IEC-KO mice, we collected colon tissue for sequencing.
WT=Tak1f/f, HO=TAK1 IEC-KO, DHO=TAK1 IEC-KO Ripk1 D138N/D138N
DPI= days post induction


title: Cooperation of TRADD- and RIPK1-dependent Cell Death Pathways in Maintaining Intestinal Homeostasis

overall_design:
To test the role of RIPK1 kinase in ileitis  in TAK1 IEC-KO mice, we collected ileum tissue for sequencing.
WT=Tak1f/f, HO=TAK1 IEC-KO, DHO=TAK1 IEC-KO Ripk1 D138N/D138N
DPI= Days post induction


title: Cooperation of TRADD- and RIPK1-dependent Cell Death Pathways in Maintaining Intestinal Homeostasis [7 DPI colon]

overall_design:
To test the role of TRADD in colitis  in TAK1 IEC-KO mice, we collected colon tissue for sequencing.
WT=Tak1f/f Ripk1 D138N/D138N, HO=TAK1 IEC-KO Ripk1 D138N/D138N, DHO=TAK1 IEC-KO Ripk1 D138N/D138N Tradd IEC-KO
DPI= days post induction


title: Transcriptome changes in 2,3-butanedione induced bronchiolitis obliterans in rats

city: Jinan

first: Xu

last: Zhou

address: No. 42, Wenhua West Road, Jinan 250011, Shandong, China

institute: Affiliated Hospital of Shandong University of Traditional Chinese Medicine

summary: Bronchiolitis obliterans (BO) is a destructive fibrotic lung disease, which can be partly induced by 2,3-butanedione (also known as diacetyl, DA). However, the mechanism underlying the effects of DA on BO is not clear. In the present study, a bioinformatics analysis was performed using DA-treated or untreated lung tissues of rats.

overall_design: Male Sprague Dawley rats (age, 8 weeks;  weight, 300 g) received 1 week of environmental adaptation, and had ad libitum access to water and food.  The rats were randomly divided into the following two groups: i) Control group, intratracheal instillation of aseptic distilled water;  ii) DA group, intratracheal instillation of 125 mg/kg DA (Macklin Inc., Shanghai, China).  Rats in the DA group accepted DA intratracheal instillation once a day for 7 days. Twenty-four hours after the last intratracheal instillation, the rats were sacrificed and lung tissues with bronchi were collected. We then performed gene expression profiling analysis using data obtained from RNA-seq of six samples per group.

first: Xingyou

last: Zhao

first: Yanning

first: Baoqing

last: Zhang


title: ATAC-Seq based chromatin accessibilty profile of GMP from mice treated with DSS, HK-EF or its excipient.

summary: Inflammatory bowel disease induced by treatment with DSS induce destruction of gut barrier inducing massive bacterial translocation to distal tissue.  Our findings suggest that gut microbiota translocation induces trained immunity in Granulocyte-monocyte progenitors that can underlie pathologies related to low-grade systemic inflammation. The identification of E.faecalis translocation to the bone marrow as a main driver of this effect and its sensing by Mincle receptor in bone marrow progenitors offers potential targets for intervention in pathophysiological settings where gut permeability is altered. Moreover, intravenously and intranasal administration  of HK-EF provided protection against systemic candidiasis in wild-type and Rag1-deficient mice lacking functional lymphocytes, indicative of innate immune-mediated protection.

overall_design: Three replicate samples (obtained pooling 2 biological replicates) of Granulocyte-monocyte progenitors from mice treated with DSS or HK-EF or its excipient were used to identify changes in the chromatin accesibility profile using ATAC-seq.


title: TMEM106B deficiency leads to alterations in lipid metabolism and obesity in the TDP-43Q331K knock-in mouse model

status: Public on Jan 21 2025

city: Ithaca

first: Cha

last: Yang

address: 100 Fairview Sq

institute: Cornell University

summary: The TMEM106B gene, encoding a lysosomal membrane protein, is closely linked with brain aging and neurodegeneration. TMEM106B has been identified as a risk factor for several neurodegenerative diseases characterized by aggregation of the RNA-binding protein TDP-43, including frontotemporal lobar degeneration (FTLD) and limbic-predominant age-related TDP-43 encephalopathy (LATE). To investigate the role of TMEM106B in TDP-43 proteinopathy, we ablated TMEM106B in the TDP-43Q331K knock-in mouse line, which expresses an ALS-linked TDP-43 mutation at endogenous levels. We found that TMEM106B deficiency leads to glial activation, Purkinje cell loss, and behavioral deficits in TDP-43Q331K mice without inducing typical TDP-43 pathology. Interestingly, ablation of TMEM106B results in significant body weight gain, increased fat deposition, and hepatic triglyceride (TG) accumulation in TDP-43Q331K mice. In addition, lipidomic and transcriptome analysis shows a profound alteration in lipid metabolism in the liver of TDP-43Q331KTmem106b-/- mice. Our studies reveal a novel function of TMEM106B and TDP-43 in lipid metabolism and provide new insights into their roles in neurodegeneration.

overall_design: RNA-seq is used to identify the gene expression changes caused by TMEM106B deficiency and TDP-43 Q331K in mouse brain and liver.

first: Gwang

middle: B

last: Lee

first: Ling

last: Hao

first: Feng

middle: H

last: Hu


title: Analysis of the BadR regulon in Borrelia burgdorferi

city: Tampa

first: Zhiming

last: Ouyang

state: FL

address: 12901 Bruce B Downs Blvd

department: Molecular Medicine

institute: University of South Florida

summary: Borrelia burgdorferi, the causative agent of Lyme disease, relies on tightly coordinated gene expression to quickly adapt and survive in the tick vector and mammalian host. BadR, an ROK (repressor, open reading frame, kinase) family transcriptional regulator, binds directly to B. burgdorferi promoter DNA, however, many questions concerning the function of BadR in gene regulation remain unanswered. In particular, there are conflicting reports concerning what genes are regulated by BadR in B. burgdorferi. Furthermore, previous studies have suggested important roles for BadR in unfed ticks, but the BadR regulon has not been defined under such conditions. Additionally, although BadR regulates rpoS expression in a growth phase-dependent manner, it remained unknown whether BadR regulates other genes during different growth phases. To address these questions, we cultivated a B. burgdorferi badR mutant and wild-type strain under various conditions and analyzed the transcriptome using RNA-sequencing. When spirochetes were grown at 37°C and collected at the mid-logarithmic and stationary phase of growth, 211 and 272 genes were differentially expressed in the badR mutant, respectively. A total of 79 genes were differentially expressed when spirochetes were grown at 23°C. Combined, this work supports a role for BadR in global B. burgdorferi gene regulation by modulating expression of different sets of genes at different stages of the enzootic lifecycle.

overall_design: To investigate the genes regulated by BadR in Borrelia burgdorferi, a badR knockout was generated by homologous recombination. The badR mutant and wild-type strains were cultivated in vitro under conditions mimicking the unfed tick and mammalian host. Under mammalian host conditions, cells were collected at two growth phases to determine if BadR regulates genes in a growth-phase dependent manner.

