title: Distinct role of perlecan plays in mesenchymal tissue regeneration through genetic and epigenetic modification

city: Morgantown

first: Gangqing

last: Hu

state: West Virginia

address: 64 Medical Center Drive

department: MicroBiology, Immunology, and Cell Biology

institute: West Virginia University

summary: Perlecan is a critical component of basement membrane proteins in tissue development. However, there are few studies investigating direct impact of perlecan on mesenchymal tissue differentiation using genetic modification (gain- and loss-of-function mutations) and epigenetic modification (matrix microenvironment). In this report, Cas9-mediated knockout (KO) of heparin sulfate proteoglycan 2 (HSPG2) and overexpression (OE) in human fetal nucleus pulposus stem/progenitor cells (NPSCs) and adult infrapatellar fat pad-derived stem cells (IPFSCs) were evaluated for the influence on mesenchymal differentiation. Furthermore, decellularized ECM (dECM) deposited by fetal NPSCs with either HSPG2 KO or OE was evaluated for its rejuvenation effect on adult NPSCs in proliferation and mesenchymal differentiation. We found that adult IPFSCs confirmed the unique role of perlecan in mesenchymal differentiation of fetal NPSCs; dECM with perlecan overexpression significantly promoted adult NPSCs in chondrogenic capacity rather than osteogenic and adipogenic capacities, indicating a potential application of perlecan in engineering smart biomaterial for cartilage regeneration.

overall_design: RNA-seq of adult infrapatellar fat pad-derived stem cells (IPFSCs) or human fetal nucleus pulposus stem/progenitor cells (NPSCs) under four treatment conditions: without transduction (CTR), transduced with lentivirus carrying GFP (GFP), transduced with lentivirus to knock out HSPG2 (KO1 and KO 2), and transduced with lentivirus to overexpress HSPG2 (OE).

first: Bin

last: Wang

first: Zhihua

last: Lu

first: Gongming

last: Gao

first: Elmira

last: Mikaeiliagah

first: Lei

first: Qingqing

last: Yu

first: Zhuo

first: Song

last: Chen

first: Xiaobing

last: Zhang

first: Ming

last: Pei


title: NAT10 and N4-Acetylcytidine restrain R-loop levels and related inflammatory responses

status: Public on Mar 26 2025

city: Austin

first: Blerta

last: Xhemalçe

state: Texas

address: 2500 Speedway

department: Molecular Biosciences

country: USA

institute: University of Texas at Austin

summary: N4-acetylcytidine (ac4C) is an evolutionarily conserved RNA modification that is deposited on diverse RNAs by N-acetyltransferase 10 (NAT10), a protein with high biological significance for aging and cancer. Here, we performed a comprehensive survey of ac4C using metabolic labeling, sodium cyanoborohydride chemical treatment coupled to next generation sequencing (NGS) and ac4C antibody-based cell and molecular biology techniques. Our NGS analysis confirms robust ac4C modification of rRNA and specific tRNA species in a NAT10-dependent manner, but suggests low or spurious ac4C acetylation in mRNA. Analysis of RNA-seq data also revealed an induction of inflammatory responses as well as mutagenesis at transcriptionally active sites in NAT10-KO cells. This finding led us to further explore the role of NAT10 in R-loops, which have recently been shown to induce APOBEC3B-mediated mutagenesis. Our analysis revealed that R-loops are modified with ac4C in a NAT10-dependent manner. Furthermore, NAT10 restrains the levels of R-loops at a subset of differentially expressed genes in a manner dependent on its catalytic activity. Together with cellular biology data showing ac4C containing RNA in endosomal structures, we propose that increased levels of ac4C-unmodified RNAs, likely derived from R-loops, in endosomal structures induce inflammatory responses.

overall_design: DRIP-seq: DNA/RNA hybrid Immunoprecipitation of HeLa Control+Mock, Control + RNase H, NAT10-KO, NAT10-KO +RNase H samples.

first: Turja

middle: K

last: Debnath

first: Nathan

last: Abell

first: Yi

middle: R

last: Li

first: Sravan

last: Devanathan

first: Enrique

last: Navedo


title: Chromatin interaction and histone mark signatures associated with TBXT expression in metastatic lung cancer

city: London

first: Reuben

middle: Mangi

last: Yaa

address: Sir Michael Uren Hub, White City Campus, 86 Wood Ln, London

department: Brain Sciences

country: United Kingdom

institute: Imperial College London

summary: The role of TBXT in cancer formation and progression is well studied, as is its expression regulation during embryonic development, but how its expression is activated and maintained in lung cancers remains elusive. Circularized chromosome capture combined with sequencing (4C-seq) was employed to analyze physical chromatin interactions with the TBXT promoter in the lung cancer cell line H460, a high TBXT expressing cell line, compared to H358 and A549, which do not express TBXT. Our analysis identified distinct patterns of potential cis-regulatory elements (pCREs) associated with the TBXT promoter, with increased near-cis pCRE enrichment in TBXT-expressing cells. Integration of pCREs with epigenetic histone modification revealed TBXT expression-specific regulatory landscapes, suggesting a role for epigenetic mechanisms in TBXT control. Two unique pCREs in TBXT-expressing H460 cells enriched with active histone mark H3K27ac, harbored binding sites for transcription factors of the forkhead box, zinc finger, and musculoaponeurotic fibrosarcoma families that are linked to cancer metastasis. Our findings shed light on the intricate control of TBXT expression in lung cancers, pointing to specific DNA elements and regulatory proteins that may be involved. This knowledge paves the way for understanding TBXT expression dynamics at the onset and progression of metastatic cancers and ultimately developing new therapeutic strategies.

overall_design: To investigate this, we employed circularized chromosome capture combined with sequencing (4C-seq) to analyze physical chromatin interactions with the TBXT promoter in lung cancer cell line H460, a TBXT expressing cell line compared to H358 and A549, non-TBXT expressors

middle: M


title: The transcriptional changes in the transcriptome of Control and Bacillus sphaericus(Bs)-treated Anopheles dirus mosquitoes

city: Chongqing

first: Shasha

address: 30 Gaotanyan street

country: China

institute: Army medical university

type: Expression profiling by high throughput sequencing

summary: We report the RNA-seq based analyses of the transcriptional changes in the transcriptome of Control and Bacillus sphaericus(Bs)-treated Anopheles dirus mosquitoes

overall_design: Comparison of the transcriptome of Anopheles dirus females at 24h past blood feeding in Control and Bs-treated mosquitoes


title: Pioneer transcription factor Runx2 affecting biological function of Schwann Cells via self-inducible reconstruction of super-silencer to remodel chromatin [Smart-seq mRNA]

city: Guangzhou

first: Zhaowei

last: Zhu

address: NO. 58 Zhongshan 2nd Rd, Guangzhou

department: Department of Plastic Surgery

institute: The First Affiliated Hospital of Sun Yat-sen University

summary: The expression of Runx2 in Schwann cells (SCs) increases significantly in the early stage of injury, but its underlying regulating mechanism remained unclear. After up-regulating Runx2 expression in SCs, the proliferation and migration ability of SCs decrease with increased cell apoptosis rate, and no obvious accelaration of axon regeneration rate could be seen when co-cultured with dorsal root ganlia. Through genome-wide mapping of the chromatin landscape, Runx2 displayed features of pioneer transcription factor (pTFs), which could independently action without histone modification, and execute positive feedback up-regulating gene expression of itself. In the meanwhile, it could construct the epigenetic mechanism of silencer to limit interactions of other transcription factors, then finally affect the fate of SCs, expecially SCs’ ability to deal with lipoproteins and form myelin. This study provides noval insight into key function of Runx2 in SCs as pTFs relying on its ability to reorganize the chromatin landscape within SCs, and interfering with the metabolism and synthesis of myelin sheaths by forming super silencers, therefore affecting the regeneration process of peripheral nerves.

overall_design: SCs from neonatal 3-5 day SD rats were isolated and purified. The purified P0 SCs were randomly divided into the Runx2-OE group and EGFP (empty vector) group. SCs were transfected with Runx2-OE or EGFP adenovirus at a concentration of MOI = 200. After 72 hours of incubation in the incubator, the changes in cell morphology were observed under laser confocal microscope. Smart-seq was performed as described previously.

first: Bo

last: He


title: Pioneer transcription factor Runx2 affecting biological function of Schwann Cells via self-inducible reconstruction of super-silencer to remodel chromatin [TF CUT&Tag]

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: SCs from neonatal 3-5 day SD rats were isolated and purified. The purified P0 SCs were randomly divided into the Runx2-OE group and EGFP (empty vector) group. SCs were transfected with Runx2-OE or EGFP adenovirus at a concentration of MOI = 200. After 72 hours of incubation in the incubator, the changes in cell morphology were observed under laser confocal microscope. CUT&TAG assay was performed as described previously with modifications.


title: Early life stress modulates neonatal somatosensation and the transcriptional profile of immature sensory neurons

city: Cincinnati

first: Mark

middle: L

last: Baccei

state: OH

address: 231 Albert Sabin Way

department: Anesthesiology

institute: University of Cincinnati

summary: Early life stress (ELS) is associated with an increased risk of experiencing chronic pain during adulthood, but surprisingly little is known about the short-term influence of ELS on nociceptive processing in the immature nervous system and the concomitant effects on somatosensation in the neonate.  Here we investigate how ELS modulates pain in neonatal mice as well as the transcriptional and electrophysiological signatures of immature dorsal root ganglia (DRG). Shortly after the administration of a neonatal limiting bedding (NLB) paradigm from postnatal days (P)2 to P9, both male and female pups exhibited robust hyperalgesia in response to mechanical, pressure and noxious cold stimuli compared to a control group housed under standard conditions, with no change in their sensitivity to noxious heat. Bulk RNA-seq analysis of L3-L5 DRGs at P9 revealed significant alterations in the transcription of pain- and itch-related genes following ELS, highlighted by a marked downregulation in Sst, Nppb, Chrna6, Trpa1 and Il31ra. Nonetheless, ex vivo whole cell patch-clamp recordings from putative A- and C-fiber sensory neurons in the neonatal DRG found no significant changes in their intrinsic membrane excitability following NLB. Overall, these findings suggest that ELS triggers hyperalgesia in neonates across multiple pain modalities that is accompanied by transcriptional plasticity within developing sensory neurons. A better understanding of the mechanisms governing the interactions between chronic stress and pain during the neonatal period could inform the future development of novel interventional strategies to relieve pain in infants and children who have experienced trauma.

overall_design:
Neonatal mouse pups underwent neonatal limited bedding protocol (stress) or control bedding protocol (control) from postnatal day (P)2-9
Lumbar L3-L5 DRG were harvested at P9, and total RNA was extracted for sequencing.
Differential gene analysis was performed to identify stress-induced transcriptional changes.

