title: Txnip deficiency causes a susceptibility to acute cold stress with brown fat dysfunction in mice [RNA-seq]

summary: Mammals adaptively regulate energy metabolism in response to environmental changes such as starvation and cold circumstances. Thioredoxin-interacting protein (Txnip), known as a redox regulator, serves as a nutrient sensor regulating energy homeostasis. Txnip is essential for mice to adapt to starvation, but its role in adapting to cold circumstances remains unclear. Here, we identified Txnip as a pivotal factor for maintaining non-shivering thermogenesis in mice. Txnip protein levels in brown adipose tissue (BAT) were upregulated by the acute cold exposure. Txnip-deficient (Txnip-/-) mice acclimated to thermoneutrality (30°C) exhibited significant BAT enlargement and triglyceride accumulation with downregulation of BAT signature and metabolic gene expression. Upon acute cold exposure (5°C), Txnip-/- mice showed a rapid decline in BAT surface temperatures with the failure of increasing metabolic respiration, developing lethal hypothermia. The BAT dysfunction and cold susceptibility in Txnip-/- mice were corrected by acclimation to 16°C, protecting the mice from life-threatening hypothermia. Transcriptomic and metabolomic analysis using dissected BAT revealed that despite preserving glycolysis, the BAT of Txnip-/- mice failed to activate the catabolism of branched-chain amino acids and fatty acids in response to acute cold stress. These findings illustrate that Txnip is required for maintaining basal BAT function and ensuring cold-induced thermogenesis.

overall_design: To explore the physiological role of Txnip in brown adipose tissue, Txnip-deficient (Txnip-/-) and wild tylpe (WT) mice were housed at room temperature (RT, 23°C) or thermoneutrality (TN, 30°C), and the interscapular BAT was dissected for gene expression profiling analysis using RNA-seq

first: Meng

last: Zou

first: Kikuko

last: Amo-Shiinoki

first: Akihiko

last: Taguchi


title: In vivo CRISPR screen reveals interplay of epigenetic regulators underlies kinetics of hepatobiliary reprogramming [CUT&Tag]

status: Public on Jan 01 2025

city: Tokyo

first: Takeshi

last: Katsuda

address: 7-chōme-3-1 Hongō

department: Department of Chemical System Engineering

country: Japan

institute: University of Tokyo

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Under chronic liver injury, a small fraction of hepatocytes undergoes reprogramming to biliary epithelial cells (BECs) through a multistep process of chromatin and transcriptional remodeling in which the hepatocyte phenotype is repressed and the biliary phenotype is activated. However, the epigenetic basis underlying this phenomenon is largely unknown. Here, using a lineage-traced model of cholestatic liver injury, we investigated the role of histone posttranslational modification in the kinetics of biliary reprogramming. In this study, we performed CUT&Tag to profile the changes in histone landscape throughout reprogramming and explored the impact of Nsd1 and Kdm2a knockout.

overall_design: Mice were challenged with DDC for 8 weeks, then reprogrammed cells and biliary cells were harvested. After DDC-induced cholestatic injury, GFP+CD24-EPCAM- cells (DDC-injured hepatocytes/early reprogrammed cells), GFP+CD24+EPCAM- cells (intermediate reprogrammed cells) and GFP+CD24+EPCAM+ cells (late reprogrammed cells) were isolated, along with noninjured hepatocytes and BECs from Kdm2a-KO, Nsd1-KO, or empty vector (EV) control mice.

first: Jonathan

middle: H

last: Sussman

first: Ben

middle: Z

last: Stanger


title: In vivo CRISPR screen reveals interplay of epigenetic regulators underlies kinetics of hepatobiliary reprogramming [RNA-Seq]

type: Expression profiling by high throughput sequencing

summary: Under chronic liver injury, a small fraction of hepatocytes undergoes reprogramming to biliary epithelial cells (BECs) through a multistep process of chromatin and transcriptional remodeling in which the hepatocyte phenotype is repressed and the biliary phenotype is activated. However, the epigenetic basis underlying this phenomenon is largely unknown. Here, we profiled cells after knockout of Nsd1 or Kdm2a at two reprogramming stages to assess the transcriptional differences imposed by epigenetic dysregulation.

overall_design: Mice were challenged with DDC for 8 weeks, then reprogrammed cells and biliary cells were harvested. After DDC-induced cholestatic injury,  GFP+CD24- cells (DDC-injured hepatocytes/early reprogrammed cells) and GFP+CD24+ cells were isolated, including the intermediate and late stages of reprogrammed hepatocytes, from Kdm2a-KO, Nsd1-KO, or empty vector (EV) control mice.