first: Sierra

last: George


title: Overlapping and separable activities of BRA-2 and HIM-17 promote occurrence and regulation of pairing and synapsis during Caenorhabditis elegans meiosis

city: Pittsburgh

first: Uma

last: Chandran

state: PA

address: 5607 Baum Blvd

institute: University of Pittsburgh

summary: Faithful meiotic segregation requires pairwise alignment of the homologous chromosomes and their synaptonemal complex (SC) mediated stabilization. Here, we investigate on new factors that promote and coordinate these events during C. elegans meiosis. We identify BRA-2 (BMP Receptor Associated family member 2) as an interactor of HIM-17, previously shown to promote double-strand break formation. We found that loss of bra-2 impairs synapsis elongation without affecting homolog recognition, chromosome movement or SC maintenance. Epistasis analyses reveal previously unrecognized activities for HIM-17 in regulating homolog paring and SC assembly in a partially overlapping manner with BRA-2. We show that removing bra-2 or him-17 restores nuclear clustering, recruitment of PLK-2 at the nuclear periphery and abrogation of ectopic synapsis in htp-1 mutants, suggesting intact CHK-2-mediated signaling and presence of a barrier that prevents SC polymerization in absence of homology. Our findings shed new light on the regulatory mechanisms ensuring faithful pairing and synapsis.

overall_design: BRA-2::AID protein with germline-specific TIR1 driver was depleted with auxin for 24 hours prior to dissection of the germ line.  Controls were the TIR-1 driver alone and both lines without auxin

first: Judith

last: Yanowitz

first: Michelle

last: Scuzzarella

first: Nicola

last: Silva


title: Parasitic plasmids are anchored to inactive regions of eukaryotic chromosomes through a nucleosome signal [shotgun_sequencing]

city: Paris

first: Axel

last: Cournac

address: 28 rue du docteur Roux

country: France

institute: Institut Pasteur

summary: Natural plasmids are common in prokaryotes, but few have been documented in eukaryotes. The natural 2µ plasmid present in the yeast Saccharomyces cerevisiae is one of these best-characterized exceptions. This highly stable genetic element has coexisted with its host for millions of years, faithfully segregating at each cell division through a mechanism that remains unclear. Using proximity ligation methods (such as Hi-C, Micro-C) to map the contacts between 2µ plasmid and yeast chromosomes under dozens of different biological conditions, we found that the plasmid is tethered preferentially to regions with low transcriptional activity, often corresponding to long, inactive genes. These contacts do not depend on common chromosome-structuring factors, such as members of the structural maintenance of chromosome complexes (SMC) but depend on a nucleosome-encoded signal associated with RNA Pol II depletion. They appear stable throughout the cell cycle and can be established within minutes. This chromosome hitchhiking strategy may extend beyond the 2µ plasmid/S. cerevisiae pair, as suggested by the binding pattern of the natural eukaryotic plasmid Ddp5 along silent chromosome regions of the amoeba Dictyostelium discoideum.

overall_design: We explored the anchoring of parasitic plasmids with main chromosomes by performing Hi-C on various hosts and in different conditions for S.cerevisiae.

first: Fabien

last: Girard

first: Romain

last: Koszul


title: Spatial transcriptomics of murine eyelid

first: Sarah

middle: E

last: Millar

address: Icahn Building, Floor 13 Room 020C, 1425 Madison Ave

department: Developmental, Cell and Regenerative Biology

institute: Icahn School of Medicine at Mount Sinai

summary: Meibomian glands (MGs) are specialized holocrine sebaceous glands embedded in the tarsal plate of the eyelids. The cell heterogeneity of MGs is not well characterized. In this study, we performed a spatial transcriptomic analysis of murine eyelid using the 10X Visium platform.

overall_design: A pair of upper and lower eyelids from an 8-week-old male C57BL/6J mouse was dissected, sectioned at 10 μm, and mounted onto the assay slide containing four capture areas (6.5mm x 6.5mm) with spatially barcoded poly T capture probes. The subsequent procedures were conducted using the 10x Visium Spatial Gene Expression kit, according to the manufacturer’s instructions.

first: Xuming

last: Zhu

first: Mingang

last: Xu


title: snRNA-seq of murine tarsal plates

summary: The eyelid is a complex environment containing, containing epidermal, hair follicles, and conjunctival cells closely adjacent to the Meibomian glands (MGs), making it challenging to isolate pure MG cells or to identify MG contributions in bulk RNA-seq analyses. To analyze transcription in MG subpopulations, and to overcome difficulties in isolating cytoplasmic mRNAs from lipid-rich acinar cells, we utilized snRNA-seq which allows for identification of cell sub-populations within a complex mix.

overall_design: The dorsal portion of the eyelids and the surrounding tissues near the tarsal plates were removed using a scalpel. The tarsal plates were digested with collagenase A and the MGs were dissected using forceps. Nuclei were analyzed from the pooled samples of four 8-week-old and four 21-month-old male mice using two replicates for each age (16 mice analyzed in total).


title: Bulk RNA-sequencing of Meibomian glands from K5-rtTA tetO-GLI2DN mice and littermate controls

summary: The role of Hh signaling in adult Meibomian gland (MG) is unclear. In this study, we overexpressed an activated form of GLI2 (GLI2ΔN) in basal epithelial cells, including those in the MG, in a doxycycline-dependent manner. Four days post doxycycline treatment, GLI2ΔN+ MG acinar basal cells expand and replace differentiating PLIN2+ meibocytes. We laser-captured control and GLI2ΔN+ MG tissues and performed bulk RNA-seq. Our data showed that the most significantly upregulated genes were those involved in regulation of cell proliferation, such as Ki67 and Ccnd1. Expression of Hh target genes Gli1, Ptch1, and Hhip was also significantly elevated in GLI2ΔN-expressing MGs compared to controls. The most significantly downregulated genes were those associated with control of lipid metabolism such as Pparg, and meibocyte differentiation including Scd3, Scd4, and Plin2. Notably, cell populations expressing MG stem cell markers Lrig1 and Lgr6 were expanded upon GLI2ΔN expression in MGs. These data suggest that forced GLI2ΔN expression in MG epithelium promotes proliferation, expands the number of cells expressing Lrig1 and Lgr6, and, either directly or indirectly, suppresses meibocyte differentiation

overall_design: Three pairs of control and K5-rtTA tetO-GLI2ΔN mice were fed with doxycycline water for 4 days. Their eyelids were dissected and frozen sectioned. The MG tissues were laser-captured and collected for RNA extraction. The resulting RNA samples were subjected to bulk RNA-seq.


title: Identification of differential expression gene of LncRNA interacting with NLRP3 protein in RAW264.7 cells infected with Salmonella typhimurium and uninfected cells

city: Beijing

first: Zhiyuan

last: An

address: No.8 gongti south Road

department: Medical Research Center

institute: Beijing Chaoyang Hospital

summary: We detected the differential expression gene of LncRNA interacting with NLRP3 protein in RAW264.7 cells infected with Salmonella typhimurium and uninfected using RNA immunoprecipitation sequencing

overall_design: Collect suspension of infected Salmonella typhimurium and uninfected RAW264.7 cells, extract total RNA directly interacting with NLRP3 protein in cells using RNA immunoprecipitation method, and then perform LncRNA sequencing.

first: Wenyi

last: Ding


title: ZNF252P promotes tumorigenesis by activating c-Myc-mediated positive feedback loop

city: Nanjing

first: Erbao

last: Zhang

address: longmian da dao

department: Department of Epidemiology

institute: Nanjing Medical University

summary: Here we screened and identified a novel oncogenic lncRNA, ZNF252P, which is upregulated in a variety of cancer types. ZNF252P promotes cancer cell proliferation and tumorigenicity in vitro and in vivo by activating c-Myc-mediated positive feedback loop. The pivotal regulatory axis of ZNF252P/c-Myc gives it the potential to be the promising biomarker and therapeutic target in cancer development.

overall_design: RNA-seq analysis was performed after knockdown of ZNF252P in A549 cell line.

first: Tianyu

last: Qu

first: Chang

first: Xiyi

last: Lu


title: Cell cycle checkpoint proficiency of endometrial cancer cells [WES]

city: Boston

middle: J.