first: Kyle

last: Harbour

first: Fady

last: Eid

first: Elizabeth

last: Serafin

first: Madailein

last: Hayes


title: Exploring the functional, protective, and transcriptomic effects of GIP on cytokine-exposed human pancreatic islets and EndoC-βH5 cells

city: Herlev

first: Joachim

last: Størling

address: Borgmester Ib Juuls Vej 83,

country: Denmark

institute: Steno Diabetes Center Copenhagen

summary: Immune-mediated beta cell destruction and lack of alpha cell responsiveness to hypoglycaemia are hallmarks of type 1 diabetes pathology. The incretin hormone glucose-dependent insulinotropic polypeptide (GIP) may hold therapeutic potential for type 1 diabetes due to its insulinotropic and glucagonotropic effects as well as its beta cell protective effects shown in rodent islets. Here, we examined the functional and transcriptomic effects of GIP treatment upon diabetogenic cytokine exposure to interleukin (IL)-1β ± interferon (IFN)-γ in human EndoC-βH5 beta cells and isolated human islets, respectively.  GIP dose-dependently augmented glucose-stimulated insulin secretion from EndoC-βH5 cells and increased insulin and glucagon secretion from human islets during high and low glucose concentrations, respectively. The insulinotropic effect of GIP in EndoC-βH5 cells was abrogated by KN-93, an inhibitor of calcium/calmodulin-dependent protein kinase 2 (CaMK2). GIP did not prevent cytokine-induced apoptosis or cytokine-induced functional impairment of human EndoC-βH5 cells. GIP also did not prevent cytokine-induced apoptosis in human islets. GIP treatment of human islets with or without cytokines for 24 hours did not significantly impact the transcriptome. GIP potentiated cytokine-induced secretion of IL-10 and c-c motif chemokine ligand (CCL)-2 from human islets while decreasing the secretion of c-x-c motif chemokine ligand (CXCL)-8. In EndoC-βH5 cells, GIP reduced IFN-γ-induced secretion of tumor necrosis factor (TNF)-α, IL-2, IL-6, and IL-10, but increased the secretion of CXCL8, CCL2, CCL4, and CCL11.  In conclusion, our results suggest that the insulinotropic effect of GIP is CaMK2-dependent. Furthermore, our results indicate that GIP does not provide substantial cytoprotective effects against diabetogenic cytokine challenge or significantly modulate the transcriptome of human islets when applied at a supraphysiological level. GIP may, however, still exert selective inflammation-modulatory effects upon diabetogenic cytokine exposure.

overall_design: Human pancreatic islets from cadaveric donors (n=4) were obtained from Prodo Labs, Inc. Islets were incubated/exposed/treated ± IL-1β and IFN-γ ± GIP for 24 hours

first: Kristine

last: Henriksen

first: Anne

last: Jørgensen

first: Simranjeet

last: Kaur


title: Pioneer transcription factor Runx2 affecting biological function of Schwann Cells via self-inducible reconstruction of super-silencer to remodel chromatin [hitone CUT&Tag]


title: RNA-Seq shows that HAVCR1 knockdown does not affect JEG3 interferon signaling response

city: Xiamen

first: Jun

last: Tao

state: Fujian

address: No. 4221, South Xiang'an Road

institute: XMU

summary: To investigate whether HAVCR1 knockout (KO) affects the response of the interferon signaling pathway in JEG3 cells, RNA-Seq was performed on HAVCR1 KO and control JEG3 cells under both basal conditions and stimulation with double-stranded RNA mimic poly I:C. The results indicated that HAVCR1 knockdown does not affect the interferon signaling pathway response in JEG3 cells. Both groups exhibited comparable activation of the interferon signaling pathway.

overall_design: The experimental design involves comparing HAVCR1 knockout (KO) JEG3 cells and control JEG3 cells under two conditions: basal conditions and stimulation with the double-stranded RNA mimic poly I:C

first: Wenzhe

last: Yu


title: Identification of modifications to the maternal-fetal interface transcriptome associated with placental insufficiency and a novel gene therapy for the treatment of fetal growth restriction (Subplacenta)

city: Gainesville

first: Rebecca

last: Wilson

address: 1345 Center Dr

institute: University of Florida

summary: The etiology of fetal growth restriction (FGR) is multifactorial, although many cases often involve placental insufficiency. Placental insufficiency is associated with inadequate trophoblast invasion that results in an environment with high resistance to blood flow, decreased availability of nutrients to the placenta, and increased hypoxia. Despite accumulating knowledge linking inadequate trophoblast invasion with placental insufficiency and FGR, current treatment options are limited to iatrogenic delivery, usually preterm. We have developed a non-viral, polymer-based nanoparticle that facilitates transient human insulin-like 1 growth factor (hIGF1) gene delivery specifically to placental trophoblast. Using the established guinea pig maternal nutrient restriction (MNR) model of placental insufficiency and FGR, the aim of the study was to identify novel pathways in the sub-placenta/decidua that will provide insight into the underlying mechanism driving FGR, and may be corrected with hIGF1 nanoparticle treatment. Pregnant guinea pig dams underwent ultrasound-guided sham or hIGF1 nanoparticle treatment at mid-pregnancy, and sub-placenta/decidua tissue was collected 5 days later. Transcriptome analysis was performed using RNA Sequencing on the Illumina platform. Histological assessment of the sub-placenta/decidua demonstrated fewer maternal spiral arteries lined by trophoblast in the MNR sub-placenta/decidua which was associated with downregulation of genes involved in epithelium development and the regulation of cell migration. hIGF1 nanoparticle treatment results in marked changes to transporter activity in the MNR + hIGF1 sub-placenta/decidua when compared to sham MNR. Under normal growth conditions however, hIGF1 nanoparticle treatment in Control + hIGF1 sub-placenta/decidua was associated with downregulation of kinase signaling and increased proteolysis indicative of homeostasis. Overall, this study identified changes to the sub-placenta/decidua transcriptome that likely result in inadequate trophoblast invasion and that contribute to placental insufficiency. Additionally, this study has increased our understanding of pathways that hIGF1 nanoparticle treatment acts on in order to restore or maintain appropriate placenta function.

overall_design:
Female (dams) Dunkin-Hartley guinea pigs were assigned to either the control diet group (n = 6) or MNR diet group (n = 7). Control dams were provided food  and water was ad libitum; MNR dams were provided water ad libitum however, food intake was restricted to 70% per kilogram body weight of the Control group.
At GD30-33, dams underwent an ultrasound-guided, transuterine, intra-placental injection of either sham or hIGF1 nanoparticle. Dams were sacrificed five (GD35-38) days after hIGF1 nanoparticle treatment. Sub-placenta/decidua tissue from two fetuses per litter (one female and one male) were blunt dissected away from the rest of the maternal-fetal interface and snap-frozen in liquid nitrogen for RNA extraction.
RNA-Seq Libraries for each gender and experimental group were generated using the Illumina Stranded mRNA Prep Kit. RNA-Seq Libraries were sequenced using the Illumina NovaSeq platform.

first: Helen

middle: N

last: Jones

first: Baylea

last: Davenport


title: Identification of modifications to the maternal-fetal interface transcriptome associated with placental insufficiency and a novel gene therapy for the treatment of fetal growth restriction  (Placenta)

summary: Fetal growth restriction (FGR) affects between 5-10% of all live births. Placental insufficiency is a leading cause of FGR, resulting in reduced nutrient and oxygen delivery to the fetus. Currently, there is no effective in utero treatment options for FGR, or placental insufficiency. We have developed a gene therapy to deliver human insulin-like 1 growth factor (hIGF1) to the placenta via a non-viral nanoparticle delivery mechanism as potential treatment of FGR. Using a guinea pig maternal nutrient restriction (MNR) model of FGR, we aimed to understand the transcriptional changes within the placenta associated with placental insufficiency that occur at the beginning stages of FGR (mid-pregnancy), and the immediate impact of hIGF1 nanoparticle treatment on the placental transcriptome. Using RNAsequencing, we analyzed protein coding genes of three experimental groups: Control dams and MNR receiving a sham treatment, and MNR dams receiving hIGF1 nanoparticle treatment. Pathway enrichment analysis comparing MNR placentas to Control revealed upregulation of pathways associated with degradation and repair of genetic information and downregulation of pathways associated with transmembrane transport. Similarly, differentially expressed genes that were decreased in MNR + hIGF1 placentas compared to Control demonstrated downregulation in pathways relating to transporter activities, but upregulation in pathways associated with positive regulation of phosphorylation and kinase activity. When compared to MNR placentas, MNR + hIGF1 placentas demonstrated changes to genes associated with transmembrane transporter activity including ion, vitamin and solute carrier (SLC-mediated) transport. Overall, this study identifies the key signaling and metabolic changes occurring in the placenta that contribute to placental insufficiency, and increases our understanding of the pathways that increasing placental IGF1 expression acts on and corrects.

overall_design:
Female (dams) Dunkin-Hartley guinea pigs were assigned to either the control diet group (n = 4) or MNR diet group (n = 7). Control dams were provided food  and water was ad libitum; MNR dams were provided water ad libitum however, food intake was restricted to 70% per kilogram body weight of the Control group.
At GD30-33, dams underwent an ultrasound-guided, transuterine, intra-placental injection of either sham (Control and MNR) or hIGF1 nanoparticle (MNR). Dams were sacrificed five (GD35-38) days after hIGF1 nanoparticle treatment. Placenta tissue from two fetuses per litter (one female and one male) was blunt dissected away from the rest of the maternal-fetal interface and snap-frozen in liquid nitrogen for RNA extraction.
RNA-Seq Libraries for each gender and experimental group were generated using the Illumina Stranded mRNA Prep Kit. RNA-Seq Libraries were sequenced using the Illumina NovaSeq platform.


title: Protein tyrosine phosphatase delta is a STAT3-phosphatase and suppressor of metabolic liver disease (HepaRG cells).

city: Strasbourg

first: Julien

last: Moehlin

address: 3 Rue Koeberlé

country: France

institute: Institute for Translational Medicine and Liver Disease (ITM)

summary: Background & Aims: Protein tyrosine phosphatase delta (PTPRD) is suppressed in several diseases including HCV infection. To identify hepatic pathways responsive to PTPRD function and their role in non-viral liver disease, we analyzed liver transcriptomic and clinical data from patients and established a Ptprd-deficient liver disease mouse model. Methods: Healthy patients were classified according to hepatic PTPRD expression and transcriptomic analysis was performed to identify signaling pathways associated with low PTPRD levels. We combined an animal model for metabolic dysfunction-associated steatohepatitis (MASH) with genetically impaired PTPRD expression (Ptprd+/-) to assess its impact on the liver transcriptome and metabolic function. Identified pathways were validated by perturbation studies in primary human hepatocytes and differentiated HepaRG cells. Substrate specificity was validated by pull-down assay. The clinical relevance was explored in a cohort of patients with fatty liver disease by ranking individuals according to hepatic PTPRD expression and analyzing its association with metabolic disease markers. Results: In healthy individuals and Ptprd+/- mice, PTPRD levels associated with hepatic glucose/lipid metabolism and peroxisomal function. Hepatic PTPRD expression is impaired in metabolic liver disease. Moreover, we revealed PTPRD as a STAT3 phosphatase in the liver, which is a regulator of peroxisomal function. Silencing of STAT3 in HepaRG cells treated with free-fatty acids was able to rescue the expression of genes implicated in lipid metabolism. During MASH, low hepatic PTPRD led to increased liver steatosis in the Ptprd+/- animals and pronounced unfolded protein response, which impacts insulin signaling. Silencing of PTPRD in PHH blunted insulin-induced AKT phosphorylation. In line with this, obese patients with low hepatic PTPRD expression exhibit increased levels of clinical markers associated with metabolic disease. Conclusion: Our data suggest an important regulatory role of the hepatic PTPRD-STAT3 axis in maintaining glucose/lipid homeostasis and how its impaired expression is associated with clinical manifestations of metabolic liver disease.

