title: Inhibition of hypoxic exosomal miR-423-3p decreases glioma progression by restricting autophagy in astrocytes

city: Jinan

first: Ziyi

last: Tang

state: Shandong

address: 107 Wenhuaxi Rd

institute: Shandong University

summary: The tumor microenvironment (TME) of gliomas comprises glioma cells and surrounding cells, such as astrocytes, macrophages, T cells, and neurons. In the TME, glioma cells can activate normal human astrocytes (NHAs) through the secretion of exosomes and the activation of astrocytes can further improve the progression of glioma, leading to a poor prognosis for patients. However, the molecular mechanisms underlying NHAs activation by gliomas remain largely unknown. It this study, glioma-derived exosomes (GDEs) play an important role in the modulation of autophagy and activation of NHAs. Compared with normoxic glioma-derived exosomes, hypoxic glioma-derived exosomes (H-GDEs) further improved autophagy and activation of astrocytes, which strongly promoted the progression of glioma cells. In an miRNA array between two types of exosomes from gliomas, miR-423-3p was highly expressed in H-GDEs and played an important role in autophagy, resulting in the activation of NHAs. The mechanism by which hypoxic glioma cells react with NHAs to create an immunosuppressive microenvironment was identified and 15d-PGJ2 was established as an effective inhibitor of miR-423-3p to suppress NHA activation. These findings provide new insights into the diagnosis and treatment of gliomas by targeting autophagy and miR-423-3p expression.

overall_design: normal human astrocyte were treated with normoxia and hypoxia glioma-derived exosomes, and transfected with miR-423-3p


title: Reconstruction of complex vascular networks: Bionic penile corpus cavernosum

status: Public on Jan 17 2025

city: Guangzhou

first: Zhenxing

last: Wang

state: Guangdong

address: Guangzhou, P. R. China.

department: School of Materials Science and Engineering

country: China

institute: South China University of Technology

type: Expression profiling by high throughput sequencing

summary: The purpose of conducting transcriptome research in this project is to explore the level of inflammatory response of the graft at the site of corpus cavernosum defect and the influence of the graft on vascularization and tissue regeneration.

overall_design: This experiment is divided into five groups, namely implantation A group , defect group, normal group, implantation B group, and ECs+ implantation group （n=3）. Among them, implantation A group , defect group, normal group are compared and analyzed pairwise, while implantation B group, and ECs+ implantation group are compared and analyzed.


title: DEmiRNA-mRNA regulatorynetwork reveals miR-122-5p as aregulatory factor of argininemetabolism in necrotizingenterocolitis

city: Shanghai

first: Ting

last: Guo

state: Shanghai

address: Shanghai Putuo District,Luding road No.355

department: Department of General surgery

institute: Shanghai Children's Hospital

type: Non-coding RNA profiling by high throughput sequencing

summary: To further explore the mechanism of miR-122-5p in mice with NEC.

overall_design: The miRNA sequencing was obtained from intestinal tissues (control group, NEC group).

first: Zhili

last: Ding

first: Zhibao

last: Lv

first: Li

last: Lu


title: Transcriptomic Profile of Neuroinflammation in Alzheimer’s Mouse Brain Treated With and Without Extracellular Vesicles

city: Irvine

first: Robert

middle: Paul

last: Krattli Jr

state: CA

address: 811 Health Sciences Road, MedSci B-120

department: Anatomy and Neurobiology

country: USA

institute: University of California, Irvine

summary: Expression of 757 genes related to neuroinflammation was analyzed across 4 treatment groups of male mice to investigate the neuroprotective effects of either human neural stem cell (hNSC) or induced pluripotent stem cell microglial-like (iMGL) cell-derived extracellular vesicles (EVs) in 5xFAD mouse model of Alzheimer’s Disease (AD).

overall_design: Samples from 18 male mice, ages 25-26 weeks at the time of perfusion, were analyzed using the commercially available nCounter® Mouse Neuroinflammation Panel from NanoString™, with 4-5 samples for each of the 4 treatment groups. The 5xFAD transgenic mouse line was selected for the rapid neuropathological progression of beta-amyloid plaque deposition and subsequent cognitive dysfunction. 5xFAD mice were randomly assigned to one of three AD treatment groups that would receive 4 weekly retroorbital injections of either: hNSC-derived EVs  or iMGL-derived EVs at a dose concentration of 2.25E+07 EVs/50 ul injection; or vehicle consisting of phosphate buffered saline without calcium or magnesium (PBS -/-). C57BL/6J mice served as WT controls and also received 4 weekly retroorbital injections of vehicle.

first: Munjal

last: Acharya

last: Krattli

first: Mineh

last: Markarian


title: Tissue-resident memory CD8 T Cell Diversity is Spatiotemporally Imprinted

city: La Jolla

first: Miguel

last: Reina-Campos

state: California

address: 9420 Athena Circle

institute: La Jolla Institute for Immunology

summary: Tissue-resident memory CD8 T cells (TRM) provide protection from infection at barrier sites. In the small intestine, TRM cells are found in at least two distinct subpopulations: one with higher expression of effector molecules and another with greater memory potential. However, the origins of this diversity remain unknown. We proposed that distinct tissue niches drive TRM phenotypic heterogeneity. To test this, we leveraged spatial transcriptomics of human samples, a murine model of acute systemic viral infection, and a newly established strategy for pooled optically-encoded gene perturbations to profile the location, interaction, and transcriptome of pathogen-specific TRM differentiation at single-transcript resolution. We developed computational approaches to capture cellular locations along three anatomical axes of the small intestine and to visualize the spatiotemporal distribution of cell types and gene expression. Our study reveals that the intestinal architecture’s regionalized signaling supports two distinct TRM cell states: differentiated TRM and progenitor-like TRM cells, located in the upper versus lower villus, respectively. This diversity is mediated by distinct ligand-receptor activities, cytokine gradients, and specialized cellular contacts. Blocking TGF or Cxcl9/10-sensing by antigen-specific CD8 T cells revealed a model consistent with anatomically delineated early fate specification. Ultimately, our framework for the study of tissue immune networks has revealed that T cell location and functional state are fundamentally intertwined.

overall_design: Spatial transcriptomics (VisiumHD) of mouse small intestine at day 8 after infection with LCMV and adoptive transfer of P14 CD8 T cells

first: Reina-Campos

last: Miguel

first: Monell

last: Alex

first: Heeg

last: Maximilian

first: Goldrath

last: Ananda


title: Single-cell Transcriptome and Crosstalk Analysis Reveals Immune Alterations and Key Pathways in the Bone Marrow of Knee Osteoarthritis Patients

city: Atlanta

first: Krishnendu

last: Roy

state: Georgia

address: 345 ferst drive NW, marcus nanotechnology building, room no 3102B, 3rd floor

department: IBB

institute: Georgia tech

summary: Knee osteoarthritis (OA) is a significant medical and economic burden. To understand systemic immune effects, we performed deep exploration of bone marrow aspirate concentrates (BMAC) from knee-OA patients via single-cell RNA sequencing and proteomic analyses from a randomized clinical trial (MILES: NCT03818737). We found significant cellular and immune alterations in the bone marrow, specifically in MSCs, T cells and NK cells, along with changes in intra-tissue cellular crosstalk during OA progression. Unlike previous studies focusing on injury sites or peripheral blood, our probe into the bone marrow—an inflammation and immune regulation hub—highlights remote organ impact of OA, identifying cell types and pathways for potential therapeutic targeting. Our findings highlight increased cellular senescence and inflammatory pathways, revealing key upstream genes, transcription factors, and ligands. Additionally, we identified significant enrichment in key biological pathways like PI3-AKT-mTOR signaling and IFN responses, showing their potentially crucial role in OA onset and progression.

overall_design: scRNAseq data genration and analysis to find knee-OA related changes in patient bone marrow

first: Paramita

last: Chatterjee

first: Hazel

last: Stevens

first: Linda

last: Kippner

first: Greg

last: Gibson

first: Krish


title: Tissue-resident memory CD8 T Cell Diversity is Spatiotemporally Imprinted [snRNA-Seq]


title: Skeletal muscle-derived myostatin is a major endocrine driver of follicle-stimulating hormone synthesis

city: Los Angeles

first: Michel

middle: A

last: Zamojski

address: 8700 Beverly Boulevard

department: Regenerative Medicine Institute

institute: Cedars-Sinai Medical Center

summary: Myostatin is a paracrine myokine that regulates muscle mass in a variety of species, including humans. Here, we report a functional role for myostatin as an endocrine hormone directly promoting pituitary follicle-stimulating hormone (FSH) synthesis and thereby ovarian function. Previously, this FSH-stimulating role was attributed to other members of the transforming growth factor β family, the activins. The results both challenge activin’s eponymous role in FSH synthesis and establish an endocrine axis between skeletal muscle and the pituitary gland. The data also suggest that efforts to antagonize myostatin to treat muscle wasting disorders or lean mass loss in individuals treated with GLP-1 receptor agonists may have unintended consequences on fertility.

overall_design: Pituitary glands were dissected from 8-12 weeks old female or male mice. Samples were flash frozen and  processed for snRNA-seq.

first: Xiang

last: Zhou

first: Luisina

last: Ongaro

first: Hailey

last: Schultz

first: Frederique

last: Ruf-Zamojski

first: Stuart

middle: C

last: Sealfon

first: Daniel

middle: J

last: Bernard


title: A single-cell atlas of human fetal lung development between 14 and 19 weeks of gestation

city: Ottawa

first: Laurent

last: Renesme

state: Ontario

address: 501 Smyth Road

country: Canada

institute: OHRI

summary: Rationale. Human lung development has been mainly described in morphologic studies and the potential underlying molecular mechanisms were extrapolated from animal models. Therefore, there is a need to gather knowledge from native human lung tissue. In this study we describe changes at a single-cell level in human fetal lungs during the pseudoglandular stage. Methods. We report the cellular composition, cell trajectories and cell-to-cell communication in developing human lungs with single-nuclei RNA sequencing (snRNA-seq) on 23,251 nuclei isolated from nine human fetuses with gestational ages between 14 to 19 weeks of gestation.  Results. We identified nine main lung cell populations and 21 sub-populations, including a rare pulmonary neuroendocrine cells population. For each cell type, marker genes are reported, and selected marker genes are used for spatial validation with fluorescent RNA in situ hybridization. Enrichment and changes in cell state analysis across the different gestational ages provide insight into molecular mechanisms and signaling pathways within individual cell clusters according to gestational age. Lastly, ligand-receptor analysis highlights determinants of cell-to-cell communication among the different cell types through the pseudoglandular stage, including general developmental pathways (NOTCH and TGFB), as well as more specific pathways involved in vasculogenesis, neurogenesis, and immune system regulation.  Conclusion. These findings provide a clinically relevant background for research hypotheses generation in projects studying normal or impaired lung development and help to develop and validate surrogate models to study human lung development, such as human lung organoids.

overall_design: Expression profiling by high throughput sequencing on human fetla lung tissue from 14 to 19 weeks of gestation

first: Flore

last: Lesage

first: David

middle: P

last: Cook

first: Adithya

last: Achuthan

first: Shumei

last: Zhong

first: Satu

middle: M

last: Hanninen

first: Olli

last: Carpen

first: Ivana

last: Mizik

first: Bernard

last: Thebaud


title: Modeling pre-implantation development using totipotent blastomere-like stem cells [Cut&tag]

city: Beijing

first: Bing

last: Peng

state: Beijing

address: Yiheyuan Road

department: School of Life Sciences

institute: Peking University

summary:
we have established TBLC-based differentiation systems to recapitulate the entire pre-implantation development
Cleavage under target & tagmentation (CUT&Tag)-sequencing (CUT&Tag) for H3K4me3,H3K27ac,H3K9me3,RNA polymerase II(Pol2) and h3k27me3 in mPSC sand mTBLCs

overall_design: Comparative epigenetic profiling analysis of H3K4me3,H3K27ac,H3K9me3,RNA polymerase II(Pol2) and h3k27me3 data for mPSCs and mTBLCs

first: Qingyi

last: Wang

first: Feixiang

last: Zhang


title: Modeling pre-implantation development using totipotent blastomere-like stem cells [ATAC-seq]