last: Hill

address: 450 Brookline Ave., Dana Building, Room 1410C

department: Medical Oncology

institute: Dana-Farber Cancer Institute

type: Genome variation profiling by high throughput sequencing

summary: Endometrial cancers (EC) are the most common gynecologic malignancy in the US. Most ECs harbor limited targetable somatic alterations, and are often grouped by histology (endometrioid, serous, or clear cell), mismatch repair, or TP53 status, none of which perfectly predict therapeutic response. A better mechanistic understanding of the key functional defects in ECs and more therapies with which to engage those targets in advanced stage EC are needed. Here we utilize functional, transcriptomic and genomic assays on a panel of EC cell lines and patient-derived organoids across EC histologic and genomic subtypes to characterize the TP53 and RB1 cell cycle regulatory proficiency and therapeutic vulnerabilities in this disease. We were surprised to find that TP53 genomic and functional status has little predictive capacity for EC therapeutic response. Rather, RB regulatory status correlated better with response to G1/S targeted therapies.

overall_design: For both cell lines and organoids, the cells were grown to 80% confluence in regular growth medium. Cell lines were trypsinized and organoids were scraped, and the pellets were stored at -80°C until DNA was prepared. DNA was prepared using Qiagen's DNA Mini kit (Qiagen Cat. # 51304) according to the manufacturer's protocol. DNA was snap frozen and sent to Novogene (Sacramento, CA) for standard whole exome sequencing according to their protocol. Specifically, at Novogene, Agilent SureSelect Human All Exon V6 was utilized for library preparation. Sequencing occurred on the NovaSeq 6000 with a PE150 sequencing strategy.

first: Helena

last: Jun


title: In vitro reconstitution of meiotic DNA double-strand break formation

status: Public on Jan 20 2025

first: Yu

last: Liang

address: Yueyang Road 320

department: SIBCB

institute: CEMCS, CAS

summary: The Spo11 complex catalyzes the formation of DNA double-strand breaks (DSBs), initiating meiotic recombination-a process essential for fertility and genetic diversity. Although Spo11’s function has been known for 27 years, previous efforts to reconstitute DSB formation in vitro have been unsuccessful. Here, we biochemically characterize mouse SPO11 and TOP6BL protein complex and demonstrate that this complex cleaves DNA and covalently attaches to the 5' terminus of DNA breaks in vitro. Using a point-mutation strategy, we reveal that Mg2+ is essential for DNA cleavage activity of this complex in vitro, as confirmed by knock-in mice carrying a point mutation in SPO11 that disrupts its binding to Mg2+, thereby abolishing DSB formation. However, the activity of the SPO11 complex is ATP-independent. We also present evidence that the mouse SPO11 complex is biochemically distinct from the ancestral topoisomerase VI. Our findings establish a mechanistic framework for understanding the initial steps of meiotic recombination.

overall_design: We employed END-seq to map DNA cleavage sites with high resolution, aiming to elucidate whether the SPO11-TOP6BL complex exhibits DNA sequence specificity in its activity.

first: Xinzhe

last: Tang

first: Zetao

last: Hu

first: Jian

first: Meixia

last: Wu

first: Ping

last: Guan

first: Yawei

last: Song

first: Yue

last: Yin

first: Wei

first: Jinbiao

last: Ma

first: Ying

last: Huang

first: Ming-Han

last: Tong


title: Exploring the mechanism of transcription factor TAF7 promoting the progression of triple-negative breast cancer

city: Wuhan

first: 婉君

last: 张

state: Hubei

address: hongshan street

department: College of Life Science and Technology

institute: huazhong university of science and technology

summary: To further study the regulation of TAF7 on cells, the transcriptome sequencing on MDA MB-231 cells, MDA MB-231/shTAF7 cells, MDA 231-LM2 and MDA 231-LM2/shTAF7 cells were performed.

overall_design: We then performed gene expression profiling analysis using data obtained from RNA-seq of MDA MB-231 cells, MDA MB-231/shTAF7 cells, MDA 231-LM2 and MDA 231-LM2/shTAF7 cells to find the genes regulated by TAF7.

first: Wanjun

first: Jun

last: Wang


title: IL-16 promotes anti-tumor immunity via establishing a Th1-dominant tumor microenvironment [RNA-seq]

city: Hangzhou

first: Peng

last: Xiao

address: No.3 East Qingchun Road

institute: Zhejiang University

summary: Overcoming immunosuppression in tumor microenvironment (TME) is fundamental to the development of novel cancer immunotherapies. Herein, we revealed an unrecognized role of IL-16 in shaping anti-tumor immunity. Compared with healthy subjects, cancer patients had impaired production of IL-16, which was correlated with inferior patient prognosis. In multiple murine cancer models, IL-16 administration augmented the anti-tumor immune responses and thus restrained tumor growth. Mechanistically, IL-16 potentiated the polarization of T helper 1 (Th1) cells and the production of their effector cytokine IFN-γ, Mechanistically, IL-16 blocked glutamine catabolism by downregulating glutaminase (GLS) expression in CD4+ T cells. The IL-16-established Th1 tumor microenvironment further increased the expression of CXCR3 ligands in tumor-associated macrophages, which improved the therapeutic efficacy of immune checkpoint blockade (ICB). In cancer patients who received anti-PD1 therapy, high IL-16 levels correlated with better responsiveness. Finally, we found that the impaired production of histamine by mast cells was a causative factor for IL-16 downregulation in TME. Thus, IL-16 administration may serve as a potential approach to augment anti-tumor immunity, and also improve the outcome of ICB therapy in cancer patients. Collectively, we provided new insights into the biological function of IL-16 and thus highlighted its potential clinical value in cancer immunotherapy.

overall_design: Naïve CD4+ T cells isolated from mouse spleen were culture under Th1 condition in the presence of PBS or IL-16. Five days later, cells were harvested and subjected to RNA-sequencing analysis. Gene expression levels were compared between PBS-treated or IL-16-treated Th1 cells.


title: Cell cycle checkpoint proficiency of endometrial cancer cells [bulk RNA-Seq]

first: Sarah

overall_design:
For cell lines, doubling times were calculated. For HEC1B and ARK1 models, cells were treated for one doubling time (approximately 26 hours for ARK1 and 33.5 hours for HEC1B) with either 0.25µM Abemaciclib methanesulfonate (referred to as "Abemaciclib" henceforth) or media containing DMSO vehicle (used for analysis as batch 1). For organoids, a subset of the organoids were treated with media containing either 0.25µM Abemaciclib or media containing DMSO vehicle for 24 hours (used for analysis as batch 2). For additional cell lines and organoids to be used only for baseline transcriptomic comparisons, the cells were treated with media containing DMSO at 1:100,000 for 24 hours (used for analysis as batch 3). After treatments, cell lines and organoids were harvested and stored at -80°C until RNA was prepared. RNA was prepared using Qiagen's RNEasy Mini kit (Qiagen Cat #74104) with on-column DNASE digest (Qiagen Cat. #79254) according to the manufacturer's protocol. RNA was snap frozen and sent to GENEWIZ (South Plainfield, NJ) for Standard RNA sequencing. Each individual sample was sequenced in duplicate to provide technical replicates for validation. The following library preparation and sequencing methods in quotation marks are provided verbatim by GENEWIZ and may represent a standard pipeline used there.

"RNA samples were quantified using a Qubit 2.0 Fluorometer (Life Technologies, Carlsbad, CA, USA), and RNA integrity was checked using an Agilent TapeStation 4200 (Agilent Technologies, Palo Alto, CA, USA).