overall_design: DMSO-differentiated HepaRG cells were transfected with siRNAs against STAT3 or/and PTPRD. Cells were cultured for 48 h and subsequently treated with free-fatty acids (800 µM oleic acid and 400 µM palmitic acid) for 24 h. Total RNA was extracted from the samples using the Direct-zol kit (Zymo Research) in order to perform RNA-seq.

first: Armando

last: Andres Roca Suarez

first: Thomas

middle: F

last: Baumert

last: Lupberger


title: Selenoprotein-Mediated Redox Regulation Shapes the Cell Fate of HSCs and Mature lineages [bulkRNA-seq]

city: Suita

first: Masaki

last: Nomura

state: Osaka

address: 2-2, Yamadaoka

department: Department of Cancer Pathology

country: Japan

institute: Graduate School of Medicine, The University of Osaka

summary: The maintenance of cellular redox balance is crucial for cell survival and homeostasis and is disrupted with aging. Selenoproteins, comprising essential antioxidant enzymes, raise intriguing questions about their involvement in hematopoietic aging and potential reversibility. First, we observed a notable mRNA downregulation of key antioxidant selenoproteins in aged human hematopoietic stem cells (HSCs). Therefore, we employed tRNAsec gene (Trsp) knockout (KO) mouse model to simulate disrupted selenoprotein synthesis, revealing the insights into the protective roles of selenoproteins in preserving HSC stemness and facilitating B cell maturation, despite negligible effects on myeloid cells. Notably, Trsp KO exhibited B lymphocytopenia and reduced HSCs’ self-renewal capacity, reminiscent of aged phenotypes, along with the upregulation of aging-related genes in both HSCs and Pre-B cells. While Trsp KO activated an antioxidant response transcription factor NRF2 across all hematopoietic cells, we delineated a lineage-dependent phenotype driven by lipid peroxidation, which was exacerbated with aging yet ameliorated by ferroptosis inhibitors such as vitamin E. Interestingly, the myeloid genes were ectopically expressed in Pre-B cells of Trsp KO mice and the KO Pre-B/Pro-B cells displayed differentiation toward CD11b+ fraction in the transplant model, suggesting that disrupted selenoprotein synthesis induces the potential of B to myeloid switch. Collectively, our findings underscore the critical role of selenoprotein-mediated redox regulation in maintaining balanced hematopoiesis and highlight the preventive potential of selenoproteins against aging-related hematopoietic alterations.

overall_design: For 7 weeks after birth, the mice were injected 500 µg/mouse polyinosine-polycyticylic acid (pI-pC) every other day for a total of 3 doses. 9 weeks after the last pIpC injection, the mice were sacrificed for sample preparation.

first: Yumi

last: Aoyama

first: Weijia

last: Zang

first: Hiromi

last: Yamazaki

first: Daichi

last: Inoue


title: Gentle, Effective, and Timely Isolation of Extracellular Vesicles that is Amenable for Transcriptomics Using Next-Generation Aqueous Two-Phase System Technology

city: Toronto

first: Gobi

last: Thillainadesan

state: Ontario

address: 2075 Bayview Ave., S1-18

department: Biology

country: Canada

institute: Sunnybrook Research Institute

type: Non-coding RNA profiling by high throughput sequencing

summary: The isolation of extracellular vesicles (EVs) using currently available methods frequently compromises purity and yield to prioritize speed. Here, we present a next generation aqueous two-phase system (ATPS) for isolation of EVs regardless of scale and volume and is superior to conventional methods such as ultracentrifugation and commercial kits. This is made possible by the two aqueous phases, one rich in polyethylene glycol (PEG), and the other rich in dextran (DEX), whereby fully encapsulated lipid vesicles preferentially migrate to the DEX-rich phase to achieve a local energy minimum for the EVs. Isolated EVs as found in the DEX-rich phase are more amenable to biomarker analysis such as nanoscale flow cytometry when using various pre-conjugated antibodies specific for CD9, CD63, and CD81. TRIzol RNA isolation is further enabled by the addition of Dextranase, a critical component of this next generation ATPS method. This negates the use of specialized EV RNA extraction kits. The use of Dextranase also enables more accurate immunoreactivity of pre-conjugated antibodies for detection of EVs by nanoscale flow cytometry. Transcriptomics of isolated EVs with ATPS revealed strong overlap between micro RNA (miRNA), circular RNA (circRNA), and small nucleolar RNA (snoRNA) profiles of EVs isolated compared to UC and was superior to other kits. Overall, this ATPS method stands out as a rapid and highly effective approach to isolate EVs, ensuring optimal extraction and analysis of nucleic acids.

overall_design: Media was collected from various cells and extracelluar vesicles (Evs) were isolated using either ultracentrifuge, the aqueous two-phase system (ATPS), or Qiagen exoRNeasy Midi Kit. Total RNA was extracted using Trizol and small RNA library was prepared and sequenced.

first: Boyang

last: Su

first: Morteza

last: Jeyhani

first: Reese

last: Wunsche

first: Minzhi

last: Sheng

first: Thamara

last: Dayarathna

first: Kristin

last: Cimolai

first: Hanyi

last: Weng

first: Katerzyna

last: Jerzak

first: Stanley

last: Liu

first: Scott

middle: S

last: Tsai

first: Hon

last: Leong


title: Effect of vitamin K on gene expression in erastin induced ARPE-19 cells.

city: Shanghai

first: Xiaochan

last: Dai

address: Handan Road, No.220

institute: Fudan University

summary: We report here the effect of vitamin K1 on gene expression in ARPE-19 cells with erastin induced iron death injury, in order to explore the important role and potential mechanism of vitamin K in iron death injury in RPE.

overall_design: We treated ARPE-19 cell lines with Erastin and pre-treated them with/without vitamin K in different groups

first: Xi

last: Yang


title: Selenoprotein-Mediated Redox Regulation Shapes the Cell Fate of HSCs and Mature lineages [scRNA-seq]

first: Masaki

last: Nomura

first: Yumi

last: Aoyama


title: A Drosophila model of mucopolysaccharidosis IIIB

city: Greenwood

first: Bibhu

last: Simkhada

state: SC

address: 114 Gregor Mendel Cir

department: Department of Genetics and Biochemistry

institute: Clemson University

summary: Mucopolysaccharidosis IIIB (MPS IIIB) is a rare genetic disorder caused by defects in α-N-acetylglucosaminidase, which breaks down extracellular heparan sulfate. Symptoms progress from hyperactivity and sleep disruptions during childhood to severe neurodegeneration during adolescence. To generate an MPS IIIB model in the fruitfly, Drosophila melanogaster, we introduced mutations in the corresponding fly enzyme that correlate with patient mutations. The fly model recapitulates sleep fragmentation and hyperactivity observed in patients and enlarged lysosomes in the brain. Genes associated with synaptic function and neurodevelopment showed altered expression in the brain. The Drosophila model can enable future development of therapies for MPS IIIB.

overall_design: To investigate the differential gene expression in MPS IIIB progression, including the effects of various mutations, sex and age

first: Nestor

middle: O

last: Nazario-Yepiz

first: Patrick

last: Freymuth

first: Rachael

middle: A

last: Lyman

first: Vijay

last: Shankar

first: Robert

last: Anholt

first: Trudy

last: Mackay


title: Recapitulating cartilaginous organoids and hypoxic metabolic switching via adaptive stem cell-matrix interactions promotes histone lactylation and cartilage regeneration

city: Hong Kong

first: Boguang

address: Sha Tin

country: Hong Kong

institute: The Chinese University of Hong Kong

summary: The formation of cartilaginous organoids fostered by the cell–hydrogel adaptation creates a hypoxic microenvironment and induces metabolic switching to favor glycolysis and the accumulation of lactate in the MSC spheroids, leading to significantly elevated histone lysine lactylation (Kla). We further show that the increased H3K18la enhances chondrogenesis and cartilaginous matrix deposition by regulating the transcriptional accessibility of chondrogenic genes.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone modification H3K18la in hMSCs encapsulated in the hydrogels with different dynamic properties.


title: E-cadherin has a cell-type specific role in maintaining the integrity of pulmonary airways

city: San Diego

first: Sunyoung

last: Lee

state: CA

address: 9500 Gilman Drive

department: Pediatrics

institute: University of California, San Diego

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

overall_design: Refer to individual Series


title: E-cadherin has a cell-type specific role in maintaining the integrity of pulmonary airways [scRNA-seq]

summary: We performed single cell RNA sequencing to investigate differentially expressed genes in each airway cell type after Cdh1 inactivation in Club cells.

overall_design: Immune cells and endothelial cells were excluded by depletion and epithelial cells were selected for by EPCAM expression and then sorted into GFP+ and GFP- populations.

first: Randee

last: Young


title: Comprehensive Analysis of Phenotypes and Transcriptome Characteristics Reveal the Best Atopic Dermatitis Mouse Model Induced by MC903

city: Nanjing

first: shan

last: zhang

state: Jiangsu

address: 12 Jiangwangmiao St

institute: Hospital for Skin Diseases, Institute of Dermatology, Chinese Academy of Medical Sciences and Peking Union Medical College

summary: Although several mouse models of exogenous-agent–induced atopic dermatitis (AD) are currently available, the lack of certainty regarding their similarity with human AD has limited their scientific value. Thus, comprehensive evaluation of the characteristics of mouse models and their similarity with human AD is essential. The DNFB plus OVA model showed the highest disease severity, while the OVA model showed the lowest severity. The MC903 and MC903 plus OVA models showed high expression of T-helper(Th)2- and Th17-related genes; the DNFB and DNFB plus OVA models showed upregulation of Th1-, Th2-, and Th17-related genes; while the S. aureus inoculation model showed more enhanced Th1 and Th17 immune responses. In contrast to the other models, the OVA-induced model showed the lowest expression levels of inflammation-related genes, while the MC903 model shared the largest overlap with human AD profiles. Each AD mouse model exhibited different characteristics. The MC903 model was the best to recapitulate most features of human AD among these exogenous-agent–induced AD models.

overall_design: To compare six different exogenous-agent–induced AD mouse models and find out the optimum models for study. Female BALB/c mice underwent induction of AD-like dermatitis by MC903 alone or in combination with ovalbumin (OVA), dinitrofluorobenzene (DNFB) alone or in combination with OVA, OVA alone, or Staphylococcus aureus.


title: UBR7 in concert with EZH2 inhibits the TGF-β signalling leading to extracellular matrix remodelling [RNA-seq]

city: Bangalore

first: madavan

last: vasudevan

state: Karnataka

address: Kasturi Nagar

country: India

institute: Theomics International Private Limited

summary: The intricate interplay between resident cells and the extracellular matrix (ECM) wields a profound influence on cancer progression. In Triple Negative Breast Cancer (TNBC), ECM architecture evolves primarily due to the enrichment of lysyl oxidase, fibronectin, and collagen that eventually promote distant metastasis. Here, we have uncovered a pivotal transcription regulatory mechanism of the epigenetic regulator UBR7 in conjunction with histone methyltransferase EZH2 in regulating TGF-b/Smad signalling-axis thereby fine-tuning the ECM genes. Remarkably, UBR7 loss leads to a dramatic reduction in the facultative heterochromatin mark H3K27me3 in chromatin, leading to ECM genes firing. Apart from playing a seminal role in matrix deposition in adherent-cancer cells and spheroids, UBR7 alters the total collagen content and lysyl oxidase activity thereby directly impacting the matrix stiffness and consequently its invasive properties. These results can be further translated to TNBC-patients, where reduced RNA/protein levels of UBR7 is accompanied by heightened expression of the ECM-components and their functional-activity which is responsible for fibrosis-mediated matrix stiffness. Thus, UBR7 acts as a master regulator of TNBC matrix-stiffening thereby impacting its metastatic potential.