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
we have established TBLC-based differentiation systems to recapitulate the entire pre-implantation development
Assay for transposase accessible chromatin with high-throughput sequencing in mESC sand mTBLCs

overall_design: Comparative chromatin accessibility analysis of more open and closed signals in ATAC-seq for mPSCs and mTBLCs


title: Modeling pre-implantation development using totipotent blastomere-like stem cells [scRNA-seq]

summary: we have established TBLC-based differentiation systems to recapitulate the entire pre-implantation development

overall_design: Poly(A) RNA-seq is performed on mTBLCs cultured in 2MYCP medium


title: TaqMan Array Human Cytokine Network 96-well Plate for M0 human primary macrophages

city: Pecs

first: Hairui

last: Ou

address: Rokus

country: Hungary

institute: University of Pecs

summary: Peripheral blood was acquired from the Pécs Regional Blood Transfusion Center (Pécs, Hungary) and processed using Lymphocyte Separation Medium 1077 (Promocell, Heidelberg, Germany) in accordance with the manufacturer's instructions. Subsequently, the viable mononuclear cells were isolated, counted, and seeded into 6-well plates at a density of 1,000,000 cells per well.   The culture medium was specifically formulated using RPMI-1640 medium (Biowest, Nuaillé, France), enriched with 1% L-glutamine (Lonza, Basel, Switzerland), 2% penicillin/streptomycin (Lonza, Basel, Switzerland), and 10% exosome-depleted FBS (Thermo Fisher Scientific, Waltham, USA) to minimize EV interference from bovine serum, further supplemented with 100 ng/mL Peprotech M-CSF 300-25 (Gibco, Waltham, USA). The cells were cultured for 5 days in a 5% CO2 incubator at 37 °C, resulting in the differentiation of all monocytes into macrophages (M0) (Cao et al., 2019).

overall_design: Differentiated macrophages were treated with MEV (Milk-derived extracellular vesicles) from healthy and mastitis groups separately to achieve the final concentration of 2×10^11 particles/mL. Additionally, untreated cell cultures with the same volume of PBS were maintained as negative controls. After 48 hours, macrophage supernatants were collected and stored at -80 °C (for a maximum of 3 months) for future experiments.

first: Ou

last: Hairui

first: Kvell

last: Krisztian


title: Targeting cIAP2 in a novel senolytic strategy prevents glioblastoma recurrence after radiotherapy

city: San Antonio

first: Luis Fernnado

last: Macedo Di Cristofaro

state: Texas

address: 7703 Floyd Curl Drive,

department: Neurosurgery

institute: UTHSCSA

summary: Glioblastomas (GBM) are routinely treated with high doses of ionizing radiation (IR), yet these tumors recur quickly, and the recurrent tumors are highly therapy resistant. Here, we report that IR-induced senescence of tumor cells counterintuitively spurs GBM recurrence, driven by the senescence-associated secretory phenotype (SASP). We find that irradiated GBM cell lines and patient derived xenograft (PDX) cultures senesce rapidly in a p21-dependent manner. Senescent glioma cells upregulate SASP genes and secrete a panoply of SASP factors, prominently interleukin IL-6, an activator of the JAK-STAT3 pathway. These SASP factors collectively activate the JAK-STAT3 and NF-kB pathways in non-senescent GBM cells, thereby promoting tumor cell proliferation and SASP spreading. Transcriptomic analyses of irradiated GBM cells and the TCGA database reveal that the cellular inhibitor of apoptosis protein 2 (cIAP2), encoded by the BIRC3 gene, is a critical survival factor for senescent glioma cells. Senescent GBM cells not only upregulate BIRC3 but also induce BIRC3 expression and promote radioresistance in non-senescent tumor cells. We find that SMAC mimetics targeting cIAP2 act as novel senolytics that trigger apoptosis of senescent GBM cells with minimal toxicity towards normal brain cells. Finally, using both PDX and syngeneic mouse models of GBM, we show that the SMAC mimetic birinapant, administered as an adjuvant therapy after radiotherapy, can eliminate senescent GBM cells and prevent the emergence of recurrent tumors. Taken together, our results clearly indicate that significant improvement in GBM patient survival may become possible in the clinic by eliminating senescent cells arising after radiotherapy

overall_design: Total RNA was extracted from GBM cells LN229 and A172 mock-irradiated and radiation-induced senescent cells.. Three biological replicates of mock-irradiated and IR-induced senescent cells were analyzed by bulk-RNAseq .


title: Activation of Notch2 signaling drives cyst growth in kidney

city: Tianjin

first: Huiwen

last: Ren

address: 22 Qixiangtai Road

institute: Tianjin Medicial University

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

overall_design: Refer to individual Series


title: DNA methylation in mouse cancer model with ketogenic diet and mifepristone treatment

city: Innsbruck

first: Martin

last: Widschwendter

address: Innrain 52

country: Austria

institute: University of Innsbruck

type: Methylation profiling by genome tiling array

summary: DNA methyaltion analysis using Illumina Infinium Mouse Methylation Beadchips in different groups of mice with and without exposure to medroxyprogesterone acetate and 7,12-dimethylbenzanthracene MPA/DMBA (which induces mammary gland tumours), with and without mifepristone (an antiprogestin) treatment, and with and without a ketogenic diet.

overall_design: A total of 86 mice are organised into three experimental sets. The discovery set consists of 9 mice without exposure to MPA/DMBA and 9 mice with MPA/DMBA exposure (of which 8 developed a mammary gland tumour). These mice were on a normal diet and were not treated with mifepristone. Tissue samples from normal mammary gland, mammary tumor, cervix, oviduct, blood, adipose tissue, kidney, liver, lung, and spleen were collected from both groups. The intervention set conists of of 35 mice fed with normal diet which were randomized into four groups: without MPA/DMBA exposure and without mifepristone, without MPA/DMBA exposure and with mifepristone, with MPA/DMBA exposure and without mifepristone, and with MPA/DMBA exposure and with mifepristone. Tissue samples from normal mammary gland, mammary tumours, cervix, oviduct and blood were taken from all subgroups. The validation set conists of of 33 mice fed with ketogenic diet which were randomized into four groups: without MPA/DMBA exposure and without mifepristone, without MPA/DMBA exposure and with mifepristone, with MPA/DMBA exposure and without mifepristone, and with MPA/DMBA exposure and with mifepristone. Tissue samples from normal mammary gland, mammary tumours, cervix, oviduct and blood were taken from all subgroups.

first: James

last: Barrett


title: Activation of Notch2 signaling drives cyst growth in kidney [RNA-seq: R26NICD2 KspCre]

summary: In order to investigate how NICD2 promotes cyst growth, we carried out bulk RNA-Seq analysis of kidneys from PKD mouse models R26NICD2/+KspCre.

overall_design: mRNA profiles of 3-day old wild type and R26NICD2/+Kspcre

first: Ying

last: Yu


title: Activation of Notch2 signaling drives cyst growth in kidney [RNA-seq: R26NICD2 Pax-rtTA]

summary: In order to investigate how NICD2 promotes cyst growth, we carried out bulk RNA-Seq analysis of kidneys from PKD mouse model R26NICD2/+ Pax8-rtTA mice.

overall_design: mRNA profiles of 8-week old wild type and R26NICD2/+pax8cre


title: Modeling pre-implantation development using totipotent blastomere-like stem cells [bulk RNA-seq]

summary:
we have established TBLC-based differentiation systems to recapitulate the entire pre-implantation development
Poly(A) RNA-seq is performed on mTBLCs cultured in 2MYCP medium

overall_design: Comparative gene expression profiling analysis of RNA-seq data for mPSCs and mTBLCs


title: Adipose-derived Leptin and Complement Factor D mediate osteoarthritis severity and pain

status: Public on Jan 16 2025

city: St Louis

first: Tiandao

last: Li

state: MO

address: 4444 Forest Park Ave

institute: Washington University

summary: Obesity is a risk factor for osteoarthritis (OA), and leptin is among the adipokines implicated in obesity-induced OA. However, the specific role of leptin in OA severity and pain is not known. Using lipodystrophic (LD) mice, we show fat-secreted factors are required for knee OA development, implicating a fat-cartilage crosstalk. Fat pad implantation or systemic leptin restoration in LD mice reintroduced structural OA and pain, whereas implantation of leptin-deficient fat pad did not change OA susceptibility. Isochronic parabiosis and spatial transcriptomics confirmed that fat-joint crosstalk likely occurred via soluble mediators. Global unsupervised multiomics of conditioned media from fat implants revealed that leptin exerts a regulatory effect on adipsin (or complement factor D), the activity of which modulates contrastive OA structural and pain phenotype. These findings suggest adipokines influence OA pathogenesis, providing conclusive evidence of a fat-joint crosstalk and implicating OA as a systemic disease of adipose tissue.

overall_design: Male LD mice received either a MEF transplant (MEF-rescue) from WT, leptin heterozygous (MEF-HET) or leptin homozygous knockout (MEF-KO) pups as previously described, which developed into adipose-like tissue between 3 and 5 weeks of age. The injection was delivered subcutaneously to the sternal aspect of a donor LD mouse (3–5 weeks old) under 2% isoflurane with a 27 gauge needle.  At 16 weeks of age, surgical animals underwent destabilization of the medial meniscus surgery in their left knee joints, and the right limb served as a nonsurgical contralateral control. Mice were sacrificed at 28 weeks of age to assess OA severity, bone microstructure, and serum and SF inflammatory profiles. Genotype: Vis (Visceral), KO (Leptin Knockout), Sub (Subcutaneous), and WT (Wildtype). Treatment: EVFN (endogenous visceral fat nonsurgical), MKM (MEF knockout fat males + surgery), ESM (endogenous subcutaneous fat + male surgery), MKF (MEF knockout fat + surgery female), EVM (endogenous visceral fat + surgery), MWF (MEF WT fat females + surgery), MWM (MEF WT fat males + surgery), and ESF (endogenous subcutaneous fat + female surgery)

first: Collins

last: Kelsey

first: Zhang

last: Bo

first: Guilak

last: Farshid


title: Spray-Dried Porcine Plasma Promotes the Association Between Metabolic and Immunological Processes at Transcriptional Level in Gilthead Sea Bream (Sparus aurata) Gut.

city: Santiago

first: Felipe

middle: E.

last: Reyes-López

state: Santiago

address: Av. Libertador Bernado O'Higgins 3363

country: Chile

institute: Universidad de Santiago de Chile

summary: This study aimed to unveil the biological mechanisms modulated in response to the dietary administration of SDPP in the gilthead sea bream gut.