RNA sequencing libraries were prepared using the NEBNext Ultra II RNA Library Prep Kit for Illumina using the manufacturer's instructions (NEB, Ipswich, MA, USA). Briefly, mRNAs were initially enriched with Oligod(T) beads. Enriched mRNAs were fragmented for 15 minutes at 94 °C. First-strand and second-strand cDNA were subsequently synthesized. cDNA fragments were end-repaired and adenylated at 3'ends, and universal adapters were ligated to cDNA fragments, followed by index addition and library enrichment by PCR with limited cycles. The sequencing library was validated on the Agilent TapeStation (Agilent Technologies, Palo Alto, CA, USA), and quantified by using Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA) as well as by quantitative PCR (KAPA Biosystems, Wilmington, MA, USA).

The sequencing libraries were clustered on a flow cell. After clustering, the flow cell was loaded on the Illumina instrument according to the manufacturer’s instructions. The samples were sequenced using a 2x150bp Paired-End (PE) configuration, targeting 30 M reads/sample. The control software conducted image analysis and base calling. Raw sequence data (.bcl files) generated by the sequencer were converted into fastq files and de-multiplexed using Illumina's bcl2fastq 2.20 software. One mismatch was allowed for index sequence identification."


title: The regulatory mechanism of chlorogenic acid on fatty acid metabolism in human embryonic stem cells

city: Wenzhou

first: LIjun

last: Chen

state: Other

address: Wenzhou Medical University, Wenzhou City, Zhejiang Rrovince,China

institute: Wen Zhou Medical University

summary: Chlorogenic acid (CGA) is a crucial bioactive dietary polyphenol found in plants and serves as the primary ingredient in numerous traditional Chinese medicine preparations. Due to its beneficial impact on treating metabolic syndrome, it has garnered significant attention in medical research. Previous studies have demonstrated that chlorogenic acid possesses antioxidative, antibacterial, antiviral, anti-tumor, lipid-lowering, glucose-lowering, and immunomodulatory properties, indicating its potential role in regulating metabolism. Human embryonic stem cells (hESCs) exhibit self-renewal and multi-directional differentiation capabilities. Any element within the culture system can alter the microenvironment and influence hESC metabolism. Therefore, hESCs provide a valuable and high-throughput model for investigating CGA's regulatory mechanism on metabolism in vitro. While previous studies have explored CGA's important regulatory role in human metabolism, its specific impact on hESCs metabolism remains undisclosed. Metabolites of hESCs are critical targets for enhancing stem cell function regulation. Prior research has emphasized the essential nature of de novo synthesis and β-oxidation of fatty acids for regulating hESCs; however, CGA's roles and mechanisms in regulating fatty acid metabolism within hESCs remain unelucidated. This study utilizes transcriptome analysis combined with epigenetics to investigate CGA's regulation of fatty acid metabolism within hESCs while analyzing the molecular mechanisms involved—providing a theoretical foundation for future regulation of HESC metabolism by CGA and offering potential optimizations for hESCs culture systems.

overall_design: In the RNA-seq analysis on hESCs, ~50 cells were used for per reaction and two or three replicates were performed for each group.  In the CUT&Tag on hESCs, ~10∧3 cells were used for per reaction and two or three replicates were performed for each group.

first: Wenpin

last: Cai

first: Ming

last: Zong

first: Jingzhang

last: Ji

first: Lijun


title: Histone H3 lysine 9 tri-methylation is associated with pterygium

status: Public on Jan 19 2025

city: Daegu

first: Hong-Yeoul

last: Ryu

state: Buk-Gu

address: Daehak-ro 80

department: Biotechnology

country: South Korea

institute: Kyungpook National University

summary: Pterygium, abnormal growths of conjunctival tissue onto the cornea, are common ocular surface conditions with a high risk of recurrence after surgery and potential ophthalmic complications. The exact cause of pterygium remains unclear, and the triggers are still unknown. In this study, we investigate the epigenetic profiles of patients with pterygium, focusing on histone H3 lysine 4 (H3K4) and lysine 9 (H3K9) trimethylation (me3). While H3K4me3 levels showed no significant genome-wide change, they were significantly altered in genes related to development and ocular diseases. Conversely, H3K9me3 levels were markedly elevated genome-wide, particularly at the promoters of 82 genes involved in developmental pathways. Furthermore, we identify six genes, ANK2, AOAH, CBLN2, CDH8, CNTNAP4, and DPP6, with decreased gene expression correlated with substantially increased H3K9me3, suggesting their potential as biomarkers for pterygium. This study represents the first report linking histone modification to pterygium progression, providing valuable insights into therapeutic strategies and potential drug targets.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone modification H3K4me3 and H3K9me3 in conjunctiva and pterygium samples.

first: Dahee

last: Choi

first: Ann-Yae

first: Seok-Won

last: Jeoung

first: Yun-Hee

first: Nayoon

last: Park

first: Hyun-Sun

first: Hyun-Shik

last: Lee

first: Dong-Hyung

last: Cho

first: Dong

last: Kim


title: MyoD1 localization at the nuclear periphery is mediated by association of WFS1 with active enhancers

city: Vienna

first: Roland

last: Foisner

address: Dr.-Bohr-Gasse 9

department: Center for Medical Biochemistry

institute: Max Perutz Labs, Medical University of Vienna

summary:
Spatial organization of the mammalian genome influences gene expression and cell identity. While association of genes with the nuclear periphery is commonly linked to transcriptional repression, also active, expressed genes can localize at the nuclear periphery. The transcriptionally active MyoD1 gene, a master regulator of myogenesis, exhibits peripheral localization in proliferating myoblasts, yet the underlying mechanisms remain elusive. Using a newly generated reporter cell line, we demonstrate here that peripheral association of the MyoD1 locus is independent of mechanisms involved in heterochromatin anchoring. We identify a set of nuclear envelope transmembrane proteins, particularly WFS1, that actively tether MyoD1 to the nuclear periphery. WFS1 primarily associates with active distal enhancer elements upstream of MyoD1, and with a subset of enhancers enriched in active histone marks genome-wide, which are linked to expressed myogenic genes. Overall, our data identify a novel mechanism involved in tethering active genes to the nuclear periphery.

This research was funded in whole or in part by the Austrian Science Fund (FWF) [P29713-B28, P32512-B and P36503-B] to Roland Foisner and a doctorate program funded by the Austrian Science Fund (FWF) [W1261-B28].

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for WFS1 in wildtype and WFS1-FlagTag expressing C2C12 myoblasts in the proliferating stage sonicated for 30 and 36 cycles.

first: Konstantina

last: Georgiou

first: Fatih

last: Sarigol

first: Tobias

last: Nimpf

first: Christian

last: Knapp

first: Daria

last: Filipczak

first: Nana

last: Naetar


title: Integrative proteo-transcriptomic characterization of advanced fibrosis in chronic liver disease across etiologies

city: Solna

first: Hong

last: Yang

state: Stockholm

address: Tomtebodavägen 23A

department: Systems Biology

country: Sweden

institute: KTH Royal Institute of Technology/Science For Life Laboratory

summary: While technological advancements have improved our understanding of fibrogenesis, identifying advanced fibrosis in the general population remains challenging due to clinical heterogeneity influenced by underlying causes, comorbidities, and lifestyle factors. A systems-level characterization of metabolic and signaling dysregulation could effectively capture the signatures driving liver fibrosis across different etiologies. In this study, we utilized a data-driven multi-omics approach, encompassing liver transcriptomics and plasma proteomics, to thoroughly characterize patients across the pathological spectrum, from early-stage fibrosis to cirrhosis and associated HCC.

overall_design: 178 snap-frozen explant liver samples and/or biopsies were processed for RNA sequencing on the Illumina NovaSeq6000 system.