overall_design: RNA sequencing of UBR7 knockdown in MDA-MB-231 cells. RNA sequencing of stable cells transfected with scrambled RNA and UBR7-Sh vector.

first: Chandrima

last: Das

first: Swagata

last: Adhikary

first: Madavan

last: Vasudevan


title: Homeodomain-containing protein PRRX1 senses DNA double-strand breaks and anchors the Ku heterodimers to promote non-homologous end-joining

city: wuhan

first: zhao

last: chen

state: Hubei

address: shuiguohu

department: BIOLOGY

institute: wuhan university

summary: The DNA-PK complex is essential for non-homologous end-joining (NHEJ) to repair DNA double-strand breaks (DSB) in a template-independent way. The association of Ku70/80 with the DSB ends facilitates the formation of the DNA-PK holoenzyme. Many questions remain regarding the dynamics that DNA-PK is attached and stabilized at broken DNA ends. Here we revealed that PRRX1, a homeodomain containing protein, mediates chromatin localization and subsequent activation of the DNA-PK. PRRX1 oligomerizes to enable its simultaneous associations with both double-strand DNA and the SAP domain of Ku70, thereby enhancing Ku anchoring at DSBs to stabilize DNA-PK for efficient NHEJ repair. Low expression and pathogenic mutations of PRRX1 are associated with genomic instability and defective NHEJ repair. The peptide interfering PRRX1 oligomerization compromises NHEJ and hampers cell survival on irradiation. These findings deepen our understanding of how the NHEJ machinery is activated and have implications for optimizing cancer therapies.

overall_design: E14.5 primary MEFs were generated from wild‑type，Prrx1+/‑ or Prrx1‑/‑ C57BL/6 littermates.Total RNA isolation of MEFs was performed using RNAiso Plus and  converted to cDNA.RNA-seq of MEFs was performed and analyzed in Novogene

first: Yan

last: Wang

first: Fuyuan

last: Shen

first: Chen

last: Zhao

first: Jiali

last: Li

first: Wen

first: Yamu

first: Fangyu

first: Haojian

last: Zhang

first: Xuefeng

last: Chen

first: Qiang

first: Ying

last: Liu

last: Zhou


title: Psychological stress-induced ALKBH5 deficiency promotes tumour innervation and pancreatic cancer progression via extracellular vesicle transfer of m6A-modified RNAs

first: Zhou

last: yifan

state: 周一帆

address: 74 Zhongshan Second Road

institute: Sun Yat-sen University Cancer Center

summary: Pathological role and the mechanism of psychological stress in cancer progression are little known. Here, we show in a mouse model that psychological stress drives pancreatic ductal adenocarcinoma (PDAC) progression via stimulating tumor nerve innervation. We demonstrate that nociception and other stressors activate sympathetic nerve to release Noradrenaline, downregulating tumor cell RNA demethylase alkB homolog 5 (Alkbh5). Alkbh5 deficiency causes cancer cell aberrant m6A modification of RNAs, which are packed to extracellular vesicles and delivered to neurons in the tumor microenvironment, enhancing hyperinnervation and PDAC progression. ALKBH5 levels are reversely correlated with tumor innervation and survival time in PDAC patients. Animal experiments identify a natural flavonoid Fisetin preventing neurons from taking in m6A-RNA contained EVs and suppress PDAC tumor excessive innervation and progression. Together, our study shed light on a novel molecular mechanism for neuro-cancer crosstalk linking psychological stress and cancer progression and raise a potential strategy for PDAC therapy.

overall_design:
We performed RNA sequencing data, proteomics data and assay for transposase-accessible chromatin with sequencing data obtained from isolated PADC cells to elucidate the mechanism for the stress to induce tumor progression. We performed RNA m6A-sequencing to elucidate RNA m6A methylation change in cell RNA and EV RNA after NA treatment or Alkbh5 KO. To evaluate the effects of EV-RNAs on neuronal transcriptional profiles, we performed RNA sequencing of neurons after transfection of EV-RNAs from Alkbh5-KO or control KPC cells. We also separated m6A-modified RNAs and non-m6A-modified RNAs extracted from Alkbh5-KO KPC cell produced EVs by RNA immunoprecipitation and then delivered them into neurons to elucidate the change of neuronal transcriptional profiles.
This dataset includes RNA-seq of isolated tumor cell from orthotopic tumor tissue.

first: Jian

last: Zheng

first: Ziming


title: UBR7 in concert with EZH2 inhibits the TGF-β signalling leading to extracellular matrix remodelling [ChIP-seq]

first: madavan

overall_design: Chromatin immunoprecipitation DNA sequencing (ChIP seq) for histone modifications H3K27me3 in stable cells transfected with scrambled RNA and UBR7-Sh vector.

first: Chandrima

first: Swagata


title: Expression data from murine liver after knocking out Cdkl3

city: Shenyang

first: qiu

middle: zhong

last: pang

address: 195 Chuangxin Road, Baita Street

institute: Northeastern University

summary: We first found that Cyclin-Dependent Kinase Like 3 (CDKL3) plays an important role in Non-Alcoholic Fatty Liver Disease (NAFLD). Using the Cre-Loxp system, we constructed the murine model with a liver-specific Cdkl3 deletion and then comprehensively evaluated the effects of liver Cdkl3 deficiency through high-throughput sequencing.

overall_design: After knocking out Cdkl3 in the liver (C57BL/6), male mice (control and knockout group) were selected and fed with a chow diet for 10 months, followed by sequencing transcriptome profiling of whole livers.

first: Zhongqiu

last: Pang

first: Ren

last: Sheng


title: Dynamic Single Cell Transcriptomics Defines Kidney FGF23/KL Bioactivity and Novel Segment-Specific Inflammatory Targets [RNA-seq]

city: Indianapolis

first: Kenneth

last: White

state: IN

address: 635 Barnhill Drive

institute: Indiana University School of Medicine

summary: FGF23 via its coreceptor αKlotho (KL) provides critical control of phosphate metabolism, which is altered in rare and very common syndromes. However, the spatial-temporal mechanisms dictating renal FGF23 functions remain poorly understood, thus developing approaches to modify specific FGF23-dictated pathways has proven problematic. Herein, wild type mice were injected with rFGF23 for 1, 4 and 12h and renal FGF23 bioactivity was determined at single cell resolution. Computational analysis identified distinct epithelial, endothelial, stromal, and immune cell clusters, with differential expressional analysis uniquely tracking FGF23 bioactivity at each time point. FGF23 actions were sex independent but critically relied upon constitutive KL expression mapped within proximal tubule (S1-S3) and distal tubule (DCT/CNT) cell sub-populations. Temporal KL-dependent FGF23 responses drove unique and transient cellular identities, including genes in key MAPK- and vitamin D-metabolic pathways via early- (AP-1-related) and late-phase (EIF2 signaling) transcriptional regulons. Combining ATACseq/RNAseq data from a cell line stably expressing KL with the in vivo scRNAseq pinpointed genomic accessibility changes in MAPK-dependent genes, including the identification of FGF23-dependent EGR1 distal enhancers. Finally, we isolated unexpected crosstalk between FGF23-mediated MAPK signaling and pro-inflammatory TNF receptor activation via NF-κB, which blocked FGF23 bioactivity in vitro and in vivo. Collectively, our findings have uncovered novel pathways at the single cell level that likely influence FGF23-dependent disease mechanisms.

overall_design: To investigate unbiased FGF23-dependent genomic and transcriptional reprogramming  in a kidney cell line, HEK293 cells transfected with stable membrane-bound KL receptor were treated with human FGF23 protein or vehicle control for 4 hrs, in quadriplicate. These samples were then used for RNAseq.

first: Rafiou

last: Agoro

middle: E


title: Human CAL51 Transcriptome in response to p53 activation

first: Xiang

address: DongAn Road 270

overall_design: Treatment of CAL51 cells with DMSO, 5-FU, DDP or Nutlin

first: Mingming

last: Cao

first: Quan

last: Liao


city: Guangzhou

overall_design:
We performed RNA sequencing data, proteomics data and assay for transposase-accessible chromatin with sequencing data obtained from isolated PADC cells to elucidate the mechanism for the stress to induce tumor progression. We performed RNA m6A-sequencing to elucidate RNA m6A methylation change in cell RNA and EV RNA after NA treatment or Alkbh5 KO. To evaluate the effects of EV-RNAs on neuronal transcriptional profiles, we performed RNA sequencing of neurons after transfection of EV-RNAs from Alkbh5-KO or control KPC cells. We also separated m6A-modified RNAs and non-m6A-modified RNAs extracted from Alkbh5-KO KPC cell produced EVs by RNA immunoprecipitation and then delivered them into neurons to elucidate the change of neuronal transcriptional profiles.
This dataset includes ATAC sequencing of isolated tumor cell from orthotopic tumor tissue.


title: Genome-wide transcriptional silencing and mRNA stabilization allow the coordinated expression of the meiotic program in mice

city: Rennes

first: Frédéric

last: Chalmel

state: France

address: 9 avenue du Pr. Léon Bernard

department: Physiology and physiopathology of the urogenital tract

institute: Inserm U1085-Irset

summary: The transcriptional dynamic of mammalian cells when these transit from the ubiquitous mitotic to a meiotic specific program is key to understand this switch central to sexual reproduction. By quantifying active RNA polymerase II and nascent transcripts using single cell dataset and ethynyl-uridine pool-down with sorted cells from synchronised testes, we detailed the transcriptional activity of murine male germ cells. When spermatogonia differentiate, transcription slows down, reaching minimal activity at meiotic entry and resumes during pachytene stage. This event, we termed EMLT (for early meiotic low transcription), is distinct from the silencing of sex chromosomes as it is independent of Setdb1 though it is accompanied by the same chromatin mark, H3K9me3. EMLT is delayed in Stra8KO but occurs in mutants altering meiotic chromosome structure or DSB formation or repair.

overall_design: By comparing transcript abundance and neotranscription we unveil a massive event of mRNA stabilization that parallels EMLT. Altogether our data indicate that meiosis is initiated with a nearly silent genome and we propose that the stabilization of transcripts at that time facilitates the meiotic entry by synchronising the expression of several meiotic sub-programs.

first: Laura

last: Bellutti

first: Edith

middle: C

last: Sock Pen

first: Gabriel

last: Livera


title: E-cadherin has a cell-type specific role in maintaining the integrity of pulmonary airways [RNA-seq]

first: Sunyoung

last: Lee

overall_design:
We performed bulk RNA sequencing to investigate differentially expressed genes in each airway cell type after Cdh1 inactivation in Club cells.
Immune cells and endothelial cells were excluded by depletion and epithelial cells were selected for by EPCAM expression and then sorted into GFP+ and GFP- populations.