overall_design: Fish (BWi = 10.6   0.1 g, n = 400, mean   standard deviation, SD) were fed during 95 days with C and SDPP diets using automatic feeders (ARVO-TEC T Drum 2000; Arvotec, Huutokosk, Finland) at a rate of 2.5% of the stocked biomass, which approached apparent satiation. The daily feed ratio was evenly distributed in 7 meals per day from 08:00 to 18:00 h. During the trial, water temperature and pH (pH meter 507; Crison Instruments, Barcelona, Spain), salinity (MASTER-20T; ATAGO Co., Ltd., Tokyo, Japan), and dissolved oxygen (OXI330; Crison Instruments) was measured. The water flow rate was maintained at approximately 9.0–10.1 L/min in a recirculation system (IRTAmar; Spain) that provided the experimental tanks with adequate water quality (total ammonia and nitrite) through UV, biological, and mechanical filtration. Photoperiod followed natural changes according to the season of the year (November to February; 40°37'41'' N). Fish were regularly sampled monthly to evaluate their growth in BW and adjust the feeding ratio to stocked biomass.  At the end of the trial, fish were anesthetized as described above. All fish in experimental tanks were measured for final BW and SL as indicated. Then, 10 fish per tank were sacrificed with an overdose of MS-222 (200 mg/L) for tissue sampling purposes. The intestinal cavity was exposed, and the intestine was removed with a scalpel. A small piece of the anterior intestine (0.5 cm) was dissected and immediately transferred into RNAlater (Ambion) incubated at 4 C overnight and frozen at -80°C until further RNA extraction.

first: Eva

last: Vallejos-Vidal

middle: E

first: Enric

last: Gisbert


title: Pervasive formation of double-stranded RNAs by overlapping sense/antisense transcripts in budding yeast mitosis and meiosis

city: Paris

first: Maxime

last: Wery

address: 26 rue d'Ulm

country: France

institute: Institut Curie

summary:
Previous RNA profiling studies revealed co-expression of overlapping sense/antisense (s/a) transcripts in pro- and eukaryotic organisms. Functional analyses in yeast have shown that certain s/a mRNA/mRNA and mRNA/lncRNA pairs form stable double-stranded RNAs (dsRNAs) that affect transcript stability. Little is known, however, about the genome-wide prevalence of dsRNA formation and its potential functional implications during growth and development in diploid budding yeast. To address this question, we monitored dsRNAs in a Saccharomyces cerevisiae strain expressing the ribonuclease DCR1 and the RNA binding protein AGO1 from Naumovozyma castellii. We identify dsRNAs at 347 s/a loci that express partially or completely overlapping transcripts during mitosis, meiosis or both stages of the diploid life cycle. We associate dsRNAs with s/a loci previously thought to be exclusively regulated by antisense interference, and others that encode antisense RNAs, which downregulate sense mRNA-encoded protein levels. To facilitate hypothesis building we developed the Sense/Antisense double-stranded RNA (SensR) expression viewer. Users are able to retrieve different graphical displays of dsRNA and RNA expression data using genome coordinates and systematic or standard names for mRNAs and different types of stable or cryptic long non-coding RNAs (lncRNAs). Our data are a useful resource for improving yeast genome annotation and for work on RNA-based regulatory mechanisms controlling transcript and protein levels. The data are also interesting from an evolutionary perspective, since natural antisense transcripts that form stable dsRNAs have been detected in many species from bacteria to humans.
The SensR viewer is freely accessible at https://sensr.genouest.org.

overall_design: Small RNA sequencing in cells of S. cerevisiae with or without reconstituted RNAi pathway.

first: Ugo

last: Szachnowski

first: Emmanuelle

last: Becker

first: Igor

last: Stuparevic

first: Olivier

last: Sallou

first: Mateo

last: Boudet

first: Anthony

last: Bretaudeau

first: Antonin

last: Morillon

first: Michael

last: Primig


title: Modeling pre-implantation development using totipotent blastomere-like stem cells

first: Bing

last: Peng


title: Cancer-associated fibroblasts serve as decoys to suppress NK cell anti-cancer cytotoxicity [NMFs]

city: Rehovot

first: Aviad

last: Ben-Shmuel

state: Other

address: Nella and Leon Benoziyo Building for Biological Sciences, Herzl

department: Biomolecular Sciences

country: Israel

institute: Wiezmann Institute

summary: Cancer associated fibroblasts (CAFs) are one of the most abundant components of the breast tumor microenvironment (TME) and major contributors to immune modulation in the TME. CAFs are well known to regulate the activity of diverse types of immune cells including T cells, macrophages and dendritic cells, however little is known about their interaction with Natural killer (NK) cells. NK cells constitute an important arm of anti-tumor immunity, yet the regulation of NK cell activity by CAFs in solid tumors is poorly understood. Here we find, using mouse models of cancer and ex-vivo cocultures, that immunosuppressive CAF subsets severely inhibit NK cell cytotoxicity towards cancer cells. We unravel the mechanism by which this suppression occurs, through CAF-mediated downregulation of the NK-surface receptors, Natural Killer Group 2D (NKG2D) and DNAX Accessory Molecule-1 (DNAM-1). Ligands for these receptors are known to be expressed by cancer cells and minimally expressed in healthy tissue. Here we find that CAFs also upregulate ligands for NKG2D and DNAM-1. Ligand-receptor engagement between NK cells and CAFs leads to CAF cytolysis, which in turn diminishes the expression of NKG2D and DNAM-1 on NK cells via a negative feedback loop, and promotes cancer escape from NK cell surveillance. These results reveal a CAF-mediated immunosuppressive mechanism with implications for treatment of solid tumors.

overall_design: To investigate the baseline transcriptomic differences between normal mammary fibroblasts (NMFs) from C57BL6 and BALB/C mice, we isolated fibroblasts from the mammary fat pad of healthy female mice using magetic seperation (3 biological replicates each) and subsequently cultured NMFs for four days prior to bulk RNA sequencing

first: Avad

first: Yael

last: Gruper

first: Ruth

last: Scherz-Shouval


title: Profiling of RNAs associated with LINE-1 ORF1p in pancreatic ductal adenocarcinoma

city: Charlestown

first: Eunae

last: You

state: MA

address: 149, 13th street

department: Center for Cancer Research

institute: Massachusetts General Hospital

summary: Repeat element viral mimicry is a common feature in pancreatic ductal adenocarcinoma (PDAC) that require mechanisms to manage this repeat “viral” load and attenuate innate immune responses. Here, we show that the LINE-1 ORF1 protein (ORF1p) in PDAC cells plays a role in shielding repeat RNAs from activating a pathogen recognition receptor (PRR)-mediated antiviral response that is independent of retrotransposition. Suppression of ORF1p using short hairpin RNA induces innate immune responses through the dsRNA sensors RIG-I and MAVS. Low ORF1p PDAC cell lines have suppressed expression of PRRs demonstrating convergent mechanisms to suppress innate immune signaling. Localization of ORF1p in processing bodies (P-bodies) with the dsRNA helicase MOV10 were found important for these antiviral responses. Loss of ORF1p resulted in significant growth reduction in tumorspheres and mouse xenografts with an enriched epithelial cell state, and high ORF1p expression was associated with worsened survival in a cohort of human PDAC patients.

overall_design: RNA immunoprecipitation was conducted in three PDAC cell lines (PDAC3, 6, and 8) cultured in 3D tumorspheres for 7 days, followed by RNA isolation.

first: Bidish

middle: K

last: Patel

first: Alexandra

middle: S

last: Rojas

first: Siyu

last: Sun

first: Natalie

middle: I

last: Ho

first: Ildiko

last: Phillips

last: Raabe

first: Yuhui

last: Song

first: Katherine

middle: H

last: Xu

first: Joshua

middle: R

last: Kocher

first: Peter

last: Richiery

first: Phoebe

last: Shin

first: Martin

last: Taylor

first: Linda

middle: T

last: Nieman

first: Benjamin

middle: D

last: Greenbaum

first: David

last: Ting


title: LINE-1 ORF1p immune evasion mechanisms in pancreatic ductal adenocarcinoma

overall_design: This dataset include control and ORF1, MOV10 shRNA-treated human pancreatic cancer cell lines (biological replicates = 3).

first: Michael


title: Histone deacetylase-1 is required for epigenomic stability in Neurospora crassa [ChIP-seq]

city: Athens

first: Zachary

last: Lewis

state: GA

address: 120 E. Green St.

department: Microbiology

institute: University of Georiga

summary: Polycomb group (PcG) proteins assemble into chromatin modifying complexes that regulate stem cell identity and multicellular development. The trimethylation of lysine 27 on histone H3 (H3K27me3) is catalyzed by the conserved Polycomb Repressive Complex 2 (PRC2). In Neurospora crassa, the typically-subtelomeric pattern of H3K27me3 depends on other chromatin regulators including the H3K36 methyltransferase ASH1, the chromatin remodeling enzyme IMITATION SWITCH, and components of constitutive heterochromatin including the H3K9 methyltransferase DEFECTIVE IN METHYLATION-5 (DIM-5) and the H3K9me3-binding protein HETEROCHROMATIN PROTEIN 1 (HP1). How constitutive heterochromatin impacts PRC2 activity is unclear. We performed a genetic screen to identify histone deacetylase genes required to repress PRC2-methylated genes in N. crassa. We found that HISTONE DEACETYLASE-1 (HDA-1) is required for transcriptional repression in PcG-repressed domains and for normal patterns of H3K27me3. In the absence of HDA-1, H3K27me3 is lost from typical PRC2-methylated domains and instead is enriched at a subset of constitutive heterochromatin domains normally marked for H3K9me3. In addition to altered H3K27me3 patterns, HDA-1-deficient mutants displayed aberrant patterns of H3K9me3, H3K36me3, and lysine acetylation (Kac). Mutants lacking the HP1/HDA-1-interacting protein CHROMO DOMAIN PROTEIN-2 (CDP-2) displayed a similar but non-identical phenotype, and we observed variable patterns of aberrant H3K7me3 enrichment in genetically identical ∆hda-1 strains, raising the possibility that loss of HDA-1 activity leads to progressive changes in H3K27me3 enrichment. Indeed, a newly constructed ∆hda-1 deletion strain displayed a near-wild type H3K27me3 chromatin modification profile that decayed over multiple strain passages corresponding to hundreds of nuclear divisions. Together, our data indicate that HDA-1 is a critical regulator of epigenome stability in N. crassa.

overall_design: Zachary Lewis

first: Aileen R.

last: Ferraro

first: Farh

last: Kaddar

first: Clayton

last: Hull-Crew

first: Ashley W.

last: Scadden

first: Andrew D.

last: Klocko

first: Zachary A.

first: Felicia

last: Ebot-Ojong


title: Effect of GPX4 knockdown on gene expression profile of EBV positive gastric cancer cell line AGS-EBV.

city: Qingdao

first: Duo

last: Shi

address: Qingdao University, 308 Ningxia Road, Qingdao

department: School of Basic Medicine

institute: Qingdao University

summary: Reactive oxygen species (ROS) mediated oxidative stress plays an increasingly extensive role in tumorigenesis. Oxidative stress is also an important mechanism for host cells to resist infection by pathogenic microorganisms. Viruses have a similar defense mechanism. Studies have shown that ROS levels are elevated during in vitro and in vivo EBV infection. In nasopharyngeal carcinoma, EBV latent membrane protein LMP1 "REDOX" nasopharyngeal carcinoma cells by up-regulating NRF2 and NOX2, acquiring new REDOX homeostasis to tolerate higher levels of ROS. In gastric cancer, EBV upregulates NOX2 by down-regulating miR34a via EBNA1, thereby enhancing the antioxidant capacity of host cells. In addition, oxidative stress is involved in the lytic reactivation of EBV, but the specific mechanism is not clear. GPX4 is an important ROS scavenger that plays a role in maintaining cellular REDOX homeostasis. Our study showed that GPX4 is relatively highly expressed in EBV-positive gastric cancer cell lines, which can promote tumor proliferation and migration, and we found that GPX4 helps maintain latent infection of EBV in gastric cancer. In order to further explore the specific mechanism, we performed transcriptome sequencing analysis after knocking down GPX4 expression in AGS-EBV cells.

overall_design: AGS-EBV cells were transfected with lentivirus, a cell line with stable knockdown of endogenous GPX4 (AGS-EBV-shGPX4) and a control cell line (AGS-EBV-shNC) were constructed, and transcriptome sequencing was performed.