first: Mujdat

last: Zeybel

first: Dila

last: Atak

first: Adil

last: Mardinoglu


title: Drosophila Modulo is Essential for Transposon Silencing and Developmental Robustness.

city: Charleston

first: Vamsi

last: Gangaraju

address: 173 Ashley Ave

institute: Medical University of South Carolina

type: Non-coding RNA profiling by high throughput sequencing

summary: Transposable element (TE) silencing in the germline is crucial for preserving genome integrity; its absence results in sterility and diminished developmental robustness. The Piwi-interacting RNA (piRNA) pathway is the primary small non-coding RNA mechanism by which TEs are silenced in the germline. Three piRNA binding proteins promote the piRNA pathway function in the germline– P-element-induced wimpy testis (Piwi), Aubergine (Aub), and Argonaute 3 (Ago3). Piwi mediates transcriptional silencing of TEs by promoting the deposition of the heterochromatin mark Histone 3 lysine nine trimethylation (H3K9me3) at TE genomic sites. Aub and Ago3 facilitate post-transcriptional silencing of TEs. Proteins and mechanisms that promote piRNA function in TE silencing are still being discovered. This study demonstrates that the Drosophila Modulo protein, a homolog of mammalian Nucleolin and an epigenetic regulator, is crucial for the enrichment of H3K9me3 at TEs. We show that Modulo interacts with Piwi and operates downstream of the Piwi-piRNA complex's entry into the nucleus. Lack of Modulo function impairs Piwi-interacting protein Panoramix's ability to target transposon RNAs. Furthermore, the reduced function of Modulo in the mother undermines developmental robustness and exacerbates neomorphic Kr[If-1]-induced ectopic eye outgrowths in the offspring. Maternal Modulo enhances developmental robustness by inhibiting TE activation and transcriptome variability associated with intrinsic genetic variation. Thus, Modulo is an essential component of the mechanism that operates in the maternal germline to facilitate TE silencing and ensure developmental robustness in the ensuing generation.

overall_design:
RNAseq profiling of mod[07570/L8], mod[07570]/+, and mod[L8]/+ ovaries to test the role of the Modulo protein in transposon silencing
RNAseq profiling of heterozygous control mod[07570]/+, and transheterozygous mod[07570]/[L8] heads to test the role of the Modulo protein in transposon silencing in the heads
RNAseq profiling of mod[07570]/[L8] heads in the presence and absence of ectopic outgrowths.
ChIP-seq of heterochromatin mark H3K9me3 from transheterozygous mod[07570]/[L8] and heterozygous control mod[07570]/+
Sequencing of Piwi-bound small RNAs in transheterozygous mod[07570]/[L8] and heterozygous control mod[07570]/+.


title: Human MAIT10 and MAIT17 profiles are effector states directed by the cytokine milieu

city: Huddinge

first: Johan

last: Sandberg

address: Alfred Nobels allé 8

department: Department of Medicine

institute: Karolinska Institutet

summary: Mucosa-associated invariant T (MAIT) cells are unconventional MR1-restricted T cells with rapid innate-like response characteristics. MAIT cells can mediate broad effector functions, including bacterial cytolysis and production of a range of pro-inflammatory cytokines. However, how the MAIT cell antigen response profile is controlled is unclear. Incorporating functional and transcriptomic analyses we define three distinct MAIT cell effector programs driven by the cytokine milieu during antigen recognition. Activation by TCR signaling together with IL-12 or IL-23 drives a program biased towards IL-10 through the transcription factor c-MAF and the expression of the inhibitory receptors TIM-3, LAG-3 and PD-1 (MAIT10). Co-activation with IL-18 supports responses dominated by IL-17, GM-CSF, IFNγ and TNF (MAIT17). The IL-18-driven inflammatory and tissue-repair program vetoes IL-10 expression. On the contrary, TCR activation without cytokine co-activation drives primarily cytolytic arming. These activation states are transient and reflect the adaptation of MAIT cells to the local milieu. MAIT cell IL-10 production serves an autocrine regulatory function in addition to influence the state of monocytes. Finally, in active Crohn’s disease patients, the MAIT10 profile is suppressed. Together, these findings demonstrate that MAIT cells adapt their effector response programs to the local cytokine milieu.

overall_design:
Single cell sequencing of FACS-sorted peripheral MAIT cells unstimulated or stimulated with 5-OP-RU alone or in combination with IL-12, IL-18 or IL-23.
NOTE FROM SUBMITTER: Pease note that, due to the privacy of human donors, in accordance with EU GDPR legislation the raw data is not provided

first: Caroline

last: Boulouis

first: Elli

last: Mouchtaridi

first: Thomas

last: Müller

first: Jeffrey

middle: Y

last: Mak

first: David

middle: P

last: Fairlie

first: Peter

last: Bergman

first: Jakob

last: Michaëlsson

first: Jonas

last: Halfvarsson

first: Jenny

last: Mjösberg

first: Marcus

last: Buggert

middle: K


title: Effects of AIF3 on gene expression in MEF cells

city: Dallas

first: Yingfei

last: Wang

state: TX

address: 5323 Harry Hines BLVD

department: Pathology

institute: UT Southwestern Medical Center

summary: AIF3 splicing differentially affects gene expression involved in different pathways.

overall_design: To study the effect of AIF3 splicing on gene expression, MEF cells were isolated from AIF3 floxed embryos and infected with or without CRE lentivirus. Single-cell clones expressing either AIF or AIF3 splicing were established as WT and AIF3 cell lines, respectively. The AIF3 single clone C3 was used in this study. Additionally, a rescue cell line expressing exogenous AIF in AIF3 C3 clone, designated as AIF3-AIF, was established.

first: Mi

last: Zhou

first: Weibo

last: Luo


title: Epigallocatechin Gallate (EGCG) Modulates Senescent Endothelial Cell-Monocyte Communication in Age-Related Vascular Inflammation

status: Public on Jan 18 2025

city: Toronto

first: Kai

middle: Howson

last: Ellis

state: ON

address: 686 Bay St

department: Genetics and Genome Biology

country: Canada

institute: The Hospital for Sick Children

summary: Aging significantly affects intercellular communication between vascular endothelial cells (ECs) and hematopoietic cells, leading to vascular inflammation and age-associated diseases. This study determined how senescent ECs communicate with monocytes, whether extracellular vesicles (EVs) released from senescent ECs affect monocyte functions, and investigated the potential for epigallocatechin-3-gallate (EGCG), a flavonoid in green tea, to reverse these effects. Human umbilical vein endothelial cells (HUVECs) were treated with Etoposide (10 µM, 24h) to induce senescence, followed by EGCG (100 µM, 24h) treatment to evaluate its potential as a senotherapeutic agent. The interaction between ECs and monocytes was analyzed using a co-culture system and direct treatment of monocytes with EC-derived EVs. EGCG reduced senescence-associated phenotypes in ECs, as evidenced by decreased senescence-associated (SA)-β-Gal activity and reversal of Etoposide-induced senescence markers. Monocytes co-cultured with EGCG-treated senescent ECs showed decreased pro-inflammatory responses compared to those co-cultured with untreated senescent ECs. Additionally, senescent ECs produced more EVs than non-senescent ECs. EVs from senescent ECs enhanced lipopolysaccharide (LPS)-induced pro-inflammatory activation of monocytes, whereas EVs from EGCG-treated senescent ECs mitigated this activation, maintaining monocyte activation at normal levels. Our findings reveal that EGCG confers anti-senescent effects via modulation of the senescent EC secretome (including EVs) with the capacity to modify monocyte activation. These findings suggest that EGCG could act as a senotherapeutic agent to reduce vascular inflammation related to aging.