first: Randee

last: Young


title: RNA sequence profile of WT and SAMD4A knockdown in human embryonic stem cell

first: Li

last: Li

address: Zhen Nan Road 500

department: School of Medicine

institute: Tongji University

summary: To identify the function of RBP SAMD4A in hESC self-renewal and lineage specification. We constructed SAMD4A knockdown hESCs. We performed RNA-sequencing (RNA-seq) on WT and SAMD4A knockdown human embryonic stem cells and analyzed the gene expression profiling using the data.

overall_design: To explore the function of RBP SAMD4A in hESC self-renewal and lineage specification. We constructed SAMD4A knockdown hESC line, and analyzed the differentially expressed genes (DEGs) in WT and SAMD4A knockdown hESCs (three replicates for each).

last: Yi


title: Hepatitis B Treatment Impact on lncRNA and Immune Signaling in Maternal-Fetal Tissues

city: Nanning

first: wang

last: Cuimin

address: No. 6, Taoyuan Road, Qingxiu District

institute: Guangxi Academy of Medical Sciences

summary: To assess the impact of anti-hepatitis B virus (HBV) therapy on lncRNA expression and immune signaling pathways in umbilical cord blood and placental tissue, with the goal of identifying therapeutic targets for intrauterine infection prevention.

overall_design: Umbilical cord serum and placental tissue were collected from six HBV carriers. Three pregnant women underwent antiviral therapy, while three served as untreated controls. LncRNA microarray analysis and bioinformatics were used to evaluate therapy effects on lncRNA expression and signaling pathways.

first: Wang


title: A supervised Bayesian method for time (re)annotation of transcriptomics data

city: Helsinki

first: Miia

last: Lindström

address: Agnes Sjöbergin katu 2

department: Faculty of Veterinary Medicine

country: Finland

institute: University of Helsinki

summary: We propose a Bayesian method based on Gaussian process regression modeling to solve the problem. We use it to perform time annotation in old \textit{Clostridium botulinum} microarray experiments by utilising recently collected RNA-Seq time series. Additionally, we apply the method to time annotate RNA-Seq experiments and test its robustness toward noise on synthetically generated data.

overall_design: Time series experiments with wild type and various mutants

first: Elio

last: Nushi

first: François

middle: P

last: Douillard

first: Katja

last: Selby

first: Nigel

last: Minton

first: Antti

last: Honkela


first: Zhou

last: yifan

overall_design: We performed RNA sequencing data, proteomics data and assay for transposase-accessible chromatin with sequencing data obtained from isolated PADC cells to elucidate the mechanism for the stress to induce tumor progression. We performed RNA m6A-sequencing to elucidate RNA m6A methylation change in cell RNA and EV RNA after NA treatment or Alkbh5 KO. To evaluate the effects of EV-RNAs on neuronal transcriptional profiles, we performed RNA sequencing of neurons after transfection of EV-RNAs from Alkbh5-KO or control KPC cells. We also separated m6A-modified RNAs and non-m6A-modified RNAs extracted from Alkbh5-KO KPC cell produced EVs by RNA immunoprecipitation and then delivered them into neurons to elucidate the change of neuronal transcriptional profiles.

first: Jian

last: Zheng

first: Ziming

last: Chen


title: RNA-seq and acRIP-seq in NAT10-knockdown PDAC cells

first: Leyi

last: Huang

state: Guangdong

address: 107 Yan Jiang West Road, Guangzhou, P.R. China

institute: Sun Yat-sen University

summary: Pancreatic cancer is a lethal diease with high tendency of metastasis. Howerver,  the mechanisms of pancreatic cancer are sitill unclear. To explore the roles of N4-acetylation (ac4C) RNA modification and its involved N-Acetyltransferase 10 (NAT10) in pancreatic ductal adenocarcinoma (PDAC), we performed profiling by high throughput sequencing.

overall_design: NAT10 stably knockdown or ctrl PANC-1 cells were used to investigate the different expression genes which were correlated to NAT10.

first: Quanbo

last: Zhou


title: Dynamic Single Cell Transcriptomics Defines Kidney FGF23/KL Bioactivity and Novel Segment-Specific Inflammatory Targets [scRNA-Seq]

first: Kenneth

last: White

summary: Wild type mice were injected with rFGF23 for 1, 4 and 12h and renal FGF23 bioactivity was determined at single cell resolution. UMAP plots identified distinct epithelial, endothelial, stromal, and immune cell clusters, with differential expressional analysis uniquely tracking FGF23 bioactivity at each time point.

overall_design: C57BL/6J male and female mice were purchased from the Jackson Laboratory at 9-weeks of age and housed in the IUSM Laboratory Animal Resource Center (LARC) for 1 week prior to the start of the experiments. At 10-weeks of age, mice were treated with a single intraperitoneal injection of 10 μg/mouse body weight of rhFGF23 (R&D Systems, Inc.) and euthanized at baseline (0h), 1, 4, 12, or 24h. For scRNAseq experiment, male-female kidney pairs were digested, and cells were isolated as previously described.

first: Rafiou

last: Agoro


title: Endogenous viral elements constitute a complementary source of antigens for personalized cancer immunotherapy

city: Hoersholm

first: Christian

last: Garde

address: Dr Neergaards Vej 5F

institute: Evaxion Biotech

summary: We present a pipeline, ObsERV, for the design of personalized cancer immunotherapies based on epitopes derived from endogenous retroviral elements (EVEs). We show that EVE-derived peptides are presented as antigens on tumors and can be predicted by ObsERV. Preclinical testing of ObsERV demonstrates elicitation of poly-functional CD4+ and CD8+ T-cell responses as well as long-term tumor protection and persistence of the responses.

overall_design: Murine tumors (B16F10 and CT26) were grown and harvested for RNA-seq. RNA-seq was leveraged to design EVE based vaccines using the ObsERV pipeline.

first: Thomas

last: Trolle

first: Pablo

last: Garces

first: Michail

last: Pavlidis


title: Root transcriptome of CLE3 overexpressing lines and clv1-15 mutant lines

city: East Lansing

first: Hideki

last: Takahashi

state: Michigan

address: 1066 Bogue Street

department: Biochemistry and Molecular Biology

institute: Michigan State University

summary:
Nitrogen availability significantly impacts the development of plant root system architecture as plants must optimize growth for efficient acquisition of this nutrient. A signaling pathway involving the CLE family of small signaling peptides and the CLV1 leucine-rich repeat receptor kinase has been shown to repress lateral root development when plants are undergoing nitrogen starvation. Targets of this pathway remain unstudied and may be implicated in the coordination of N-responsive root growth. We obtained the clv1-15 mutant in Landsberg erecta (Ler) background and have generated transgenic lines overexpressing CLE3 in Columbia-0 (Col-0) background for further analysis.
To identify downstream components of CLE-CLV1 signaling, we analyzed the transcriptome profiles of CLE3 overexpressing lines and the clv1-15 mutant.

overall_design: Root tissue from Landsberg erecta (Ler) and Columbia-0 (Col-0) wild-type accessions, two CLE3 overexpression lines in Col-0 background, and clv1-15 mutant in Ler background were used for this analysis. Plants were grown for 10 days on MGRL media supplemented with 0.1 mM KNO3. Experiments were performed in triplicate for Ler, Col-0, and clv1-15 and in duplicate for CLE3 overexpression lines. Samples were hybridized to the ATH1 Array (Affymetrix).

first: Katerina

last: Lay-Pruitt

first: Takao

last: Araya

first: Ricardo

last: Giehl

first: Rashed

last: Abualia

first: Eva

last: Benkova

first: Nicolaus

last: von Wiren


title: Impaired protein tyrosine phosphatase delta is a driver of hepatic metabolic syndrome

city: Strasbourg

first: Frank

last: Jühling

address: 3 Rue Koeberlé

institute: Université de Strasbourg, Inserm, Institut de Recherche sur Les Maladies Virales et Hépatiques UMR_S1110

summary:
Background & Aims: The protein tyrosine phosphatase delta (PTPRD) is suppressed in several diseases including cancers and hepatitis C virus (HCV) infection. To identify hepatic pathways responsive to PTPRD function and their role in liver disease, we analyzed liver transcriptomic and clinical data from patients and established a Ptprd-deficient liver disease mouse model.
Methods: Based on liver transcriptomic data, healthy patients were classified according to PTPRD expression and gene set enrichment analysis (GSEA) was performed to identify signaling pathways associated with low PTPRD expression. We established a liver disease mouse model combining impaired PTPRD expression (Ptprd+/-) and high-fat diet to assess its impact on liver transcriptomics and hepatic metabolism. Identified pathways were validated by perturbation studies in primary human hepatocytes. The clinical relevance was validated in a cohort of patients with fatty liver disease by ranking of individuals according to hepatic PTPRD expression prior analyzing the association with metabolic disease markers.
Results: In healthy individuals and Ptprd+/- mice fed with normal diet, low PTPRD levels associated with a gene expression pattern involved in glucose/lipid metabolism in the liver. During diet-induced liver disease, low hepatic PTPRD led to increased blood glucose and hepatic lipid levels in the Ptprd+/- animals. Moreover, Ptprd+/- animals exhibited a pronounced unfolded protein response, which has a potential impact on hepatic insulin signaling. Indeed, silencing of PTPRD in PHH blunted the insulin-induced AKT phosphorylation. Accordingly, obese patients with low hepatic PTPRD expression exhibit increased levels of clinical markers associated with metabolic disease.
Conclusion: Our data suggests an important regulatory role of hepatic PTPRD in maintaining glucose/lipid homeostasis and how its impaired expression is associated with clinical manifestations of metabolic disease.

overall_design: mRNA profiles obtained by RNA-seq from liver tissues of either Ptprd+/+ (n=3) or Ptprd+/- (n=3) C57BL/6J mice.

first: Armando

last: Andres

first: Roca

last: Suarez


title: RNA-seq analyzes the effect of purA on transcriptome in Escherichia coli during ciprofloxacin treatment.

status: Public on Mar 25 2025

city: xiamen

first: Weiwei

last: Zhu

address: 4221-117 South Xiang-An Road

institute: Xiamen University

summary: PurA encodes an adenylosuccinate synthetase that catalyzes the first step in the de novo synthesis of AMP. A deficiency in purA has been found to confer Escherichia coli tolerance to diverse types of antibiotics. To understand the molecular basis of the increased tolerance, we performed RNA-seq analysis to compare the transcriptional profiles of wild-type and ΔpurA mutant strains before and after ciprofloxacin treatment. The data showed that the purA deficiency suppresses transcript levels from NADH:quinone oxidoreductase genes prior to stress exposure, thereby predisposing E. coli to antibiotic tolerance by reducing respiration. During ciprofloxacin exposure, a purA deficiency suppressed a surge in expression of TCA cycle and ATP synthesis genes and the accumulation of intracellular ATP and ROS, thereby conferring bacterial tolerance to diverse antibiotic stresses.

overall_design: Exponentially growing cultures of E. coli K-12 were treated with ciprofloxacin (5 MIC) for 0 min and 90 min. Cells were collected for total RNA extraction and subsequent RNA-seq. Differential expression of various conditions was analyzed.