title: Quantitative profiling of differentially expressed miRNAs in exosomes of siCTL- and siPRDX3-transfected HCT 116 cells

city: Seoul

first: GooTaeg

last: Oh

address: 52, Ewhayeodae-gil

department: Department of Life Sciences

country: South Korea

institute: Ewha Womans University

type: Non-coding RNA profiling by high throughput sequencing

summary: Purpose: The goal of this study is to compare exosomal small RNA transcriptome of HCT116 cells to identify the target of PRDX3 under basal or knock down conditions by utilizing miRNA-seq. Methods: miRNA profilies of siCTL or siPRDX3 transfected HCT 116 exosoems were generated by illumina sequencing method, in triplicate. After sequencing, the raw sequence reads are filtered based on quality.Sequence reads were mapped with the bowtie2 software tool, which yielded bam files. Mature miRNA sequences were used as references for mapping. Read counts mapped to a mature miRNA sequence were extracted from the alignment file using bedtools v2.25.0 and Bioconductor, which use the R statistical programming language. Read counts were used to determine the expression level of miRNAs. The CPM+TMM normalization method was used for between-sample comparison. Results: We identified known miRNA in species (miRDeep2) in the HCT116 exosome transfected with siCTL or siPRDX3. The expression profile of mature miRNA is used to analyze differentially expressed miRNA(DE miRNA). Conclusions: Our study represents the first analysis of HCT116 exosomal miRNA profiles affected by PRDX3 knockdown with biologic replicates.

overall_design: The HCT-116 cells were cultured in RPMI 1640 supplemented with 10% fetal bovine serum at 37 °C in 5% CO2 saturated humidity. The PRDX3 siRNA, control siRNA were transfected using a NEON microporation system for 40hours.To isolate exosomes, cells were placed in RPMI 1640 with 10% nanoparticle-free FBS and incubated for 24 h, and exosomes were isolated using an ExoLutE exosome isolation kit according to the manufacturer’s instructions. Endothelial micro-RNA profiles of siCTL- and siOASL-transfected HUVECs were generated by miRNA-seq.


title: Effect of CRISPR activation of Zbtb7b, Vps72, Gba, or Mrpl9 in liver tumorigenesis in mice

city: Rochester

first: Kirk

last: Wangensteen

state: MN

address: GU1034H, 222 3rd Ave SW

department: Gastroenterology & Hepatology

country: USA

institute: Mayo Clinic

summary: Hepatocellular carcinomas (HCCs) undergo chromosomal amplifications with resultant increases in gene expression levels, yet the impact of these changes remains largely unknown. We sought to identify cancer driver genes at these genetic loci using CRISPR activation (CRISPRa) screening in mouse livers. We examined The Cancer Genome Atlas data for copy number-amplified and upregulated genes in HCCs. We then conducted in vivo CRISPRa tumorigenesis screens targeting the top 51 genes using dCas9 knock-in mice. Genes Zbtb7b, Vps72, Gba, and Mrpl9 emerged as putative drivers of liver tumorigenesis. Furthermore, high levels of expression of VPS72, GBA, or MRPL9, but not ZBTB7B, correlated with worse survival of HCC patients from the The Cancer Genome Atlas. For validation of the role of these genes in driving tumorigenesis, mice were injected with single CRISPRa plasmids containing Vps72-, Gba-, or Mrpl9-targeting gRNAs. This led to decreased survival and extensive liver tumorigenesis compared to non-targeting controls. We performed RNA sequencing of these tumors to understand the transcriptomic changes caused by activation of these genes.

overall_design:
To understand how activation of Vps72, Gba, Mrpl9, and Zbtb7b could drive tumorigenesis, we injected dCas9-expressing mice with a single plasmid containing (1) Myc to drive baseline hyperproliferation, (2) transcriptional activator sequences for target gene activation, and (3) a single gRNA targeting Zbtb7b (sg-Zbtb7b), Vps72 (sg-Vps72), Gba (sg-Gba), Mrpl9 (sg-Mrpl9), or non-targeting control (sg-NT).
We then performed RNA sequencing of liver tumors from dCas9-expressing Albino (Alb) mice injected with sg-NT (N=5) or sg-Vps72 (N=5) or dCas9-expressing Black (Bl) mice injected with sg-NT (N=6), sg-Zbtb7b (N=5), sg-Gba (N=5), or sg-Mrpl9 (N=5).