overall_design: RNA-seq profiling was conducted on THP1 monocytes in monoculture and co-cultured with control endothelial cells (ECs), senescent endothelial cells (senECs), or EGCG-treated senescent endothelial cells for 24 hours, followed by a 2-hour lipopolysaccharide (LPS) treatment.

first: Sarvatit

last: Patel


title: Macrophage responses to zinc-limited and zinc-replete Mycobacterium tuberculosis H37Rv

status: Public on Jan 17 2025

city: Honolulu

first: Sladjana

last: Prisic

state: HI

address: 1800 East-West Rd

institute: University of Hawaii

summary: The goal of this experiment was to determine if macrophage responses differ between macrophages infected with M. tuberculosis H37Rv grown in the standard, zinc-replete medium compared to cells grown in zinc-limited medium. The macrophage responses after 4 hours and 24 hours of infection were determined by isolating the RNA and analyzed it with the NanoString panel mouse host response panel.

overall_design: There were 15 samples total: RAW 264.7 macrophages infected with zinc-limited M. tuberculosis after 4 hours of infection, RAW 264.7 macrophages infected with zinc-replete M. tuberculosis after 4 hours of infection, uninfected RAW 264.7 macrophages after 24 hours, RAW 264.7 macrophages infected with zinc-limited M. tuberculosis after 24 hours of infection, RAW 264.7 macrophages infected with zinc-replete M. tuberculosis after 24 hours of infection, all in biological triplicate.

first: Endrei

last: Marcantonio


title: Interleukin-16 Establishes a Th1-dominant Tumor Immune Microenvironment And Potentiates Immune Checkpoint Therapies [scRNA-Seq]

first: Peng

last: Xiao

summary: Overcoming immunosuppression in tumor microenvironment (TME) is crucial for the development of novel cancer immunotherapies. In this study, we revealed a previously unrecognized role of IL-16 in shaping anti-tumor immunity. Compared to healthy individuals, cancer patients exhibited impaired production of IL-16, which was associated with inferior patient prognosis. In multiple murine cancer models, IL-16 administration augmented the anti-tumor immune responses and thus restrained tumor growth. Further investigation uncovered that IL-16 potentiated the polarization of T helper 1 (Th1) cells and the production of their effector cytokine IFN-γ. Mechanistically, IL-16 inhibited glutamine catabolism by downregulating the expression of glutaminase (GLS) in CD4+ T cells. The establishment of IL-16-dependent Th1 tumor microenvironment further increased the expression of CXCR3 ligands in tumor-associated macrophages (TAMs), thereby improving the therapeutic effectiveness of immune checkpoint blockade (ICB). In cancer patients who received anti-PD1 therapy, higher levels of IL-16 were correlated with better responsiveness. Finally, we found that the impaired production of histamine by mast cells was a contributing factor to the downregulation of IL-16 in TME. Therefore, IL-16 could potentially be utilized as a therapeutic approach to augment anti-tumor immunity, and improve the outcome of ICB therapy in cancer patients. Collectively, our research provided new insights into the biological function of IL-16, emphasizing its potential clinical significance in cancer immunotherapy.

overall_design: Fresh breast cancer tissues and the corresponding normal tissues were harvested, cell-specific gene expression was analyzed by scRNA-Seq.


title: Single-cell multiomics study of cell-type-specific neuron activation unravels context-dependent gene regulation of brain disorders

city: Evanston

first: Jubao

last: Duan

state: IL

address: 1001 University Place

department: Center for Psychiatric Genetics

institute: NorthShore University HealthSystem

summary: Genome-wide association studies (GWAS) have identified many risk genes for neuropsychiatric disorders (NPD) such as schizophrenia (SCZ). However, functional interpretation of these GWAS findings remains challenging; a major hurdle is that polygenic risk may manifest its effect conditionally upon neural activation, i.e., context-dependent. Human induced pluripotent stem cell (hiPSC)-derived neurons are a tractable cellular model for ascertaining context-dependent polygenic effects, e.g., neural activation that may mimic the physiological effects of environmental stimuli. Here, we modelled neural activation by depolarisation using high potassium chloride (KCl) in co-cultured excitatory/inhibitory neurons of 100 hiPSC lines (28 from SCZ donors), followed by assaying single-cell multiomes (scRNA/ATAC-seq) of over 700,000 nuclei from neurons at 0 hr (unstimulated), 1 hr (early response), and 6 hr (later response) of high KCI exposure. We linked genes with open chromatin peaks for each main neural subtype (GABAergic inhibitory, NEFM+ or NEFM-excitatory neurons) and confirmed a significant correlation between peak accessibility and target gene expression. Compared to static genes/peaks, dynamic ones, specifically those activity-upregulated, were more enriched for NPD GWAS risk, with the most robust enrichment for SCZ. Based on the dynamic gene expression pattern across the three time points, we found NPD risk genes tend to be continuously upregulated (i.e., with up-up dynamics). We further analysed context-specific SCZ-associated differentially expressed genes (DEGs) using MAST for single neurons of 28 SCZ cases and 72 controls. We found that 3-5%, 4-6%, and 15-23% of genes were SCZ-associated DEGs in NEFM+ excitatory, NEFM- excitatory, and GABAergic neurons, respectively, most of which were cell activity-specific. Notably, we found that SCZ-associated DEGs in high-activity neurons were more enriched for synapse-related GO terms and NPD risk genes. Interestingly, upstream regulatory sequences of the stimulation-specific SCZ-associated DEGs were enriched for binding sites of TCF4, which is a strong GWAS risk gene and was considered the master regulator of other SCZ-related genes. Moreover, the upregulated genes in SCZ cases in the stimulated (only at 6hr) NEFM+ excitatory neurons showed strong enrichment for GO terms related to cholesterol synthesis. Single-neuron DEG analysis for 28 SCZ cases and 28 matched controls gave similar results. Our study suggests that many NPD genes may only elicit disease-relevant effects upon neuronal activation, providing novel insights into how polygenic risk factors interact with environmental stimuli for NPD.

overall_design: Raw FASTQ files complies 10x Genomics multiome scARC (scRNA-seq + scATAC-seq) standard. Library generated using cultured glutamatergic and GABAergic neurons derived from iPS cells with Chromium Next GEM Single Cell Multiome ATAC + Gene Expression Reagent Kit. The output folders, including all matrices files, of Cell Ranger 2.0.2 were packaged and compressed sample-wise in according to their BioSample and SRA metadata

first: Siwei

last: Zhang


title: Stimulation of TRPV1+ sensory nerves at nape induces significant alterations in gene expression in the spleen

first: Zijing

last: Liu

address: Fengtai District, #20 East St.

institute: Beijing Institute of Biotechnology

summary: RNA-seq analysis was performed to characterize the transcriptional changes in the spleen triggered by distinct treatments. Capsaicin (CAP) treatment not only induced anti-inflammatory effects in inflamed states but also inhibited the immune response under normal physiological conditions.

overall_design: RNA-seq analysis was performed to characterize the transcriptional changes in the spleen triggered by distinct treatments, including saline control, CAP alone, LPS alone or CAP-LPS treatment.

first: Dengcen

last: Song

first: Zheng

last: Cao


title: Transcriptional response in RAW 264.7 macrophages infected with zinc-replete and zinc-limited Mycobacterium tuberculosis

summary: The goal of this experiment was to determine if macrophages can distinguish between M. tuberculosis grown in standard, zinc-replete media and those grown in zinc-limited media. In order to determine this, we performed RNA-sequencing (RNA-seq) of RAW 264.7 macrophages infected with zinc-limited Mtb and zinc-replete Mtb at 4 hours post-infection (hpi) and 24hpi. Transcriptional response in the infected macrophages was compared to uninfected controls (macrophages exposed to growth media alone) and between macrophages infected with zinc-limited Mtb and zinc-replete Mtb at each time point.