first: Xilin

last: Zhao


title: Targeting Galectin-3 to modulate inflammation in LAMA2-deficient congenital muscular dystrophy

city: Pittsburgh

first: Yonne

last: Menezes

state: Pennsylvania

address: 4401 Penn Ave

institute: University of Pittsburgh

summary: LAMA2-deficient congenital muscular dystrophy (LAMA2-CMD) is a severe neuromuscular disorder caused by LAMA2 mutations, leading to muscle degeneration, chronic inflammation, and fibrosis. Histopathological assessment of muscle biopsies from LAMA2-CMD patients and mouse models show clear evidence of inflammation, which oftentimes are regarded as one of the typical dystrophic hallmarks. However, the composition of immune cells in the laminin-deficient muscles remain understood. Consequently, targeted pharmacological intervention to reduce inflammation has never been tested. In this study, we characterized the immune landscape in dyW mouse model of LAMA2-CMD using RNA sequencing and flow cytometry. Transcriptomic analysis of dyW quadriceps identified 2,143 differentially expressed genes, with most of the upregulated genes belong to immune-related pathways. Lgals3 (Galectin-3) was significantly upregulated (log₂FC = 4.27, FDR p-value= 9.21x10-88) and identified as a key upstream regulator of the immune-related pathways. In parallel, flow cytometry analysis revealed elevated leukocyte (CD45⁺) infiltration, with macrophages as the predominant cell population. Pro-inflammatory (M1) macrophages were increased, whereas anti-inflammatory (M2) macrophages remained low, indicating persistent inflammation and impaired resolution. Interestingly, Galectin-3+ macrophages were significantly enriched, which strongly suggest that Galectin-3 drives inflammation in LAMA2-CMD. Treatment of dyW mice with TD-139, a Galectin-3 inhibitor, reduced leukocyte infiltration, decreased Galectin-3+ macrophages, and shifted macrophage polarization toward an M2 anti-inflammatory profile. In addition, RNA sequencing of TD-139-treated dyW muscles showed upregulation of muscle repair pathways and downregulation of fibrosis-related genes. These findings establish Galectin-3-expressing macrophages as an important player in LAMA2-CMD pathophysiology. Importantly, it warrants further investigation on the therapeutic potential of TD-139-mediated inhibition of Galectin-3, including long-term preclinical study, in LAMA2-CMD and potentially other dystrophic conditions driven by chronic immune activation.

overall_design: Transcriptome profile by RNA-seq from isolated quadriceps in 5 weeks old dyW (mouse model of LAMA2-CMD) was compared to healthy wildtype control mice

first: Yonne Karoline

last: Tenorio de Menezes

first: Jinseo

last: Lee

first: Marie A

last: Johnson

first: Jia Qi

last: Cheng Zhang

first: Ruvindi N

last: Ranatunga

first: Dwi

last: Kemaladewi


title: The Plasmodium falciparum ApiAP2 transcription factor PfAP2-V controls parasite proliferation and virulence in intra-erythrocytic developmental cycle

first: Yiwei

last: Zhang

address: 120 Dongling Road

institute: Shenyang Agricultural University

summary: Malaria pathogenesis, encompassing parasite invasion, egress, and antigenic variation, relies on the coordinated activity of numerous proteins, yet their molecular regulatory mechanisms remain poorly understood. Here, we define the role of PfAP2-V, a critical AP2 transcription factor in Plasmodium falciparum, during intra-erythrocytic developmental cycle. PfAP2-V displayed two distinct peaks of expression and was critical for parasite proliferation, invasion, and the regulation of virulence-associated genes. Inducible knockdown of PfAP2-V reduced parasitemia by blocking trophozoite development, which was associated with downregulated phosphorylation of virulence-associated proteins. Furthermore, PfAP2-V knockdown reduced PfEMP1 expression, impairing the adhesion of infected red blood cells to endothelial receptors. Genome-wide chromosome conformation capture and chromatin immunoprecipitation sequencing analyses revealed that PfAP2-V knockdown altered chromatin interactions and accessibility, disrupting the regulation of antigenic variant genes. These findings establish PfAP2-V as a key transcriptional regulator at distinct stages of the intra-erythrocytic cycle.

overall_design: The intrinsic mechanism of PfAP2-V regulation in P. falciparum was explored by single-cell RNA sequencing (scRNA-seq) analysis. A single-cell transcriptome profiles of PfAP2-V knockdown strains of P. falciparum before and after 5 mM treatment were constructed.


title: THP-1 infection H37Rv sequencing results

city: chongqing

first: yusheng

last: yang

address: Department of Orthopedic, Southwest Hospital, Army Medical University

institute: 陆军军医大学第一附属医院

summary: High-throughput sequencing was conducted to identify the effector molecules regulated by H37rv.

overall_design: Adherent THP-1 cells were infected with H37Rv at an MOI of 1 for 4 hours, and then washed three times with PBS to remove bacteria.Cultivate for 24 hours, extract RNA for sequencing.

first: Yusheng

last: Yang

first: Jiezhong

last: Deng


title: THP-1 infection H37Rv sequencing results [1,25(OH)2D3 treatment]

institute: 陆军军医大学第一附属医院


title: Single-copy expression of an amyotrophic lateral sclerosis-linked TDP-43 mutation (M337V) in BAC transgenic mice leads to altered stress granule dynamics and progressive motor dysfunction

city: Oxford

first: Kevin

last: Talbot

address: John Radcliffe Hospital, Headley Way

department: Nuffield Department of Clinical Neurosciences

institute: University of Oxford

summary: Mutations in the gene encoding the RNA-binding protein TDP-43 cause amyotrophic lateral sclerosis (ALS), clinically and pathologically indistinguishable from the majority of 'sporadic' cases of ALS, establishing altered TDP-43 function and distribution as a primary mechanism of neurodegeneration. Transgenic mouse models in which TDP-43 is overexpressed only partially recapitulate the key cellular pathology of human ALS, but may also lead to non-specific toxicity. To avoid the potentially confounding effects of overexpression, and to maintain regulated spatio-temporal and cell-specific expression, we generated mice in which an 80 kb genomic fragment containing the intact human TDP-43 locus (either TDP-43WT or TDP-43M337V) and its regulatory regions was integrated into the Rosa26 (Gt(ROSA26)Sor) locus in a single copy. At 3 months of age, TDP-43M337V mice are phenotypically normal but by around 6 months develop progressive motor function deficits associated with loss of neuromuscular junction integrity, leading to a reduced lifespan. RNA sequencing shows that widespread mis-splicing is absent prior to the development of a motor phenotype, though differential expression analysis reveals a distinct transcriptional profile in pre-symptomatic TDP-43M337V spinal cords. Despite the presence of clear motor abnormalities, there was no evidence of TDP-43 cytoplasmic aggregation in vivo at any timepoint. In primary embryonic spinal motor neurons and in embryonic stem cell (ESC)-derived motor neurons, mutant TDP-43 undergoes cytoplasmic mislocalisation, and is associated with altered stress granule assembly and dynamics. Overall, this mouse model provides evidence that ALS may arise through acquired TDP-43 toxicity associated with defective stress granule function. The normal phenotype until 6 months of age can facilitate the study of early pathways underlying ALS.

overall_design: The BAC vectors were retrofitted via the loxP site within the backbone of pCYPAC2 with a vector containing PhiC31 integrase attB sites and a promoterless Neomycin selection cassette. The vectors were then targeted to the Rosa26 (Gt(ROSA26)Sor) locus in embryonic stem cells (ESCs) by PhiC31 integrase-mediated cassette exchange into IDG26.10-3 ESCs, which harbour an PhiC31 attB flanked hygromycin cassette,  pre-integrated at the Rosa26 (Gt(ROSA26)Sor) locus (Chen et al., 2011). G418 resistant ESCs were screened by PCR screening and immunoblotting to 1) confirm correct exchange (integration) at the 5’ and 3’  end using the two primer combination ExPGK3 (5’-CACGCTTCAAAAGCGCACGTCTG- 3’)  and ExNeo2 (5’-GTTGTGCCCAGTCATAGCCGAATAG-3’); allBac-F1 (5’- TTAGGTCCCTCGACCTGCAGG-3’)  and Rosa3HR-R (5’- CGGGAGAAATGGATATGAAGTACTGGGC-3’), respectively and 2) to ensure all regions of the human genomic BAC were present.  Candidate TDP-43WT and TDP-43M337V ESCs were injected into C57BL/6 blastocysts and the resulting chimeric embryos transferred to  recipient pseudopregnant females. Chimeric male founder mice (F1) were subsequently crossed with C57BL/6J female mice to generate two isogenic human TDP-43 transgenic lines, differing only by the presence or absence of the M337V mutation. Mice were again screened by PCR using a battery of primers to confirm correct integration and presence of the BAC vector regions.  Retention of the TDP-43M337V mutation in exon 6 was confirmed by sequencing DNA from ESCs and subsequent chimeric founders. Founder mice were backcrossed to C57BL/6J for a minimum of 3 generations and homozygous and heterozygous transgenic lines were established. To minimise the effects of genetic background, all experimental control animals were age-matched littermates. These TDP-43M337V BAC mice are available from The Jackson Laboratory as strain JAX#029266, https://www.jax.org/strain/029266.  Five mice with M337 transgene, two mice with WT transgene and three nontransgenic C57BL/6 mice were used for this experiment, all female.

first: Talbot

last: Kevin

first: Gordon

last: David

first: Scaber

last: Jakub


title: Proteasomal activation ameliorates neuronal phenotypes linked to FBXO11-deficiency [fly]

city: Erlangen

first: Steffen

last: Uebe

address: Kussmaulallee 4

department: Humangenetisches Institut

country: Germany

institute: Uniklinikum Erlangen

summary: Haploinsufficiency of FBXO11, encoding a ubiquitin ligase complex subunit, is associated with a variable neurodevelopmental disorder. So far, the underlying nervous-system related pathomechanisms are poorly understood, and specific therapies are lacking. Using a combined approach, we established an FBXO11-deficient human stem cell-based neuronal model using CRISPR/CAS9 and a Drosophila model using tissue specific knockdown techniques. We performed transcriptomic analyses on iPSC-derived neurons and molecular phenotyping in both models. RNA-sequencing revealed disrupted transcriptional networks related to processes important for neuronal development, such as differentiation, migration and cell signaling. Consistently, we found that loss of FBXO11 leads to neuronal phenotypes such as impaired neuronal migration and abnormal proliferation/differentiation balance in human cultured neurons and impaired dendritic development and behavior in Drosophila. Interestingly, application of three different proteasome activating substances could alleviate FBXO11-deficiency-associated phenotypes in both human neurons and flies. One of these substances is the long-approved drug Verapamil, opening the possibility of drug repurposing in the future. Our study shows the importance of FBXO11 for neurodevelopment and highlights the reversibility of related phenotypes, opening an avenue for potential development of therapeutic approaches through drug repurposing.

overall_design: RNA-Seq profiling of heads of Drosophila melanogaster with a neuronal knockdown of Fbxo11.