title: Histone deacetylase-1 is required for epigenomic stability in Neurospora crassa [CUTnRUN]


title: Total B cells from Healthy Individual

city: Manchester

first: Ian

middle: Edward

last: Prise

state: Select state

address: Oxford Road

country: United Kingdom

institute: University of Manchester

summary: B cells are a heterogenous population circulating in peripheral blood. We used single cell RNA sequencing (scRNA-seq) to analyze the diversity of B cell populations in human peripheral blood.

overall_design: Live CD19+CD3- B cells were isolated by flow-cytometric cell sorting (FACS) of peripheral blood mononuclear cells (PBMC) from a healthy individual.

first: Grainger

last: John Robert

first: Younas

last: Mehwish

first: Prise

last: Ian


title: RNA sequencing(RNA-Seq)in liver tissues from different exercise interventions in NAFLD mice

city: Wuhan

first: Mingliang Wei

last: Mingliang Wei

state: Hubei

address: 299 Bayi Road, Wuchang District

institute: Wuhan University

overall_design: C57BL/6 male mice (GemPharmatech Co., Ltd) aged 6 weeks were randomly divided into normal diet control (NC) group (n=13) and HFD group (D12492, Research Diets, n=29) after one week of adaptive feeding. After eight weeks, HFD group mice were randomly divided into the following three groups (n=8): HFD, HAE (HFD + AE group), and HRE (HFD + RE group) and received at least 12 weeks of exercise training.

first: Chaorui

last: Hu

first: Chuanfei

last: Zeng

first: Huan

last: Li

first: Mingliang

last: Wei

first: Shiqian

last: Liu

first: Lianyun

first: Yong

last: Xiao

first: Du

last: Cheng

first: Min

last: Wu

first: Mingkai

last: Chen


title: Persistent large-scale changes in alternative splicing in prefrontal cortical neuron types following psychedelic exposure

city: Berkeley

first: Andrea

last: Gomez

address: 163 Weill Hall

department: Molecular and Cell Biology

institute: University of California, Berkeley

summary: The goal of this project was to assess the cell-type specific transcriptomic changes induced by psychedelics in a rodent model. We used the RiboTag technique to isolate RNA from genetically-defined neuronal subtypes in the mouse prefrontal cortex. Conditional tagging of the ribosomal protein Rpl22 was done with Pvalb-Cre, Cux2-CreERT, and Rbp4-Cre for PV interneurons, L2/3 pyramidal neurons, and L5 pyramidal neurons respectively. We deep-sequenced (~100 million paired-end reads) the RNAs using Illumina platforms. We found cell-type specific gene expression and alternative splicing changes associated with psychedelics, providing a essential resource for understanding the various effects of psychedelics on cortical cell types and functions.

overall_design: We mapped ribosome-bound RNA after treatment with saline or psychedelics DOI or psilocybin in genetically-defined neurons of the mouse prefrontal cortex, with 4-5 replicates per cell type, per time point, and per treatment.

first: Yungyi

last: Hsiao

first: Marcos

last: Fonseca


title: Gene expression differences between wild-type and ZUP1 knock-out mice in liver infected with MHV-A59

city: BEIJING

first: HAO

last: wenyan

department: School of Basic Medical Sciences

institute: Peking University Health Science Center

summary: To investigate the transcriptome differences between wild-type and ZUP1 knock-out mice in liver when they were infected with MHV, we employed MHV-A59, a hepatic and neuronal tropic murine coronavirus, to infect wild-type and Zup1 knockout mice and colleted liver tissues after MHV infection to perform RNA- sequencing.

overall_design: We then performed gene expression profiling analysis using data obtained from RNA-seq of wild-type and Zup1 knockout mice infected with MHV-A59 for 48 hours.

first: Wenyan

last: Hao

first: Mengfan

last: Guo


title: IFIT1 Knockdown Mitigates Translational Suppression, Enhancing Myogenic Differentiation and Revitalizing Mesenchymal Stem Cell Therapy for Myocardial Infarction [III]

city: Shanghai

first: Lei

last: Hou

address: 746 Zhongshan Middle Road

department: Department of Cardiology

institute: Songjiang Hospital and Songjiang Research Institute, School of Medicine, Shanghai Jiao Tong University

summary: Early transplantation of mesenchymal stem cells (MSCs) showed promise in rescuing cardiac function and alleviating ventricular remodeling post-myocardial infarction (MI) while regulatory T cells (Treg) either. However, the crosstalk between them remains fully elusive. To explore mechanisms optimizing MSC therapy from Treg immunoregulatory and develop potential therapeutic targets, adipose-derived stem cells (ADSCs), an easily accessible MSC, were applied. The Treg depletion diminished the cardioprotective effects of ADSCs, causing both systolic dysfunction and cardiac fibrosis. Tregs promoted ADSC retention by proliferation and migration activation with further myogenic differentiation. RNA-sequencing analysis revealed Treg co-culture induced a myogenic differentiation mRNA profile in ADSCs, significantly downregulating Ifit1. IFIT1 downregulation resulted in cardiomyocyte-like differentiation in ADSCs, evidenced by increased myogenic markers and spontaneous beating. Mechanistically, IFIT1 knockdown provoked upregulation of E2F1 protein without altering E2f1 mRNA levels. IFIT1 specifically bound to E2f1 mRNA to inhibit translation, reducing polysomes while knockdown introduced myogenic translational profile. The rAAV9-sh-Ifit1 injection restored systolic dysfunction, reduced fibrosis, and attenuated cardiac hypertrophy. This study uncovered a novel mechanism whereas Tregs potentiated the therapeutic efficacy of ADSCs in MI by downregulating IFIT1, thereby promoting E2f1 translation and myogenic differentiation, offering a potential target to improve cardiac repair strategies.

overall_design: RNA sequencing analysis was performed on Scrasmbled ADSCs polysomes (polysome_Sc-1, polysome_Sc-2) and sh-Ifit1 ADSC polysomes (polysome_Kd-1, polysome_Kd-2)  (n = 2/group).

first: Luyao

first: Yanan

last: Pang

first: Jiachun

last: Xia

first: Sensen

last: Zhang

first: Jumo

last: Zhu


title: IFIT1 Knockdown Mitigates Translational Suppression, Enhancing Myogenic Differentiation and Revitalizing Mesenchymal Stem Cell Therapy for Myocardial Infarction [II]

overall_design: RNA sequencing analysis was performed on Scrasmbled ADSCs (TR-Sc-1,TR-Sc-2,TR-Sc-3) and sh-Ifit1 ADSC (TR-KD-1,TR-KD-2,TR-KD-3)  (n = 3/group).


title: NLRP7 maintains the genomic stability during early human embryogenesis via mediating alternative splicing

city: Guiyang

first: Liangxia

last: Jiang

address: No. 6 Ankang Avenue, Gui'an New District

institute: Guizhou Medical University

summary: Genomic instability is the main cause of abnormal embryo development and abortion. NLRP7 dysfunctions affect embryonic development and lead to recurrent pregnancy loss (RPL) or hydatidiform moles (HM), but the underlying mechanisms remain largely elusive. In this study, we found that NLRP7 knockout affected the genetic stability, resulting in increased DNA damage in both human embryonic stem cells (hESCs) and blastoids, making embryonic cells in blastoids more susceptible to apoptosis. Mechanistically, NLRP7 could interact with factors related to alternative splicing (AS) and DNA damage response (DDR), including DDX39B, PRPF8, THRAP3 and PARP1, and its dysfunction led to abnormal AS of genes involved in Homologous recombination (HR) in hESCs, indicating that NLRP7-mediated AS is required for the maintenance of genome integrity during early human embryogenesis. Together, this study first uncovers that NLRP7 plays an essential role in the maintenance of genetic stability during early human embryonic development by regulating AS of HR-related genes.

overall_design: To investigate the involvement of NLRP7 in maintaining the genetic stability  in early human embryonic cells, we established a NLRP7 knockout cell line using CRISPR/Cas9 gene-editing technology

first: Zhongliang

last: Su

first: Qibing

last: Zeng

first: Peng

last: Luo

first: Liangzhao

last: Chu


title: BMPR1A-mediated BMP signalling ensures correct mitosis-to-meiosis transition in mouse fetal ovarian germ cells, in vivo

city: Brisbane

first: Josephine

last: Bowles

state: Queensland

address: The University of Queensland

department: School of Biomedical Sciences

country: Australia

institute: The University of Queensland

summary: A successful mitosis-to-meiosis transition in germ cells is essential for fertility in sexually reproducing organisms. In mice and humans, it is established that STRA8 expression is critical for meiotic onset in both sexes. Here we show that BMP signalling is also essential, not for STRA8 induction but for correct meiotic progression in mouse fetal ovarian germ cells. Germ cell-specific deletion of Bmpr1a caused aberrant expression of meiotic, pluripotency, and cell cycle genes in E14.5 mouse fetal ovaries. Our results verify a role for BMP signalling in instructing germ cell meiosis in female mice in vivo, and shed light on the regulatory mechanisms underlying fetal germ cell development.

overall_design: Comparative transcriptome analysis of RNA-seq data from E14.5 Bmpr1a cKO vs control mouse fetal ovaries.

first: Fiona Ka Man

last: Cheung

first: Chun-Wei

last: Feng

first: Cassy

last: Spiller


title: SINEUP RNA rescues molecular phenotypes associated with CHD8 suppression in Autism Spectrum Disorder model systems

city: Trento

first: Erik

last: Dassi

state: TN

address: Via Sommarive, 9

department: CIBIO

country: Italy

institute: University of Trento

summary: Paired-end of RNA-seq of SINEUP silencing of CHD8 in hiNPCs cells

overall_design: total RNA-seq of control (shGFP) and SINEUP-silenced CHD8

first: Marta

last: Biagioli

first: Francesca

last: Di Leva

first: Miguel

last: Pellegrini


title: Transcriptomic changes induced by acute mtDNA depletion in IMR90 cells in the absence of energetic stress

city: San Diego

first: April

middle: Elizabeth

last: Williams

address: 10010 N Torrey Pines Rd

department: IGC

institute: Salk Institute for Biological Studies

summary: The functional status of mitochondria is relayed by mitochondrial retrograde signaling (MRS) to elicit adaptive changes in nuclear gene expression. However, MRS pathways that sense the amount of the multi-copy mitochondrial DNA (mtDNA) in mammalian cells remain poorly understood. Budding yeast have “intergenomic signaling” pathways that can sense the amount of mtDNA independently of oxidative phosphorylation (OXPHOS), the primary function of genes encoded by mtDNA.  We found that mtDNA-depleted IMR90 cells can sustain OXPHOS for a significant amount of time, providing a robust model system to interrogate human intergenomic signaling. FAM43A is induced early in response to mtDNA depletion in a CHK2 kinase-dependent manner, and its depletion activates CHK2 to increase mtDNA copy number via upregulation of the p53R2 form of ribonucleotide reductase. We propose that FAM43A serves a checkpoint-like function that restricts mtDNA replication in the presence of mtDNA stress to maintain mtDNA integrity. FAM43A could be a candidate mtDNA-disease locus and a therapeutic target for mtDNA depletion syndromes.

overall_design: To investigate the transcriptomic changes that occur at different stages of mtDNA depletion, we treated IMR90 cells with 2'3'-dideoxycytidine (20 uM) for 0, 24, 48, 72, or 96 hours. We then performed differential gene expression analysis using data obtained from RNA-sequencing of untreated (n=2 biological replicates) and 2'3'-dideoxycytidine treated cells (n=3 biological replicates per time point).