overall_design: RNA was extracted and analyzed from 24 different samples: RAW 264.7 macrophages infected with zinc-replete and zinc-limited M. tuberculosis, and uninfected macrophage controls (added zinc-replete and zinc-limited media without bacteria) after 4 hours and 24 hours of infection, all in biological triplicates.

first: Yuanyuan

last: Fu

first: Vedbar

last: Khadka

first: Youping

last: Deng


title: Noncoding RNA profiling in omentum adipose tissue from obese patients and the identification of novel metabolic biomarkers

city: jinan

first: yongjiao

last: zhang

state: shandong

address: jingshi

institute: shandong second medical university

summary: Obesity, a prevalent metabolic disorder, is linked to perturbations in the balance of gene expression regulation. Noncoding RNAs (ncRNAs), including long noncoding RNAs (lncRNAs), circular RNAs (circRNAs), and microRNAs (miRNAs), play pivotal roles in regulating gene expression. The aim of this study was to identify additional ncRNA candidates that are implicated in obesity, elucidating their potential as key regulators of the pathogenesis of obesity. In our research, compared with those in healthy individuals, various lncRNAs, circRNAs, and miRNAs were significantly differentially expressed in obese subjects. We established comprehensive ncRNA profiles from human greater omentum adipose tissue and compared the profiles between obese individuals with healthy controls via whole-transcriptome sequencing. Subsequent validation via qRT‒PCR substantiated the sequencing results, reinforcing the reliability of our findings. These insights not only validate the importance of ncRNAs but also lay a theoretical foundation for further functional exploration of the roles of lncRNAs, circRNAs, and miRNAs in obesity, potentially offering novel perspectives on the pathogenesis of this condition.

overall_design: We detected differentially expressed ncRNAs in the greater omentum of obese and healthy individuals by whole transcriptome sequencing. Then we performed GO analysis, KEGG pathway analysis, real-time quantitative polymerase chain reaction (qRT-PCR), Protein-protein interaction (PPI) and hub gene analysis.

first: Yongjiao

first: Ao

last: chen

first: Fei

last: Gao

first: Sumei

last: Lu


title: Mapping snoRNA-target RNA interactions in an RNA binding protein-dependent manner with chimeric eCLIP [chimeric eCLIP]

status: Public on Jan 16 2025

city: Houston

first: Eric

last: Van Nostrand

address: 1 Baylor Plaza Rm T334

department: Therapeutic Innovation Center

institute: Baylor College of Medicine

summary: To indentify the targets of snoRNAs, we adapted an enhanced chimeric eCLIP approach,utilizing both established core proteins and lesser-known snoRNA-associated RBPs as bait. .

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: We perform chimeirc eCLIP using C/D box snoRNA binding proteins (FBL, NOP56, NO58) as bait in both mouse ESCs and human cell lines and H/ACA snoRNA binding proteins (DKC1, GAR1) as bait in human cell lines; we also performed chimeric eCLIP with the accessory snoRNA binding proteins (LARP7, NOLC1, WDR43, UPT18, ZC3H8, SMNDC1) to capture snoRNA interactions.

first: Zhuoyi

first: Bongmin

last: Bae

first: Simon

last: Schnabl

last: Yuan

first: Thareendra

last: De Zoysa

first: Maureen

last: Akinyi

first: Charlotte

last: Le Roux

first: Karine

last: Choquet

first: Amanda

last: Whipple


title: Mapping snoRNA-target RNA interactions in an RNA binding protein-dependent manner with chimeric eCLIP [eCLIP]

summary: To identify the snoRNAs, we perfromed eCLIP with all snoRNA binding proteins as baits.

overall_design: eCLIP of snoRNA binding proteins (FBL, NOP56, NOP58, SNU13, DKC1, GAR1, NHP2, NOP10)

last: Song


title: Mapping snoRNA-target RNA interactions in an RNA binding protein-dependent manner with chimeric eCLIP [RNA-seq]

summary: To quantify the effect of SNORD89 on splicing in human cells, we compared polyA mRNA-seq results in SNORD89 KD cells and control cells.

overall_design: We knock down SNORD89 with antisense oligonucleotide and perfrom polyA mRNA-seq data in none treated cells and SNORD89 KD cells. Alternative splicing was then quantified under two conditions.


title: Alveolar macrophage-specific depletion system reveals the unique roles in respiratory infections

city: Suita

first: Daisuke

last: Motooka

state: Osaka

address: 3-1, Yamadaoka

country: Japan

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

summary: Nakayama et al. generate a mouse line where alveolar macrophages are specifically labelled by fluorescent protein and diphtheria toxin receptor. Using the system, they clarify the role of alveolar macrophages in various respiratory infections and GM-CSF inhalation therapy.

overall_design: RNA sequencing showing higher expression in YFP+ Alveolar macrophages (SiglecF+ CD11c+ CD11b- Ly6G- CD45+) compared to YFP+ others in the lungs of Cx3cr1-Cre/VeDTR (ΔFRT).

first: Yuki

last: Nakayama

first: Miwa

last: Sasai

last: Okuzaki

first: Masahiro

last: Yamamoto


title: Mapping snoRNA-target RNA interactions in an RNA binding protein-dependent manner with chimeric eCLIP [Ribo_Meth-seq]

type: Methylation profiling by high throughput sequencing

summary: To determine potential snoRNA-guided methylation in rRNA and snRNA in mESC, we performed RiboMeth-seq to quantify 2'-O-methylation levels.

overall_design: 2'-O-methylation footprint in mouse ESCs was quantified by RiboMeth-seq. To identify modification sites guided by Snord89 and Snord101, we knocked down these snoRNAs with antisense oligonucleotides (ASOs) in differentiated 'induced' neurons and compared RiboMeth-seq data with non-targeting control ASO.


title: Single cell level investigations into the correlates of protection from attenuated Mtb vaccine candidate ΔsigH

status: Public on Jan 15 2025

city: Huntington

first: Sadia

last: Akter

state: WV

address: 1700 Third Ave.

department: Biomedical Sciences

institute: Marshall University

summary: Bronchoalveolar lavage (BAL) airway cells were collected from macaques and subjected to sc-RNA sequencing to determine transcriptomic changes due to vaccinations

overall_design: 23 samples from three macaque groups (unvaccinated, BCG-vaccinated, and ΔsigH-vaccinated) at pre-vaccination, post-vaccination, and post-Mtb challenge time points. Each condition has 4 samples, except "unvaccinated post-Mtb challenge" has 3 samples.

first: Deepak

last: Kaushal

first: Dhiraj

last: Singh

first: Mushtaq

last: Ahmed

first: Shaabana

last: Khader


title: Identification of Antiviral Host Factors by Functional Gene Expression Analysis using HBV Infection Assay Systems

city: Yoshida-gun

first: Takuto

last: Nosaka

state: Fukui

address: 23-3 Matsuoka, Eiheiji-cho

department: Second Department of Internal Medicine

institute: University of Fukui

summary: The in vitro HBV infection assay system was established in primary human hepatocytes (PXB cells) infected with HBV derived from the plasmid containing 1.3-mer HBV genome. Comprehensive and functional studies were performed by small interfering RNA knockdown and vector transfection experiments of the related genes and analyzed using microarray to reveal the host factors related to HBV regulation. Knockdown of STAT1 increased viral products in HBV-transfected HepG2 cells, but unexpectedly decreased in HBV-infected PXB cells. In RNA microarray analysis, STAT1 knockdown induced changes in intracellular molecules related to metabolism and protein synthesis in PXB cells, but not in HepG2 cells.