first: Anne

last: Gregor

first: Philipp

last: Kirchner

first: Arif

last: Ekici

first: Christiane

last: Zweier


title: Transcriptome analysis of cardiac tissues from mice subjected to SD model with oxygen therapy

city: Shanghai

first: Yunkai

address: 800 Xiangyin Road

institute: Naval Medical University

summary: Adequate sleep is essential for relieving stress and rejuvenating the mind; however, undesirable physiological and pathological responses resulting from sleep insufficiency or sleep deprivation (SD) are becoming increasingly common. However, the influence of sleep deficiency on gut microbiota and microbiota-associated human diseases, especially on cardiac diseases remain controversial. Here, we constructed the experimental SD model in mice and found it significantly resulted in weakness, depression-like behaviors, and multiple organs dysfunction. Intriguingly, SD mice developed pathogenic cardiac hypertrophy and fibrosis with poor ejection fraction as well as fractional shortening. 16s rRNA sequencing demonstrated that SD-induced the pathogenic effects of gut microbiota, which was also observed in mice received by fecal microbe from SD mice in fecal microbiota transplantation (FMT) assays. Next, we investigated the therapeutic effects and underlying mechanisms of oxygen therapy in gut microbiota-associated cardiac fibrosis and dysfunction. The environment of 30% oxygen concentration effectively ameliorated the pathological effects on cardiac function. Transcriptome data also found oxygen therapy targeted several hypoxia-dependent pathways and suppressed cardiac collagen production. In conclusion, these results indicated the importance of sufficient sleep in gut microbiota and may represent a potential therapeutic strategy of oxygen environment exerts protective effects in sleepless sufferings through gut microbiota.

overall_design: Sleep deprivation was conducted using a water tank-based sleep deprivation (SD) equipment, involving 18 hours of deprivation per day for 4 weeks. The starting light time (ZT0) was consistently set at 07:00, regardless of the photoperiod length, and the sleep deprivation period was scheduled from ZT7 (14:00) to the following ZT1 (08:00). During the experiment, after the mice completed sleep deprivation at ZT1, thet were placed in an oxygen chamber. 30% oxygen concentrations were adjusted using the contronlling panel, and the mice were treated for 2 hrs. At the end of the treatment, the mice were returned to their cages for rest and subsequently subjected to sleep deprivation at ZT7 for 18 hours. The blank control group did not receive any additional interventions and was maintained under normal conditions.

first: Xiaomeng

last: Ren

first: Shuqi

last: Cai


title: Characterizing extracellular RNA secreted by Caco-2 cells

city: Davis

first: Patricia

last: Baldrich Gonzalez

address: 451 Health Sciences Dr., Davis, CA 95616

department: Genome Center

institute: University of California - Davis

summary: All cells secrete extracellular RNA (exRNA), however the roles of secreted RNAs are still not fully understood. Here, we are profiling the small RNAs secreted from Caco-2 cells. ExRNAs were isolated as different fractions: total extracellular, intravesicular, extravesicular, and nonvesicular (abitrarily named as Total ExRNA P40, P100, S100).

overall_design: Small RNA seq profiling of extracellular RNA isolated from Caco-2 cells

first: Glen

last: Borchert

first: Sunita

last: Paudel

last: Baldrich


title: Microwave-Induced Tumor-Derived Microparticles Treat Lung Adenocarcinoma by Inducing Immunogenic Cell Death via HMGB1 Shuttling [RNA-Seq]

city: Wuhan

first: Xinghui

last: Cao

address: Hangkong Road 13, Wuhan, China

institute: Huazhong University of Science and Technology

summary: Tumor-derived microparticles (TMPs), a subtype of extracellular vesicles, hold great promise in tumor immunotherapies and vaccines, and have demonstrated significant potential as drug delivery systems for clinical tumor treatment. In our previous study, we utilized TMPs released from parental tumor cells upon ultraviolet irradiation (UV-TMPs) to deliver methotrexate (MTX), effectively managing malignant pleural effusion (MPE) in advanced lung cancer cases. However, concerns remain regarding the potential risks associated with nucleic acid incorporation, alongside the limited antitumor efficacy of UV-TMPs alone. Here, we introduce a microwave (MW)-assisted method for preparing TMPs, termed MW-TMPs. Brief exposure to short-wavelength MW radiation triggered various forms of programmed cell death, including apoptosis, necroptosis, and pyroptosis, promoting the release of TMPs. These resulting MW-TMPs contained fewer nucleic acids and exhibited superior antitumor activity in vivo compared to UV-TMPs. Mechanistically, MW-TMPs induced immunogenic cell death (ICD) by shuttling HMGB1 and enabled dual targeting of tumor cells by natural killer (NK) and T cells, while reprogramming of suppressive tumor immune microenvironments in LLC lung adenocarcinoma (LUAD) mouse models. Furthermore, similar effective tumor inhibition was observed in clinical MPE samples and zebrafish PDX models, suggesting the potential of MW-TMPs for clinical cancer immunotherapy. In further application studies, MW-TMPs successfully encapsulated the chemotherapeutic drug MTX, demonstrating enhanced antitumor efficacy both in vitro and in vivo, thereby establishing their potential as an innovative drug delivery platform. Moreover, combination immunotherapy with PD-L1 blockade confirmed a synergistic antitumor effect. This MW-TMP development strategy has improved upon traditional UV-TMP methods in terms of simplicity, efficiency, safety, and efficacy.

overall_design: LLC cells stimulated with UV or MW for 24 hours were collected.

first: Yang

last: Jin

first: Mengfei

last: Guo

first: Yali

last: Wu

first: Wenjuan

last: Chen

first: Jingjing


title: The atypical KRASQ22K mutation directs TGF-β response towards partial epithelial-to-mesenchymal transition in patient-derived colorectal cancer tumoroids

city: Vienna

first: Gerda

last: Egger

address: Waehringer Guertel 18-20

department: Pathology

country: Austria

institute: Medical University of Vienna

summary: Transforming growth factor beta (TGF-β) exhibits complex and context-dependent cellular responses. While it mostly induces tumor-suppressive effects in early stages of tumorigenesis, its tumor promoting properties are evident in advanced disease. This TGF-β duality is still not fully understood, and whether TGF-β supports invasion and metastasis by influencing cancer cells directly, or rather through the stromal tumor compartment remains a matter of debate. Here, we utilized a library of colorectal cancer (CRC) patient-derived tumoroids (PDTs), representing a spectrum of tumor stages, to study cancer cell-specific responses to TGF-β. Using medium conditions allowing for the differentiation of PDTs, we observed TGF-β induced tumor-suppressive effects in early-stage tumoroids. PDTs with TGF-β pathway mutations or PDTs derived from metastatic tumors were insensitive to the treatment. Notably, one tumoroid line harboring an atypical KRASQ22K mutation underwent partial epithelial-to-mesenchymal transition (EMT), associated with morphological changes and increased invasiveness. On a molecular level, this was accompanied by elevated expression of mesenchymal genes, as well as deregulation of pathways associated with matrix remodeling and cell adhesion. Our results suggest that tumor cell intrinsic responses to TGF-β are critical in determining its tumor-suppressive or -promoting effects.

overall_design: To detect which effect TGF-β1 has on the gene expression of colorectal cancer (CRC) patient-derived tumoroids (PDTs), we treated PDTs from one line for 10 days with solvent or TGF-β1 in different medium compositions (n=3) followed by mRNA sequencing. Differentially expressed genes have been extracted from follwoing comparisions ET versus ES, BS versus ES, BT versus ES, BS versus ET, BT versus ET and BT versus BS. ES= ENAS + solvent; ET= ENAS + 5ng/mL TGFbeta; BS=Basal + EGF +solvent; BT= Basal +EGF + 5ng/mL TGFbeta.

first: Theresia

last: Mair

first: Loan

last: Tran

first: Carlos

middle: U

last: Pérez M

first: Raheleh

last: Sheibani-Tezerji


title: Proteasomal activation ameliorates neuronal phenotypes linked to FBXO11-deficiency [human]

first: Steffen

last: Uebe

overall_design: RNA-Seq profiling of hIPSC-derived neurons with FBXO11 loss-of-function

first: Anne

last: Gregor

first: Philipp

last: Kirchner

first: Arif

last: Ekici

first: Christiane

last: Zweier


title: Nuclear receptor 4A1 is Critical for Neutrophil-Dependent Pulmonary Immunity to Klebsiella pneumoniae Infection

first: Brian

last: Campfield

state: PA

institute: University of Pittsburgh School of Medicine

summary: Bacterial pneumonia is a burdensome, costly disease and increasingly challenging to treat due to antibiotic resistance. Complex host-pathogen interactions regulate protective immunity, and immune-directed therapies could yield novel treatments. Neutrophils play a central role in pulmonary bacterial immunity, and mechanistic understanding of neutrophil antibacterial functions in pneumonia has potential clinical and fundamental application. Nuclear receptor 4a1 (Nr4a1), a member of the nuclear orphan receptor family, has been described to regulate inflammation and immune development in a cell type-specific manner, but its role in pulmonary host defense is not well understood. This study demonstrates that Nr4a1 is essential for protection against K. pneumoniae as Nr4a1 -/-mice showed increased lung bacterial burden, dissemination, and reduced survival. Notably, neutrophil-specific Nr4a1 expression was critical for defense against K. pneumoniae infection by regulating the neutrophil transcriptome. These findings suggest targeting neutrophil-specific Nr4a1 may be beneficial for bacterial pneumonia treatment.

overall_design: Norie, Sugitani

first: Matthew

last: Henkel

first: Taylor

last: Eddens

middle: T


title: Chemokine (C-C motif) Ligand 2 Expressing Adventitial Fibroblast Expansion During Loeys-Dietz Syndrome Aortic Aneurysm Formation [8wk mouse]

city: Palo Alto

first: Alex

last: Dalal

state: California

address: CT Surgery, 870 Quarry Rd Ext

department: Cardiothoracic Surgery

institute: Stanford University

summary: Loeys-Dietz Syndrome (LDS), caused by a spectrum of mutations in the TGF-β signaling cascade, leads to aggressive thoracic aneurysms. Given the ubiquitous nature of TGF-β signaling machinery, the exact cell type(s) responsible for initiation of LDS pathogenesis is poorly understood.

overall_design: Aortic root and ascending aortic tissue from 8-week-old Tgfbr2G357W/+ (Loeys-Dietz Syndrome) and WT littler mate control mice was subjected to tissue digestion and single-cell RNA sequencing. Multiple mice were pooled into each sample, yielding 2 samples total.

first: A

first: M

last: Fischbein


title: Single-cell RNA sequencing analysis of baicalein’s immunomodulatory effects in lung graft ischemia-reperfusion injury

city: Chengdu

first: Sen-Lin

last: Hou

address: West China Hospital, Sichuan University, 37 Guoxue Alley, Chengdu

department: Institute of Thoracic Oncology

institute: West China Hospital

summary: This dataset encompasses single-cell RNA sequencing data from lung graft samples subjected to ischemia-reperfusion injury (IRI) with and without baicalein treatment, as well as sham-operated controls. A total of 105,337 cells were initially obtained across nine samples, followed by rigorous quality control measures that yielded 83,590 high-quality cells for downstream analysis. The analysis identified 14 classical immune cell types and 36 subpopulations, highlighting significant differences in neutrophil composition and gene expression between baicalein-treated (BI) and untreated IRI (IR) groups. Specifically, the BI group exhibited a marked reduction in neutrophil abundance and a lower activation level of the AGE-RAGE signaling pathway. Differentially expressed gene analysis in neutrophils identified 263 upregulated and 337 downregulated genes in the BI group compared to the IR group, with functional enrichment analysis revealing pathways associated with immune regulation and cellular stress responses. These findings provide valuable insights into the immunomodulatory effects of baicalein during lung graft IRI and offer a resource for further investigation into therapeutic mechanisms.