first: Alva

middle: G

last: Sainz

first: Gerald

last: Shadel


title: Stem Cell-Derived Vessels-on-Chip for Cardiovascular Disease Modeling

city: Neuherberg

first: Sandra

last: Wiedenmann

state: Bavaria

address: Ingolstädter Landstraße 1

department: Helmholtz Pioneer Campus

country: Germany

institute: Helmholtz Zentrum München

summary: The metabolic syndrome is accompanied by vascular complications. Human in vitro disease models are hence required to better understand vascular dysfunctions and guide clinical therapies. Here, we engineered an open microfluidic vessel-on-chip platform that integrates human pluripotent stem cell-derived endothelial cells (SC-ECs). The open microfluidic design enables seamless integration with state-of-the-art analytical technologies, including single-cell RNA sequencing, proteomics by mass spectrometry, and high-resolution imaging. Beyond previous systems, we report SC-ECs maturation by means of barrier formation, arterial toning, and high nitric oxide synthesis levels under gravity-driven flow. Functionally, we corroborate the hallmarks of early onset atherosclerosis with low sample volumes and cell numbers under flow conditions by determining proteome and secretome changes in SC-EC stimulated with oxidized low-density lipoprotein and free fatty acids. More broadly, our organ-on-chip platform enables the modeling of patient-specific human epithelial tissue and has the potential to become a general tool for animal-free vascular research.

overall_design: Single cell RNA sequencing of endothelial cells cultivated under flow or static condition on a Vessel-on-Chip System

first: Maren

last: Marder

first: Caroline

last: Remmert

first: Julius

middle: A

last: Perschel

first: Munkhtur

last: Otgonbayar

first: Christine

last: von Toerne

first: Stefanie

last: Hauck

first: Judith

last: Bushe

first: Annette

last: Feuchtinger

first: Bilal

last: Sheikh

first: Michel

last: Moussus

first: Matthias

last: Meier


title: Single-cell transcriptomic analysis of skin sorted CD45 positive and CD45 negative population from Il2ra upstream enhancer deleted (UP_Del)  mice presenting with and without alopecia phenotype.

city: Bethesda

first: Keisuke

last: Nagao

state: MD

address: 10 Center Dr, Bldg 10, 12N240B

institute: National Institutes of Health

summary: To understand the function of CD25/IL-2Ra in the regulation of alopecia development, we sorted live CD45 + and CD45- population from the skin of Up_del mice and were subjected to single-cell RNA-sequencing.

overall_design: Whole skins from UP_Del mice with and without alopecia phenotype along with C57BL/6 WT mice were harvested. Hypodermal layers were removed and remaining epidermal and dermal layer were subjected to mincing followed by mechanical force and enzymatic treatments. Cells were liberated after enzymatic digestion of the tissues. Cells were stained with CD45 antibody and live CD45+ and CD45- population were sorted by fluorescence-activated cell sorting (FACS). Their transcriptomics changes assessed by single-cell RNA-sequencing using the 10X genomics platform.

first: Shubham

last: Goel

first: Warren

last: Leonard


city: Kashiwa

first: Fumihito

last: Miura

state: Chiba

address: Kashiwanoha 5-1-5

department: Graduate School of Frontier Sciences

country: Japan

institute: The University of Tokyo

type: Methylation profiling by high throughput sequencing

summary: Whole-genome bisulfite sequencing of Dnmt1/3a/3b  triple knockout (TKO) embryonic stem cells with or without expression of exogenous Dnmt1 before and after cell differentiation.

overall_design: Whole-genome bisulfite sequencing of TKO embryonic stem cells with or without expression of exogenous (full-length or its 602-aa N-terminal deletion mutant) Dnmt1 before and after cell differentiation. Cells were collected on day 0,  4, and 7 days after induction of differentiation, and served for whole-genome bisulfite sequencing.

first: Masako

last: Tada

first: Takamasa

last: Ito

first: Musashi

last: Kubiura-Ichimaru

first: Takashi

first: Shoji

last: Tajima


title: Transcriptomic analysis of Ganoderma Microsporum GMI-treated A549 NSCLC cell line.

city: Taipei

first: Wei-Lun

last: Hwang

address: No.155, Sec.2, Linong Street

department: Department of Biotechnology and Laboratory Science in Medicine

country: Taiwan

institute: National Yang Ming Chiao Tung University

summary: To investigate molecular mechanisms underlying anti-tumoral effects of GMI in lung cancer, we performed RNA-sequencing in GMI-treated A549 cells.

overall_design: A549 cells were treated with GMI to examine the differentially expressed profile by Illumia sequencing.

first: Tung-Yi

last: Lin

first: Wei-Jyun

last: Hua


title: Transcriptomic analysis of PLA-degrading bacteria Brevibacillus brevis grown on polylactic acid

city: Jeonju-si

first: Jaeyoung

last: Yu

address: Baekje-daero

country: South Korea

institute: Jeonbuk National University

summary: Polylactic acid (PLA) is a promising biodegradable material used in various fields, such as mulching films and disposable packaging materials. Biological approaches for completely degrading biodegradable polymers can provide environmentally friendly solutions. However, to our knowledge, no studies have performed transcriptome profiling to analyze PLA-degrading genes of PLA-degrading bacteria. Therefore, this study reports for the first time an RNA sequence approach for tracing genes involved in PLA biodegradation in the PLA-degrading bacterium Brevibacillus brevis. In the interpretation results of the differentially expressed genes, the hydrolase genes mhqD and nap and the serine protease gene besA were up-regulated by a fold change of 7.97, 4.89, and 4.09, respectively. This result suggests that hydrolases play a key role in PLA biodegradation by B. brevis. In addition, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analyses showed that genes implicated in biofilm formation were upregulated. The biodegradation of PLA starts with bacteria attaching to the surface of PLA and forming a biofilm. Therefore, it could be confirmed that the above genes were up-regulated for access to PLA and biodegradation. Our results provide transcriptome-based insights into PLA biodegradation, which pitch a better understanding of microbial biodegradation of plastics.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for PLA-degrading bacteria Brevibacillus brevis.

first: Yewon

last: Jang

first: Wooil

last: Choi

first: Sung-Kon

last: Kim

first: Jeehoon

last: Han

first: Jiho

last: Min


title: Massively parallel reporter assay investigates shared genetic variants of eight psychiatric disorders

city: Chapel Hill

first: Hyejung

last: Won

state: North Carolina

address: 116 Manning Dr, # 6110

department: Genetics and Molecular Biology

institute: UNC Chapel Hill

summary: A meta-genome-wide association study across eight psychiatric disorders has shed light on the genetic architecture of pleiotropy in major psychiatric disorders. However, the mechanisms underlying pleiotropic effects of the associated variants remain to be investigated. We conducted a massively parallel reporter assay to decipher the regulatory logic of variants with disorder-specific and pleiotropic effects. We found that pleiotropic variants differ from disorder-specific variants by manifesting chromatin accessibility that extends across diverse cell types in the neuronal lineage, as well as by altering motifs for transcription factors with higher connectivity in protein-protein interaction networks. Next, we mapped pleiotropic and disorder-specific variants to putative target genes using functional genomics approaches. CRISPR-mediated validation confirmed the functional impact of selected variants on predicted target genes. Furthermore, in vivo CRISPR perturbation of a pleiotropic and a disease-specific gene suggests that pleiotropy is mediated by the regulation of genes expressed in a broader range of neuronal types and with higher network connectivity.

overall_design: To functionally validate emVar-gene relationships, we used CRISPR droplet sequencing (CROP-seq) in human induced pluripotent stem cell (hiPSC)-derived neurons that constitutively express dCas9-KRAB and an inducible NGN2 cassette. Cells containing the CROP-seq construct were isolated by using Fluorescence-activated cell sorting (FACS) for RFP signal and analyzed via scRNA-seq.

first: Alejandro

last: Gomez


title: RNA-Seq analysis of PuCH35 overexpression line and wildtype Populus ussuriensis root tips in response to PEG stress

city: Harbin

first: Hao

middle: Cheng

last: Li

state: Heilongjiang

address: 26 Hexing Road

institute: Northeast Forestry University

summary: One week-old in-vitro plants of PuCH35 overexpression (PuC3H35-OE) lines and wildtype (WT) Populus ussuriensis were transfered to 1/2 MS medium supplemented with or without 6% PEG6000 for 7 days, and the root tips (0.5 cm long) were harvested. Samples were collected from three biological replicates (25 in-vitro plants each).

overall_design: RNA-seq analysis of the root tips (0.5 cm)

first: Chenghao

first: Yuting

last: He

first: Dandan

first: Minzhen

last: Zeng


title: Nuclear and Cytoplasmic HEK293 Cell 3' end mRNA-seq

city: Cambridge

first: Harry

last: Fischl

address: Babraham Research Campus

institute: Storm Therapeutics

summary: Alternative polyadenylation analysis on mRNA from nuclear and cytoplasmic fractions of HEK293 cells

overall_design: RNA was extracted from nuclear and cytoplasmic subcellular fractions of HEK293 cells. The 3’ ends of extracted RNA were generated into libraries using the QuantSeq 3’mRNA-Seq library kit for Ion Torrent (Lexogen). Libraries were loaded onto the Ion Chef System (ThermoFisher) for template preparation and chip loading and the resulting chips were sequenced on the Ion Proton Sequencing System (ThermoFisher). Sequences were aligned using the Ion Torrent Server TMAP aligner to genome build hg19.

first: Andre

last: Furger

first: Jane

last: Mellor


title: Distinct structural and functional heterochromatin partitioning of lamin B1 and lamin B2 revealed using genome-wide Nicking Enzyme Epitope targeted DNA sequencing.

city: Ipswich

first: Sriharsa

last: Pradhan

address: 240 County Road

institute: New England Biolabs

summary: Gene expression is regulated by chromatin DNA methylation and other features, including histone post-translational modifications (PTMs), chromatin remodelers, and transcription factor occupancy. A complete understanding of gene regulation will require the mapping of these chromatin features in small cell number samples. Here we describe a novel genome-wide chromatin profiling technology, named as Nicking Enzyme Epitope targeted DNA sequencing (NEED-seq). NEED-seq offers antibody-targeted controlled nicking by Nt.CviPII-pGL fusion to study specific protein-DNA complexes in formaldehyde fixed cells, allowing for both visual and genomic resolution of epitope bound chromatin. When applied to nuclei, NEED-seq yielded genome-wide profile of chromatin associated proteins and histone PTMs. Additionally, NEED-seq of lamin B1 and B2 demonstrated their association with heterochromatin. Lamin B1 and B2 associated domains (LAD) segregated to three different states, and states with stronger LAD correlated with heterochromatic marks. Hi-C analysis displayed A and B compartment with equal lamin B1 and B2 distribution, although methylated DNA remained high in B compartment. LAD clustering with Hi-C resulted in subcompartments, with lamin B1 and B2 partitioning to facultative and constitutive heterochromatin respectively and were associated with neuronal development. Thus, lamin B1 and B2 show structural and functional partitioning in mammalian nucleus.