overall_design: RNA microarray was performed in HBV-infected PXB cells and HepG2.D11 cells with STAT1 knockdown and HBV-infected PXB cells with IRF2 knockdown. siRNA was transfected into the PXB cells on day 0 and HBV was added on day 4 and PXB cells were collected on day 13. HepG2.D11 cells were seeded on day 0 and siRNA was transfected at medium change on day 1. HepG2.D11 cells were collected on day 5. The comparison combinations used in this analysis were 1) PXB_si-Non Target and PXB_si-STAT1, 2) HepG2_si-Non Target and HepG2_si-STAT1, and 3) PXB_si-Non Target (PXB Non E077) and PXB_si-IRF2 (PXB IRF2 E077).

first: Tatsushi

last: Naito

first: Yu

last: Akazawa

first: Kazuto

last: Takahashi

first: Hidetaka

last: Matsuda

last: Ohtani

first: Tsutomu

last: Nishizawa

first: Hiroaki

last: Okamoto

first: Yasunari

last: Nakamoto


title: DLK-dependent protein network regulates hippocampal glutamatergic neuron degeneration

city: La Jolla

first: Yishi

last: Jin

state: CA

address: 9500 Gilman Dr

institute: University of California, San Diego

summary: The conserved MAPKKK DLK plays many roles in neuronal development, axon injury, and neuronal stress responses. The outcomes of activating or inhibiting DLK signaling depend on cell-type and cellular contexts. Emerging evidence has implicated DLK signaling in several neurodegenerative diseases. However, our understanding of the DLK-dependent cellular network in the central nervous system remains limited. Here, we investigated roles of DLK in hippocampal glutamatergic neurons, using conditional knockout and overexpression mice. We find that dorsal CA1 and dentate gyrus neurons are particularly vulnerable to elevated DLK activity. We performed RiboTRAP-seq analysis and identified the DLK dependent translatome, majority of which are involved in neuronal developmental processes, neuronal stress responses, and synapse formation and function. Increasing DLK signaling is associated with disruptions of microtubules, potentially involving Stmn4. We also show that in primary hippocampal neurons DLK regulates neurite outgrowth, axon specification, and synapse formation. This study broadens our understanding of both conserved and cell-type specific effects of DLK signaling. The identification of translational targets of DLK in glutamatergic neurons has relevance to our understanding of neurodegenerative disease.

overall_design: To investigate DLK roles in hippocampal glutamatergic neurons we generated mice with conditional overexpression or conditional knockout of DLK in glutamatergic neurons using Vglut1-Cre mice. Mice were crossed to Ribotag mice and actively translated mRNA from hippocampal glutamatergic neurons from the respective genotypes was sequenced. All samples are from the P15 timepoint. Three or four biological replicates for overexpression or knockout and respective sibling controls were used.

first: Erin

last: Ritchie


title: Cell cycle checkpoint proficiency of endometrial cancer cells

city: Boston

first: Sarah

last: Hill

state: MA

address: 450 Brookline Ave., Dana Building, Room 1410C

institute: Dana-Farber Cancer Institute

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

overall_design: Refer to individual Series


title: Gene expression profile of phagocytic and non-phagocytic microglia following traumatic brain injury and nasal anti-CD3 treatment

first: Omar

last: Albastaki

address: 60 Fenwood Rd

institute: Brigham and Women's Hospital

summary: Bulk RNA-sequencing was performed to investigate the transcriptomic signature of phagocytic and non-phagocytic microglia following traumatic brain injury and nasal anti-CD3 treatment.

overall_design: TBI-induced lesion was injected with labelled apoptotic neurons on day 6 post-injury for 16 hours and on day 7 post-injury for 4 hours. We performed bulk RNA-seq on phagocytic and non-phagocytic microglia isolated at 4 and 16 hours post-injection following traumatic brain injury and nasal anti-CD3 treatment. Preparation for RNA libraries for sequencing was performed using standard Illumina protocols.


title: IL-10-dependent Treg-microglia crosstalk induced by nasal anti-CD3 mAb ameliorates traumatic brain injury by modulating microglial phagocytosis and neuroinflammation.


title: Gene expression profile of microglia following traumatic brain injury and adoptive transfer of Tregs at acute and chronic timepoints

summary: To investigate whether the immunomodulatory effects of nasal anti-CD3 on the microglial transcriptomic profile following TBI is dependent on Tregs, we employed adoptive transfer experiments

overall_design: For the first adoptive transfer experiment, splenic/cervical lymph nodes CD4+ FoxP3+ Tregs from isotype and anti-CD3 treated donor mice (CD45.2) and CD4+ FoxP3(GFP) negative population from anti-CD3 treated donor mice (CD45.2) at 7 days post-TBI were intraperitoneally transferred into injured but untreated recipient mice (CD45.1). Bulk RNA-sequencing analysis was performed on microglia isolated from the ipsilateral hemisphere of injured but untreated mice at 7 days post-TBI following adoptive transfer of CD4+Foxp3+ Tregs from TBI-Iso mice, CD4+Foxp3+ Tregs from TBI-aCD3 mice, and CD4+ T cells depleted of Tregs from TBI-aCD3 mice. For the second experiment to characterize the gene expression profile of microglia at the chronic phase of injury following adoptive transfer, total splenic CD4+ T cells from isotype and aCD3-treated donor mice (CD45.2) and CD4+ FoxP3 GFP negative population from aCD3-treated donor mice (CD45.2) at 7 days post-TBI were intraperitoneally transferred into untreated but TBI-injured recipient mice (CD45.1). Bulk RNA-sequencing analysis was performed on microglia isolated from the ipsilateral hemisphere of injured but untreated mice at 1 month post-TBI following adoptive transfer of total CD4+ T cells from TBI-Iso mice, total CD4+ T cells from TBI-aCD3 mice, and CD4+ T cells depleted of FoxP3+ cells from TBI-aCD3 mice.


title: Effect of blocking the IL-10 receptor on the microglial transcriptomic signature following chronic traumatic brain injury and nasal anti-CD3 treatment

summary: Investigating the impact of blocking the IL-10 receptor on the gene expression profile of microglia following chronic traumatic brain injury and nasal anti-CD3 treatment.

overall_design: The IL-10 receptor was blocked in vivo through intraperitoneal administration of anti-IL-10 receptor blocking antibody every 3 days post-injury. Bulk RNA-sequencing analysis was performed on sorted microglia single cell suspensions from the ipsilateral hemisphere of the mouse brains using the microglia-specific 4D4+ antibody at 30 days following traumatic brain injury. Preparation for RNA libraries for sequencing was performed using standard Illumina protocols


title: Gene expression profile of microglia following traumatic brain injury and nasal anti-CD3 treatment at acute and chronic timepoints

summary: Bulk RNA-sequencing was performed to characterize the gene expression profile of microglia at acute and chronic timepoints following traumatic brain injury and nasal anti-CD3 treatment. We further investigated how the chronic microglial transcriptomic profile is modulated following traumatic brain injury and nasal anti-CD3 treatment in female mice with severe TBI, and in male mice with a delayed administration of treatment post-injury.

overall_design: Bulk RNA-sequencing analysis was performed on sorted microglia single cell suspensions from the ipsilateral hemisphere of the mouse brains using the microglia-specific 4D4+ antibody at 7 days and 30 days following traumatic brain injury. Preparation for RNA libraries for sequencing was performed using standard Illumina protocols. For the delayed treatment experiment, the therapeutic window of nasal anti-CD3 treatment was delayed to 3 days following injury.