overall_design: To investigate the immunomodulatory effects of baicalein on lung graft IRI, a single-cell RNA sequencing experiment was conducted using lung graft samples. The experiment involved three groups: a BI group, an untreated IR group, and a Sham group, each comprising three biological replicates. Utilizing the orthotopic left LTx model in rats, lung tissue specimens from the transplanted left lung were procured. Single-cell suspensions were prepared from the lung graft tissues, followed by library preparation and sequencing using 10x Genomics technology. After rigorous quality control with Seurat and DoubleFinder, 83,590 high-quality cells were retained for analysis. Cells were clustered into 14 classical immune cell types and further subdivided into 36 subpopulations. Neutrophils showed the most pronounced differences between the BI and IR groups, with significant reductions in abundance and differential gene expression patterns in the BI group. The data provide a comprehensive view of immune cell dynamics and the impact of baicalein on neutrophil-related signaling pathways, such as the AGE-RAGE pathway, during lung graft IRI.

first: Dong

last: Tian

first: Xiang-Yun

last: Zheng

first: Zeng-Wei

last: Yu

first: Ye

last: Wu

first: Pei-Zhi

last: Liu

first: Lin-Xi

first: Yu-Xuan

first: Yang

last: Zhao

last: Li

first: Hong-Tao

last: Tang

first: Wei-Yang

first: Ya-Ling

first: Chuan-Fen

last: Zhang

first: Yun

last: Wang

first: Hong-Ying

last: Wen

first: Qiang

last: Pu

first: Masaaki

last: Sato

first: Lun-Xu


title: RNA-seq of degranulating and non-degranulating NK cells under inhibitory and stimulatory treatment

city: Biberach

first: Katrin

last: Fundel-Clemens

address: Birkendorfer 65

institute: Boehringer Ingelheim

summary: Cytotoxic cells (T CD8+ cells and NK cells) are the primary executors of tumour cell killing. However, not all cells are equally prone to kill. Within circulating lymphocytes, certain gene expression patterns may better prepare immune cells to attack.Upon co-culture with tumour cells, cytotoxic cells will undergo a process known as degranulation, which is based on the release of lysosomal-like structures, cytotoxic granules, that contain proteins specialized on disturbing tumour cell membrane integrity. This results in the killing of tumour cells. Cytolytic granules are decorated with lysosomal proteins such as the lysosome-associated membrane proteins (LAMP) family members, which in basal conditions are restricted to lysosomal compartments. Upon fusion of cytolytic granules with the cell membrane, such proteins become transitorily enriched and available to antibodies. This can be used as a tool to distinguish and sort out cell populations that have been recently activated.In our project, we use LAMP1 as a marker of NK cell activation upon challenge with tumour cells. We make use of this strategy to select which cells are better equipped to get activated, and what are the underlying transcriptional signatures involved in this phenotype. In this project we aim to identify intrinsic modulators of cytotoxic cell activation. To this end, we have the following objectives:- Definition of the NK cell activation priming signature in basal conditions.- Comparison of transcriptional signatures following treatment with stimulatory/inhibitory cytokines and the combination of both.- Uncovering new potential targets for immune therapy. - Analysis of the resulting data from the RNAseq to define the transcriptional signature of activated cells, perform gene set enrichment analysis, identification of enriched pathways and getting insight on the ups-tream regulators of the gene signatures.- Confirmation of hits in primary cells and generation of genetically modified NK cell lines/inhibitors/activators to challenge protein function.- Mechanistic study of protein hits.- TCGA study for relevance in cancer indications.- (If established) Parallel NGS study with T CD8+ cells.- Writing a manuscript containing main findings for the NK cells hits, and potentially for the T CD8+ cell study.

overall_design:
Natural killer cells isolated from 6 different healthy donors were each co-cultured with tumor cells to induce degranulation either or not in the prescence of TGFbeta, IL2, or both. Subsequently, NK cells were sorted based on NK cell (CD56, CD3e CD45) and degranulation (LAMP1) markers and processed for RNA sequencing. The total number of samples equals 6 donors * 4 treatments * 2 LAMP1 statuses = 48.

***************************************************************
Raw files for human/patient samples were not submitted to GEO due to concerns about submitting personally identifiable sequence data for open access.
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first: Marta

last: Puig-Gámez

first: Alec

last: Dick

first: John

last: Park


title: Insights into the single-cell transcriptome characteristics of porcine endometrium with embryo loss

city: Qingda

first: yadan

last: Jin

state: 山东Shandong

address: Qingdao

institute: Qingdao Agricultural University

summary: Reproductive disorders are a concern in the pig industry.  Successful gestation processes are closely related to a suitable endometrial microenvironment, and the physiological mechanisms leading to failed pregnancy during the peri-implantation period remain unclear.  We constructed single-cell transcriptome profiles of peri-implantation embryo loss and successful gestation endometrial tissues, and identified 22 cell subpopulations, with epithelial and stromal cells being the predominant endometrial cell types.  The two tissues showed marked differences in cell type composition, especially among epithelial cell subpopulations.Overall, the present study revealed the molecular features of endometrial cell transcription in embryo-lost endometrial tissues, providing deeper insights into the endometrial microenvironment of reproductive disorders, which may inform the etiological, diagnostic, and therapeutic studies of reproductive disorders.

overall_design: All experimental animals were three years old with a similar genetic background.  We selected a Yorkshire sow with an abortion experience and subsequent unsuccessful breeding with three consecutive artificial insemination matings that represented the sample of embryo loss (EL).  Another Yorkshire sow with successful artificial insemination was selected and gestated for 30 days, representing the sample of successful embryo implantation (EIS).  We defined the day of artificial insemination as day 0 of pregnancy and endometrial samples were collected from each uterine horn


title: Factors Driving Diversity in Gene Regulatory Networks at Genome Scale[ASE]

city: Cambridge

first: Jie

last: Yun

state: MA

address: 77 MASS

department: CEE

institute: MIT

summary: Gene expression is a quantitative trait under the control of genetic and environmental factors and their interaction, so-called GxE. Understanding the mechanisms driving GxE is fundamental for ensuring stable crop performance across environments, and for predicting the response of natural populations to climate change. Gene expression is regulated through complex molecular networks, however environmental and genotypic effects on genome-wide regulatory networks are rarely considered. In this study, we model genome-scale gene expression variation between two natural accessions of the model grass Brachypodium distachyon and their response to soil drying. We identified genotypic, environmental, and GxE responses in physiological, metabolic, and gene expression traits. We then identified gene regulation conservation and variation among conditions and genotypes, simplified as co-expression clusters found unique in or conserved across library types. Putative gene regulatory interactions are inferred as network edges with a graphical model approach, resulting in hypotheses about gene-gene interactions which are then found to be specific to or with higher affinity in one genotype (G regulation), one environmental treatment (E regulation), or in one genotype under treatment (GxE regulation). Some gene-gene interactions are conserved across conditions so the differential expression is accordingly transmitted to target genes. These variably detected edges cluster together in co-expression modules, suggestive of different constraints or selection strength acting on specific pathways. We further applied our graphical model approach to identify putative, E-dependent regulatory mechanisms of leaf glucose content as an exemplar metabolite. Our study highlights an approach to identify variable features of gene regulatory networks and thereby identify key components for later genomic intervention to elucidate function or modulate environmental response. Our results also suggest possible targets of evolutionary change in gene regulatory networks associated with environmental plasticity.

overall_design:
To study regulation mechanism of how different genotypes of Brachypodium (Bd21 and Bd3-1) response to soil water deficit differently, we generated hybrid of the two genotypes and applied same treatments and harvest samples for RNA sequencing. We started gradual dry-down on 33rd day with a soil water content ended up 55% for drought and 85% for control after 6 days. The youngest fully expanded leaves from 5 replicates of hybrids under drought and control treatment, and 2 control Bd3-1 samples were harvested at a single time point 1-2:30pm of the day for RNA sequencing. These along with 12 samples (each genotype and condition, replicated three times) from the main experiment with exact same treatments are sequenced.
We did gene expression profiling analysis and differential expression by fitting models using DESeq2 to infer gene regulation mechanism.

Grant: IOS 2239070
Grant title: NSF CAREER Award
Grantee: David Des Marais
Funding agency: National Science Foundation

first: Angela

last: Burnett

first: Alistair

last: Rogers

first: David

last: Des Marais


title: Identification and Functional Characterization of lncRNAs involved in Human Monocyte-to-Macrophage Differentiation

city: Santa Cruz

first: Susan

last: Carpenter

address: Red Hill Rd, Biomedical Sciences

department: MCD

institute: UC Santa Cruz

summary: Long noncoding RNAs (lncRNAs) make up the largest portion of RNA produced from the human genome, but only a small fraction have any ascribed functions. Although the role of protein-coding genes in macrophage biology has been studied extensively, our understanding of the role played by lncRNAs in this context is still in its early stages. There are over 20,000 lncRNAs in the human genome therefore, attempting to select a lncRNA to characterize functionally can be a challenge. Here we describe two approaches to identify and functionally characterize lncRNAs involved in monocyte-to-macrophage differentiation. The first involves the use of RNA-seq to infer possible functions and the second involves a high throughput functional screen. We examine the advantages and disadvantages of these methodologies and the emergence of novel research and technologies aiding us in addressing these challenges.

overall_design: For our candidate RNA-sequencing experiments we utilized our THP1-NFKB-dCas9 cell line and created two negative controls cell lines: a non-targeting sgRNA and an anti-GFP sgRNA.  We designed 3 sgRNAs and created three CRISPRi-LincJADE1 cell lines and created three CRISPRi-LincANXA3 cell lines. We cloned the top 2 performing guides from our THP1 differentiation screen (10.1073/pnas.2322524121) and created 2 CRISPRi-GATA2AS1 cell lines and 2 CRISPRi-PPP2R5C-AS1 cell lines. RNA samples were collected at 0hrs and 24hrs after being treated with 100nM of PMA.

first: Christy

last: Montano

first: Eric

last: Malekos

first: Sergio

last: Covarrubias


title: Factors Driving Diversity in Gene Regulatory Networks at Genome Scale[main]

overall_design:
To study how different genotypes of Brachypodium response to soil water deficit differently, we include two genotypes, i.e., Bd21 and Bd3-1, and two environment, i.e., drought and control samples. We started gradual dry-down on 33rd day after germination with a soil water content ended up 55% for drought and 85% for control after 6 days. The youngest fully expanded leaves from 48-59 replicated plants were harvested at a single time point 1-2:30pm of the day for RNA sequencing.
We did gene expression profiling analysis and differential expression among library types, especially interaction effect. The gene regulation network variation among library types are characterized based on gene expression clustering and its variation, infered gene regulations and its variation which is based on slope variation of multiple regression of target gene on its regulators.
Replicates are used for differential gene expression study, and variations among replicates are utilized for co-expression cluster, gene regulation inference, and multiple regression of gene expression.

Grant: IOS 2239070
Grant title: NSF CAREER Award
Grantee: David Des Marais
Funding agency: National Science Foundation


title: Systematic genetic perturbation reveals principles underpinning robustness of the epigenetic regulatory network

city: London

first: Paola

last: Scaffidi

address: 1 Midland Road

institute: Francis Crick Institute

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

overall_design: Refer to individual Series