overall_design: HT1080, NIH/3T3 and HCT116 cells were grown in 6 well plates until 70-80% confluency. After one wash with 1X PBS, cells were crosslinked in 1X PBS containing 4% formaldehyde for 10 min at room temperature (RT). Crosslinked cells were quenched using 0.25 M Glycine for 5 min at RT. Cells were de-crosslinked overnight at 58°C in 1X PBS and cytoplasm was extracted for 10-30 min on ice using CSK buffer (Supplementary File 1). After 1X PBS wash, cells were incubated with blocking buffer (Supplementary File 1) for 1 hr at RT followed by overnight incubation with primary antibodies at 4°C with gentle rocking. Next, fluorophore conjugated secondary antibodies were added for 1hr at RT after 3 washes with 1X T-PBS for 5 min at RT.  After addition of secondary antibody, cells were washes 3 times with 1X T-PBS for 5 min at RT and incubated for 1 hr at RT with 1 µl of Nt.CviPII-pGL in 800 µl NEED-seq binding buffer (Supplementary File 1) per well. Plates were then washed 3 times with NEED-seq washing buffer (Supplementary File 1) for 15 min at RT and incubated with 10 units of DNA Polymerase I, 5 M of dNTP mix containing biotinylated-dCTP for 1 hr at 37°C in 800 µl of 1X NEBufferTM 2 per reaction to allow nick translation to occur. Plates were again washed 3 times with NEED-seq washing buffer for 5-15 mins at RT with 1X PBS additional wash. DNA was extracted using Monarch genomic DNA purification kit. After DNA purification, genomic DNA was cleaved (digested) using 2 units of Nt.CviPII (NEB # R0626S) in 500 µl of 1X NEBufferTM 2 at 37°C  for 4 hrs to overnight (This ensures the nick translated 5mC conjugated cytosine in DNA be protected against digestion). NaCl 5M stock solution was added to reach 1 M final concentration (Streptavidin Binding Buffer, Supplementary File 1) along with 30 µl of Streptavidin magnetic beads for 2 hrs with end-over-end mixing at 4°C to capture biotinylated DNA fragments. Streptavidin bound DNA beads were washed twice with Streptavidin washing buffer (Supplementary File 1). A third wash was performed with 1X TE buffer. Beads were resuspended in 50 µl of TE buffer and libraries were made on beads using NEBNext® Ultra™ II DNA Library Prep Kit for Illumina. 8 to 12 PCR cycles were used to amplify barcoded DNA and DNA library was sequenced on Novaseq 6000. NEED-seq protocol described above was used with some modifications. Briefly, K562 crosslinked cells with 4% formaldehyde were de-crosslinked overnight at 58°C in 1X PBS. Cytoplasmic extraction was performed for 30 min with end-over-end mixing at 4°C. After 300 x g centrifugation at 4°C for 5 min, cell pellet (nuclei) was washed once with 1X PBS and resuspended in 1X PBS. Nuclei were then captured by adding 10 µl of preactivated Concanavalin A magnetic beads for 10 min with end-over-end mixing at RT. Bead-bound nuclei were incubated with blocking buffer (Supplementary File 1) for 1 hr at RT. Primary antibodies were added (1/400 final dilution) and incubated overnight with end-over-end mixing at 4°C. After 3 washes with 1X T-PBS (Supplementary File 1) for 5 min at RT, blocking buffer (Supplementary File 1) was added with fluorophore conjugated secondary antibody for 1 hr at RT. After 3 washes with 1X T-PBS (Supplementary File 1) for 5 min at RT, beads were resuspended in 1 ml NEED-seq binding buffer (Supplementary File 1) including fusion enzyme Nt.CviPII-pGL and incubated for 1 hr at RT. Beads were then washed 3 times with NEED-seq washing buffer (Supplementary File 1)  for 5-15 mins at RT. After 1 wash with 1X PBS, beads were incubated with DNA Polymerase I, 5 M of dNTP mix for 1 hr at 37°C in 800 µl of 1X NEBufferTM 2 with end-over-end mixing. After 3 washes with washing buffer (Supplementary File 1) for 15 min at RT, beads were resuspended in 50 mM Tris-Cl (pH 7.5), 1 % SDS, 200 mM NaCl and 2 µl of proteinase K and incubated overnight at 65°C to lyse nuclei bound to the beads. DNA was extracted using phenol-chloroform and precipitated using glycogen. 200 ng of DNA was fragmented using Nt.CviPII and used to make libraries as described above. For low number of cells, barcoded DNA was amplified using ~18 PCR cycles. To determine the dynamic range of NEED-seq, cells were serially diluted after the labeling reaction. 1 nM of size selected DNA library was sequenced on Novaseq 6000.

first: Pierre-Olivier

last: Estève

first: Sagnik

last: Sen


title: Transcriptomic characterisation of transitioning cell types in the skin of Atlantic salmon

status: Public on Jan 15 2025

city: Roslin

first: rose

middle: ruiz

last: daniels

address: The University Of Edinburgh

institute: The University of Edinburgh

summary:
Background
The skin maintains the body’s integrity and serves as the first line of defence against pathogens, stressors and mechanical injuries. Despite the global significance of salmon in aquaculture, how cells transcriptomic profile varies during wound healing, remains unexplored. Teleost contain adult pluripotent cells that differentiate into various tissues, including bone, cartilage, tendon, ligament, adipose, dermis, muscle and connective tissue within the skin. These cells are pivotal for preserving the integrity of skin tissue throughout an organism´s lifespan and actively participate in the wound healing processes. In this study, we characterise the transcriptomic profiles of putative mesenchymal stromal cells (fibroblast-like adult stem cells) in healthy Atlantic salmon tissue and during the wound healing process.
Results
Single-nucleus sequencing and spatial transcriptomics were used to detect transcriptomic changes occurring during wound healing that are commonly associated with mesenchymal stromal cells. We followed the transcriptomic activity of these cells during an in vivo wound healing time course study showing that these cells become more transcriptionally active during the remodelling stage of wound healing. The changes detected give insights into the potential differentiation pathways leading to osteogenic and fibroblast lineages in the skin of Atlantic salmon.
Conclusions
We chart the transcriptomic activity of sub-clusters of putative differentiating stromal cells during the process of wound healing for the first time, revealing different spatial niches of the various putative MSC subclusters, and setting the stage for further investigation of the manipulation of transitioning cell types to improve fish health.
KEYWORDS: Salmo salar, skin, mesenchymal stromal cells, single-cell sequencing, spatial transcriptomics, wound healing, spatial niche.

overall_design: Skin from both the body and fins of Atlantic salmon (Salmo salar) was analyzed using snRNA-seq.

first: Rose

last: Ruiz Daniels

first: Sarah

middle: J

last: Salisbury

first: Lene

last: Sveen

first: Paula

last: Rodriguez Villamayor

first: Richard

last: Taylor

first: Marianne

last: Vaadal

first: Torstein

last: Tengs

first: Aleksei

last: Krasnov

first: Sean

last: Monaghan

first: Maeve

last: Ballantyne

first: Carolina

last: Penaloza

first: Mark

last: Fast

first: James

last: Bron

first: Ross

last: Houston

first: Nicholas

last: Robinson

first: Diego

last: Robledo


title: Exploration of transcriptomic and proteomic profiles of the tumor microenvironment of marginal zone lymphoma

city: Rochester

first: Zhizhang

last: Yang

address: 200 1st ST SW

institute: Mayo Clinic

summary: The role of the tumor microenvironment (TME) and intratumoral T cells in splenic marginal zone lymphoma (sMZL) is largely unknown. In the present study, we evaluated 36 sMZL spleen specimens by single cell analysis to gain a better understanding of the TME in sMZL. Using mass cytometry (CyTOF), we observed that the TME in sMZL is distinct from that of control non-malignant reactive spleen (rSP). We found that the number of TFH cells varied greatly in sMZL, ICOS+ TFH cells were more abundant in sMZL than rSP, and TFH cells positively correlated with increased numbers of memory B cells. Treg cell analysis revealed that TIGIT+ Treg cells are enriched in sMZL and correlate with suppression of TH17 and TH22 cells. Intratumoral CD8+ T cells were comprised of subsets of short-lived, exhausted and late-stage differentiated cells, thereby functionally impaired. We observed that T-cell exhaustion was present in sMZL and TIM-3 expression on PD-1low cells identified cells with severe immune dysfunction. Gene expression profiling by CITE-seq analysis validated this finding. Taken together, our data suggest that the TME as a whole, and T-cell population specifically, are heterogenous in sMZL and immune exhaustion is one of the major factors impairing T-cell function.

overall_design: Biopsy specimens from 4 marginal zone lymphoma were sequenced using CITE-seq assay

first: Hyojin

last: Kim

first: Xinyi

last: Tang

first: Stephen

last: Ansell


title: Opto-CLIP reveals dynamic FMRP regulation of mRNAs upon CA1 neuronal activation [CLIP]

city: New York

first: Ruth

last: Singer

state: NY

address: 1230 York Avenue, TSH 314

institute: The Rockefeller University

summary: Neuronal diversity and function are intricately linked to the dynamic regulation of RNA metabolism, including splicing, localization, and translation. Electrophysiologic studies of synaptic plasticity, models for learning and memory, are disrupted in Fragile X Syndrome (FXS). FXS is characterized by the loss of FMRP, an RNA-binding protein (RBP) known to bind and suppress translation of specific neuronal RNAs. Since molecular studies have demonstrated that synaptic plasticity in CA1 excitatory hippocampal neurons is protein-synthesis dependent, together these observations have suggested a potential role for FMRP in synaptic plasticity in FXS. To explore this model, we developed a new experimental platform, Opto-CLIP, to integrate optogenetics with cell-type specific FMRP CLIP and RiboTag in CA1 hippocampal neurons, allowing investigation of FMRP- regulated dynamics after neuronal activation. We tracked changes in FMRP binding and ribosome- associated RNA profiles 30 minutes after neuronal activation. Our findings reveal a significant reduction in FMRP-RNA binding to transcripts encoding nuclear proteins, suggesting FMRP translational inhibition may be de-repressed to allow rapid translational responses required for neuronal homeostasis. In contrast, FMRP binding to transcripts encoding synaptic targets were generally stable after activation, but all categories of targets demonstrated variability in FMRP translational control. Opto-CLIP revealed differential regulation of subsets of transcripts within CA1 neurons rapidly after depolarization, and offers promise as a generally useful platform to uncover mechanisms of RBP-mediated RNA regulation in the context of synaptic plasticity.

overall_design: FMRP CLIP analyses of control and optogenetically activated CA1 excitatory neurons of the mouse hippocampus

first: Veronika

last: Rajchin

first: Robert

middle: B

last: Darnell


title: Opto-CLIP reveals dynamic FMRP regulation of mRNAs upon CA1 neuronal activation [RiboTag]

overall_design: RiboTag analyses of control and optogenetically activated CA1 excitatory neurons of the mouse hippocampus


title: Elevated levels of S100A8 and S100A9 exacerbate muscle mitochondrial fragmentation in sepsis-induced muscle atrophy

city: nanning

first: dongqin

last: huang

address: No. 50 Liangyu Avenue, Wuxiang New District,

department: aneashesiology department

institute: guang xi medical universty cancer hospital

summary: Sepsis-induced skeletal muscle atrophy is common in septic patients with the increases risk of mortality and is associated with myocellular mitochondrial dysfunction. Nevertheless, the specific mechanism of sepsis muscle atrophy remains unclear. Here we conducted a clinical retrospective analysis and observed the elevation of skeletal muscle index (ΔSMI) was an independent risk factor for 60-day mortality in septic patients. Moreover, in mouse model of sepsis, the skeletal muscle atrophy was also observed, which was associated with the upregulation of S100a8/a9-mediated mitochondrial dysfunction. Inhibition of S100a8/a9 significantly improved mitochondrial function and alleviated muscle atrophy. Conversely, administration of recombinant S100a8/a9 protein exacerbated mitochondrial energy exhaustion and myocyte atrophy. Mechanistically, S100a8/a9 binding to RAGE induced Drp1 phosphorylation and mitochondrial fragmentation, resulting in muscle atrophy. Additionally, RAGE ablation or administration of Drp1 inhibitor significantly reduced Drp1-mediated mitochondrial fission, improved mitochondrial morphology and function. Our findings indicated the pivotal role of S100a8/a9 in driving the mitochondrial fragmentation in septic muscle atrophy. Targeting S100a8/a9-RAGE-initiated mitochondrial fission might offer a promising therapeutic intervention against septic muscle atrophy. Taken together, our data provide a potential mechanism for sepsis-induced muscle atrophy.

overall_design: RNA-seq of skeletal muscle profiling of 2 independent studies: wildtype male mice at Day3 of sham or cecum ligation and puncture(CLP) operation, and wildtype male mice versus rage knockout male mice at Day3 of sham or CLP operation, respectively.

first: Dongqin

last: Huang

first: Yang

last: Li

first: Yuqian

last: Guo

first: Mengcao

last: Weng

first: Hui

last: Ye

first: Yan

last: Zhang

first: Fei

last: Lin

first: Kai

first: Xiangming

last: Fang


title: C/EBPβ suppresses keratinocyte autonomous type 1 IFN response and p53 to increase cell survival and susceptibility to UVB-induced skin cancer

city: RALEIGH

first: Dereje

last: Jima

state: NC

address: 850 Main Campus Drive

department: CHHE

institute: NCSU

summary: p53 is activated by DNA damage and oncogenic stimuli to regulate senescence, apoptosis and cell-cycle arrest, which are essential to prevent cancer. Here, we utilized UVB radiation, a potent inducer of DNA damage, p53, apoptosis and skin cancer to investigate the mechanism of CCAAT/enhancer binding protein-β (C/EBPβ) in regulating p53-mediated apoptosis in keratinocytes and to test whether the deletion of C/EBPβ in epidermis can protect mice from UVB-induced skin cancer. UVB-treatment of C/EBPβ skin conditional knockout (CKOβ) mice increased p53 protein levels in epidermis and enhanced p53-dependent apoptotic activity 3-fold compared with UVB-treated control mice. UVB increased C/EBPβ levels through a p53-dependent pathway and stimulated the formation of a C/EBPβ-p53 protein complex; knockdown of C/EBPβ increased p53 protein stability in keratinocytes. These results suggest a p53-C/EBPβ feedback loop, whereby C/EBPβ, a transcriptional target of a p53 pathway, functions as a survival factor by negatively regulating p53 apoptotic activity in response to DNA damage. RNAseq analysis of UVB-treated CKOβ epidermis unexpectedly revealed that type 1 interferon (IFN) pathway was the most highly enriched pathway. Numerous pro-apoptotic interferon stimulated genes were upregulated including some known to enhance p53 apoptosis. Our results indicate that p53 and IFN pathways function together in response to DNA damage to result in the activation of extrinsic apoptosis pathways and caspase 8 cleavage. Last, we observed CKOβ mice were resistant to UVB-induced skin cancer. Our results suggest that C/EBPβ represses apoptosis through keratinocyte autonomous suppression of the type 1 IFN response and p53 to increase cell survival and susceptibility to UVB-induced skin cancer.

overall_design: 4 replicates of K5Cre control and C/EBPB epidermial specific mice exposed to 100mJ/cm2 UVB or left  unexposed

first: Hann

last: Tam

first: Jonathan

last: Hall

first: Rob

last: Smart


title: KLF9-GRK5-HDAC6 axis aggravates osteoarthritis pathogenesis by promoting chondrocyte extracellular matrix degradation and apoptosis

city: x

first: Lunhao

last: Bai

address: x

institute: x

summary: Osteoarthritis (OA) is a degenerative joint disease that affects the cartilage and surrounding tissues. The transcription factor Kruppel-like family factor 9 (KLF9) has been identified as a regulator of tumorigenesis. However, its role in OA is still not fully understood. Herein, this study aimed to access the potential role and molecular mechanism by which KLF9 regulates OA development. KLF9 was upregulated in cartilage tissues of OA patients and medial meniscotibial ligament (MMTL)-induced OA rats, as well as in IL-1β-treated chondrocytes. Furthermore, knockdown of KLF9 inhibited OA-related cartilage injury, as evidenced by inhibiting chondrocyte extracellular matrix (ECM) degradation, increasing chondrocyte viability, and decreasing apoptosis. Conversely, overexpression of KLF9 had the opposite effect. The downstream mechanism of KLF9 was confirmed. KLF9 mediated the transcription of G protein-coupled receptor kinase 5 (GRK5) by directly targeting the GRK5 promoter. GRK5 knockdown eliminated the effects of KLF9 overexpression on chondrocyte dysfunction. It was also found that GRK5 combined with histone deacetylase 6 (HDAC6) and promoted HDAC6 phosphorylation. The use of the HDAC6 inhibitor TubastatinA also abolished the effects of GRK5 overexpression on chondrocyte ECM degradation and apoptosis. These results demonstrate that the KLF9-GRK5-HDAC6 axis plays a crucial role in promoting the progression of OA.

overall_design: Differential gene expression in patients with osteoarthritis was investigated by mRNA sequencing.

first: Xiaonan

last: Zhou

first: Peng

last: Jiang

first: Huangqi

last: Tan

first: Yanfang

last: Wang


title: Combinatorial phosphorylation on CTD of RNA polymerase II selectively controls transcription and export of protein-coding mRNAs

city: Austin

first: Svetlana

last: Panina

state: TX

address: 101 East 21st St.

department: Molecular Biosciences

institute: The University of Texas at Austin

summary:
C-terminal domain (CTD) of RNA polymerase II is crucial for recruiting transcription regulators via specific post-translational modifications (PTM), especially phosphorylation. The hypothesis of combination of PTMs, or ‘CTD code’, that can allow precise and dynamic recruitment of transcription machinery is highly attractive, yet the experimental evidence to support this hypothesis has been scarce. Here, despite lacking specific antibodies for combinatorial CTD phosphorylation, we developed an innovative approach that detects double phosphorylation patterns on the CTD in a whole-genomic fashion by leveraging the antibody masking effect with selectively removing the flanking interference. Using this method, we detected pT4pS5 double phosphosites occurring exclusively during the transcription of protein-coding genes. Furthermore, we showed that pT4pS5 marks recruit the Transcription and Export complex (TREX), which specifically facilitates mRNA processing and nucleocytoplasmic export of protein-coding mRNAs. The recruitment of TREX by pT4pS5 phosphosites is particularly important for the processing of lengthy neurogenesis-related genes. Our results provide experimental support for the notion that CTD coding system can function combinatorially and in a gene-specific manner, which encodes an exact information about the transcription of specific gene clusters.
This method can be broadly applied to map all combinatorial PTM patterns on RNA polymerase II, paving the way for a deeper understanding of gene-specific transcription regulation at the molecular level.

overall_design: Small RNA-Sequencing in T4A-CTD vs wild-type (WT) cells, two biological replicates per sample (HEK293T cells).

first: Seema

last: Irani

first: Haley

last: Hardtke

first: Renee

last: Stephenson

first: Brendan

middle: M

last: Floyd

first: Rosamaria

middle: Y

last: Moreno

first: Edward

last: Marcotte

first: Qian

first: Jessie


title: Comparison of transcriptomic data obtained from stimulated and unstimulated samples for the stratification of patients with a compromised immune system (in the context of sepsis and allogeneic hematopoietic stem cell transplantation)

city: Oullins-Pierre-Bénite

first: Marion

last: Debombourg

address: 165 chemindu grand revoyet

institute: biomerieux

summary: Recently, immune function assessment has gained prominence in clinical settings. Immune functional assays (IFAs), involving in vitro stimulation, offer a relevant approach to complement traditional immunomonitoring methods which, while widely used, do not fully capture functional immune capabilities. Despite growing interest in IFAs, their added value remains unclear.  To address this gap, our study aimed to determine if insights from IFAs could be replicated with unstimulated immunoprofiling. Using the same analytical pipeline, we compared transcriptomic profiles (Nanostring®) between stimulated (TruCulture®) and unstimulated (PaxGene™) samples from i) patients with an overstimulated immune system 3-4 days post-sepsis onset, and ii) patients undergoing immune reconstitution 6-months post-allogeneic hematopoietic stem cell transplantation (allo-HSCT).  In sepsis, post-stimulation transcriptomic profiles revealed immune clusters linked to disease severity and outcomes, surpassing traditional markers like mHLA-DR, while baseline analyses failed to generate clinically relevant stratification. Similarly, allo-HSCT patients’ post-stimulation data revealed immune heterogeneity and treatment-related alterations not detected using baseline transcriptomic or cellular profiles alone.  Our findings emphasize the value of IFAs in uncovering functional immune alterations that unstimulated assessments may miss, which could offer deeper insights into immune dysfunction. This study supports IFAs as complementary tools to current clinical practices, enhancing patient management with a functional view of immune system dynamics.

overall_design:
The study presented here is divided into 2  parts : (only the second part is represented here)
1. Sepsis context (REALISM cohort) : 28 patients sampled 3-4 days after sepsis onset as well as 10 healthy volonteers were included. These patients had already been analyzed in another study using functional immune tests and the NanoString platform, which highlighted a potential stratification of patients based on severity and clinical data through the obtained transcriptomic profiles
2. Allo-HSCT study (FIGHT cohort) : 59 patients sampled 6 months post allograft as well as 5 healthy volonteers were included. These patients had already been analyzed in another study using functional immune tests and the NanoString platform, which highlighted an association between the transcriptomic profiles of patients receiving immunosuppressive treatment and delayed immune reconstitution.

first: Chloé

last: Albert-Vega

first: William

last: Mouton


title: Dbp7 and RNA exosome subunit Dis3 interacts to mediate CENP-A loading via CENP-A chaperone Sim3

first: Fei

last: Li

state: New York

address: 24 Waverly Pl

department: Department of Biology

institute: New York University

summary: Background: Centromeres are crucial for proper chromosomes segregation during cell division. Centromeres in most eukaryotes are epigenetically specified by the histone 3 (H3) variant, CENP-A. How CENP-A precisely targets to centromeres remains elusive. Centromeres are typically transcribed into non-coding RNAs, which have been implicated in centromere functions. But the exact role of centromeric RNA in CENP-A loading is still poorly understood. In addition, centromere transcription induce formation of R-loops consisting of a DNA:RNA hybrid and a displaced single-stranded DNA. Excessive R-loops lead to genome instability. Little is known about the mechanism used for safeguarding centromeres against R-loops.  Results: Here we show that through a visual genetic screen, we identified a conserved RNA binding protein Dbp7 that plays a critical role in CENP-A loading to centromeres. Deletion of dbp7+ results in CENP-A mislocalization and defects in chromosome segregation and centromere silencing. We found that Dbp7 associates with centromere transcripts, and further showed that Dbp7 prevents accumulation of R-loops at centromeres. Our domain-deletion analysis demonstrated that the DEAD-box helicase domain and the C-terminal extension motif of Dbp7 contribute to its role in centromere function. In addition, Dbp7 interacts with the key exosome subunit Dis3. Importantly, we found that Dis3 promotes CENP-A centromere loading by interacting the CENP-A chaperone Sim3.  Conclusions: Together, this study uncovered a previously unrecognized CENP-A loading mechanism, by which Dbp7 and Dis3 bind to centromeric transcripts and in turn mediate recruitment of CENP-A via recruiting Sim3. Our work also provides mechanistic insight into R-loop resolution at centromeres.

overall_design: We analyzed dbp7Δ mutant cells and wild type carrying Cnp1-GFP by ChIP followed by high-throughput sequencing (ChIP-seq) to detect the distribution of Cnp1 in the two strains.

first: Jinxin

last: Gao

first: Fengying

first: Qianhua

last: Dong

first: Lening

last: Mao

first: Muhammad

last: Ali

first: Junwei

last: Liao

first: Jinpu

last: Yang


title: High-resolution whole-genome DNA methylation revealed unique signatures of painful diabetic neuropathy

city: Bologna

first: Paolo

last: Garagnani

state: BO

address: Via San Giacomo 12

department: Department of Medical and Surgical Sciences (DIMEC)

institute: Univeristy of Bologna

summary: This work describes the DNA methylation signature and identifies genes associated with neuropathic pain in type 2 diabetes mellitus. Genome-wide methylation data for 315 DNA (WB) samples were generated using Illumina Infinium Methylation EPIC v1.0 BeadChip. Four different selection criteria were used to identify promising pain-related genes. The findings revealed significant differences in methylation patterns between painful and painless diabetic neuropathy and identified a set of individual CpG sites of unique candidate genes associated with the painful phenotype. Several of these genes, including GCH1, MYT1L and MED16, have been previously linked to pain-related phenotypes or diabetes. Through pathway enrichment analysis, we demonstrated that specific epigenetic signatures could contribute to the complex phenotype of diabetic neuropathy and cluster analyses highlighted significant epigenetic dissimilarities between painful and painless phenotypes. Our results uncovered epigenetic differences between painful and painless diabetic neuropathy patients and identified targeted genes linked to neuropathic pain through DNA methylation mechanisms. This approach holds promise for investigating other chronic pain conditions, such as secondary chronic pain from cancer treatment, thoracic surgery, and various transplant settings.

overall_design: Bisulphite converted DNA isolated from 315 whole blood samples were hybridised to the Illumina Infiniun MethylationEPIC BeadChip following manufacture's protocols.

first: Katarzyna

last: Malgorzata Kwiatkowska


title: Comparison of transcriptomic data obtained from stimulated and unstimulated samples for the stratification of patients with a compromised immune system [REALISM cohort]

summary:
Comparison of transcriptomic data obtained from stimulated and unstimulated samples for the stratification of patients with a compromised immune system (in the context of sepsis and allogeneic hematopoietic stem cell transplantation)
Recently, immune function assessment has gained prominence in clinical settings. Immune functional assays (IFAs), involving in vitro stimulation, offer a relevant approach to complement traditional immunomonitoring methods which, while widely used, do not fully capture functional immune capabilities. Despite growing interest in IFAs, their added value remains unclear.  To address this gap, our study aimed to determine if insights from IFAs could be replicated with unstimulated immunoprofiling. Using the same analytical pipeline, we compared transcriptomic profiles (Nanostring®) between stimulated (TruCulture®) and unstimulated (PaxGene™) samples from i) patients with an overstimulated immune system 3-4 days post-sepsis onset, and ii) patients undergoing immune reconstitution 6-months post-allogeneic hematopoietic stem cell transplantation (allo-HSCT).  In sepsis, post-stimulation transcriptomic profiles revealed immune clusters linked to disease severity and outcomes, surpassing traditional markers like mHLA-DR, while baseline analyses failed to generate clinically relevant stratification. Similarly, allo-HSCT patients’ post-stimulation data revealed immune heterogeneity and treatment-related alterations not detected using baseline transcriptomic or cellular profiles alone.  Our findings emphasize the value of IFAs in uncovering functional immune alterations that unstimulated assessments may miss, which could offer deeper insights into immune dysfunction. This study supports IFAs as complementary tools to current clinical practices, enhancing patient management with a functional view of immune system dynamics.

overall_design:
The study presented here is divided into 2  parts : (only the first part is represented here)
1. Sepsis context (REALISM cohort) : 28 patients sampled 3-4 days after sepsis onset as well as 10 healthy volonteers were included. These patients had already been analyzed in another study using functional immune tests and the NanoString platform, which highlighted a potential stratification of patients based on severity and clinical data through the obtained transcriptomic profiles
2. Allo-HSCT study (FIGHT cohort) : 59 patients sampled 6 months post allograft as well as 5 healthy volonteers were included. These patients had already been analyzed in another study using functional immune tests and the NanoString platform, which highlighted an association between the transcriptomic profiles of patients receiving immunosuppressive treatment and delayed immune reconstitution.


title: Cellular cartography reveals mouse prostate organization and determinants of castration resistance [multiome]

city: Ann Arbor

first: Arul

last: Chinnaiyan

state: Michigan

address: 1500 E. Medical Center Dr

institute: University of Michigan

summary: Inadequate response to Androgen Deprivation Therapy (ADT) frequently arises in Prostate Cancer (PCa) via unclear cellular mechanisms and drives aggressive forms of the disease. Here, we integrated single-cell RNA sequencing, single-cell multiomics and spatial transcriptomics to reveal the transcriptional, epigenetic and spatial basis of cell identity and cell-specific castration response in mouse prostates, and used this reference along with meta-analysis to identify determinants of ADT-response and castration resistance in human PCa. We found that that the mouse prostate contains lobe-specific luminal epithelial cell types (LEs), as driven by unique gene regulatory modules, and anatomically-defined androgen responsive transcriptional programs, which together indicate developmental divergence. Androgen recalcitrant stem-like epithelia that bear semblance to Club and Hillock cells of the human prostate were enriched in the urethra and Ventral Prostate (VP), but rarely present in other lobes. Strikingly, the VP contained two additional LEs, Pbsn-positive and Spink1-positive (enriched in SPOP mutant PCa) androgen responsive cells, which effectively mapped to human LEs and potentially denote cell types that drive distinct PCa subtypes. Castration reorganized LE transcriptomes, with castration resistant LEs activating stress responsive (e.g. Nfe2l2) and stemness (e.g. Klf6) programs. Tumor cells of ADT-treated and Castration Resistant PCa patients were enriched for these castration programs, suggesting their role in driving the emergence and sustenance of castration resistance. Overall, our cellular cartography effort culminates in a comprehensive atlas of the mouse prostate, highlights the potential for lobe-specific PCa modeling in mice and identifies potential therapeutic targets that inhibit castration resistance.

overall_design: Prostates from sham operated or castrated mice were sequenced using 10x Chromium Next GEM Single Cell Multiome ATAC + Gene Expression to test for differentially accessible chromatin regions and differential nuclear RNA expression.

first: Hanbyul

last: Cho

first: Yuping

last: Zhang

first: Jean

last: Tien

first: Jie

last: Luo

first: Rahul

last: Mannan

first: Sathiya

middle: P

last: Narayanan

first: Somnath

last: Mahapatra

first: Greg

last: Shelly

first: Gabriel

last: Cruz

first: Lisha

last: Wang

first: Fengyun

last: Su

first: Rui

first: Saravana

last: Dhanasekaran

first: Xuhong

last: Cao

first: Evan

last: Keller

first: Sethuramasundaram

last: Pitchiaya


title: Cellular cartography reveals mouse prostate organization and determinants of castration resistance [scRNA-seq]

first: Arul

overall_design: Prostates from sham operated or castrated mice were sequenced by 10x Chromium 3`end single-cell RNA-seq to test for differentially expressed genes and differential transcription modules after castration.


title: AMPK phosphorylates LMX1b to regulate a brainstem neurogenic network important for control of breathing in neonatal mice

city: Riverside

first: Brendan

last: Gongol

state: Ca

address: 900 University Ave

institute: University of California, Riverside

summary: Ventilatory drive is modulated by a variety of neurochemical inputs that converge on spatially oriented clusters of cells within the brainstem. This regulation is required to maintain energy homeostasis and is essential to sustain life across all mammalian organisms. However, the mechanistic understanding of the regulation of these ventilatory control cellular clusters is poorly defined. This study explores the role of AMP-activated protein kinase (AMPK) and LIM homeobox transcription factor 1-beta (Lmx1b), a key player in dorsal-ventral patterning during fetal development, in the regulation of brainstem ventilatory control centers

overall_design: Brain stem tissue was isolated from AMPK+/+, AMPK-/-, LMX1B+/+, LMX1B+/-, or LMX1B-/- C57BL/6 mice and RNA sequencing expression profiling was performed.

first: Traci

last: Marin


title: Enhancing oncolytic virotherapy by extracellular vesicle mediated microRNA reprograming of the tumour microenvironment.

city: London

first: Anton

last: Patrikeev

state: London

address: Chester Beatty Laboratories, 237 Fulham Rd.

department: Targeted Therapy

institute: The Institute of Cancer Research

summary: miRNA expression is critical in driving macrophage polarisation by simultaneously regulating expression of multiple cellular targets. Reintroducing miRNA to promote an anti-tumour macrophage phenotype is a promising therapeutic approach to reverse the immunosuppressive function of tumour associated macrophages.Small RNA sequencing was used to identify differentially expressed miRNA during TAM generation and following LPS/IFNγ stimulation to induce an anti-tumour phenotype. Two differentially expressed miRNA identified, miR-155 and miR-19a, were cloned into oncolytic rhabdovirus (ORV), and anti-tumour efficacy was investigated using both in vitro and in vivo models of OvCa.

overall_design: To identify miRNA that are dysregulated during pro-tumour and anti-tumour TAM polarisation, we performed small RNA sequencing on in vitro SKOV-3 generated TAMs ± LPS/IFNg  and autologous monocytes from four healthy donors.

first: Victoria

last: Jennings


title: A multi-omics census reveals miRNA miR-let-7 as instigator of paternal mitochondrial dysfunction and in epigenetic heredity of intergenerational metabolic decline [F0_Liv_eWAT_Sperm_Small_RNAseq]

city: Odense

first: Jan

middle: Wilhelm

last: Kornfeld

address: Campusbej 50

department: Biochemistry and Molecular Biology

country: Denmark

institute: University of Southern Denmark

summary: We describe that diet-induced obesity and weight loss development in male mice causes reversible abnormalities in glucose and lipid metabolism, serum metabolomes, and adipose tissue and liver gene expression and causes specific, but reversible, reductions in activity of genes controlling mitochondrial energy dissipation in white adipose tissue. When mating obese mice with isogenic lean females, we observed highly concordant reductions in mitochondrial gene expression and translation in offspring that resemble those observed in obese fathers. When mapping small RNA responses across somatic and gametic cell types and between generations, we saw that obesity and weight loss reversible affected miRNA abundance, and observed several miR-let7 isoforms induced in obese adipose tissues of the paternal (F0) and offspring (F1) generation, as well as in sperm of F0 mice. Mechanistically, when overexpressing miR-let-7 in primary adipocytes, we found it to silence DICER1, a key rheostat required for adipose tissue adaptation in obesity as evidenced by functional deficiency in mitochondrial functioon following DICER1 loss. Upon ectopic delivery of miR-let7d/e levels into fertilized oocytes by microinjection, we elicited glucose intolerance and impairments in adipose mitochondrial gene expression in mice sired from miRNA-injected embryos, thus phenocopying key aspects of paternal obesity itself. When performing single-cell RNA-Seq in embryos, we found that miR-let7d/e delivery impaired mitochondrial gene expression, suggesting altered energy metabolism following sperm-mediated changes in zygotic miRNA pools. When studying miRNA alterations in human gametes, we found that lifestyle-induced weight loss downregulated MIR-LET7D/E in semen from human subjects, suggesting similar roles for human miR-let7d/e in gametic epigenomes and embryogenesis.

overall_design: Liver, eWAT and sperm small RNA profiles of LF, HF and HF-LF male mice in F0 generation were generated by deep sequencing using Illumina NovaSeq 6000 (n=4-6).

first: Chien

last: Huang

first: JooHyun

last: Park

first: Ali

last: Altintas

first: Natasa

last: Stanic

first: Hande

last: Batarlar

first: Rocio

last: Malmros

first: Jesper

last: Havelund

first: Nils

last: Færgeman

first: Bjørk

last: Larsen

first: Karolina

last: Szczepanowska

first: Jan-Wilm

last: Lackmann

first: Aleksandra

last: Trifunovic

first: Eva

middle: K

last: Hejbøl

first: Sönke

last: Detlevsen

first: Ida

middle: E

last: Jepsen

first: Stefanie

last: Kolstrup

first: Ricardo

last: Laguna-Barraza

first: Javier

last: Gonzalez

first: Konstantin

last: Khodosevich

first: Romain

last: Barrès

first: Jan-Wilhelm