first: Alexandra

middle: M

last: Vazquez Salgado

middle: J


title: Txnip deficiency causes a susceptibility to acute cold stress with brown fat dysfunction in mice [ChIP-seq]

city: Ube

first: Katsuya

last: Tanabe

state: Yamaguchi

address: 1-1-1 Minamikogushi

institute: Yamaguchi University

overall_design: ChIP-seq studies of H3K27ac were performed in interscapular BAT from wild-type (WT) and Txnip-deficient (Txnip-/-) mice acclimated to thermoneutrality (TN, 30°C).


title: Methylomic and transcriptomic differences in treatment-naïve profiles among black and non-Hispanic white (NHW) women

city: Chicago

first: Daniela

middle: Elena

last: Matei

state: IL

address: 303 E Superior St

department: Obstetrics and Gynecology

institute: Northwestern University

type: Methylation profiling by genome tiling array

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

overall_design: Refer to individual Series


title: Single cell RNA sequencing data of embryonal carcinoma cells in vitro differentiated into yolk-sac tumor-like cells

city: Düsseldorf

first: Daniel

last: Nettersheim

state: NRW

address: Moorenstr. 5

department: Dep. of Urology

country: Germany

institute: Medical Faculty and University Hospital Düsseldorf, Heinrich Heine University

summary: Single cell RNA sequencing of embryonal carcinoma cell lines 2102EP-lenti-MPHv2, GCT27-lentiMPHv2 and NCCIT-lenti-MPHv2 cells after yolk-sac tumor-like differentiation in vitro.

overall_design: Stable MPHv2+ embryonal carcinoma cells were genereted by infection with lentiviruses (Lenti-MPHv2 plasmind and a hygromycin resistance cassette). This hygromycin resistance was used to select the Lenti-MPHv2+ cells. All three embryonal carcinoma cell lines (2102EP-, GCT27-,NCCIT-lenti-MPHv2) were differentiated by inducing endogenous SOX17 expression (by SOX17-lenti-SAMv2-lentiviruses) and treatment with recombinant Activin-A (100 ng/ml), CHIRON (3 µM), recombinant FGF2 (25 ng/ml) and recombinant Heparin (1 µg/ml). The treatment with the recombinant protein mixture was repeated every two days for eight days in total. Then, cells were isolated and three replicates of each cell line were pooled and sequenced.


title: Methylomic and transcriptomic differences in treatment-naïve profiles among black and non-Hispanic white (NHW) women [Methylation]

summary: Epithelial ovarian cancer (EOC) is a highly lethal gynecological malignancy, with the high-grade serous ovarian cancer subtype (HGSOC) accounting for a significant proportion of cases. Despite platinum-based chemotherapies being the primary treatment for EOC, HGSOC displays high resistance to these therapies. Black women have the highest mortality/incidence ratio of all ethnic groups, and recent studies suggest significant genetic and epigenetic differences between black and non-Hispanic white women who are treatment naïve. This study aims to explore immunogenic differences and hypermethylation phenotype in black women that may contribute to poorer outcomes within this patient group.  We collected primary tumor samples from 36 black and 31 non-Hispanic white (NHW) treatment-naïve patients via biopsy and performed methylomic and transcriptomic analyses. Our analysis revealed significant differences in gene expression and methylation between black and NHW patients. Among the differentially expressed genes, we found significant pathway enrichment within p53/apoptosis signaling pathway and cholesterol/lipid modulation. Additionally, we observed mild hypermethylation in black patients vs. the NHW cohort. Furthermore, we discovered differences in the proportion of important immune cell types between the two groups, with CD8+ T-cells and CD4+ memory-resting T-cells being significantly higher proportionally in black patients.  In conclusion, our study identified significant differential expression between black and NHW patient groups, as well as mild hypermethylation and differences in the proportion of important immune cell types. These findings provide further insights into the biological mechanisms underlying the disparities in outcomes between black and NHW patients with EOC. Further exploration of the contribution of these differences to clinical outcomes and treatment response in black women is warranted.

overall_design: We assessed the transcriptomic and methylomic differences of chemotherapy-resistant HGSOC patient-samples of Black and Non-Hispanic White (NHW) patients using DNA methylation Epic-array and RNA-seq.

first: Russell

last: Keathley

first: Hao

last: Huang


title: Methylomic and transcriptomic differences in treatment-naïve profiles among black and non-Hispanic white (NHW) women [RNA-Seq]

summary: Epithelial ovarian cancer (EOC) is a highly lethal gynecological malignancy, with the high-grade serous ovarian cancer subtype (HGSOC) accounting for a significant proportion of cases. Despite platinum-based chemotherapies being the primary treatment for EOC, HGSOC displays high resistance to these therapies. Black women have the highest mortality/incidence ratio of all ethnic groups, and recent studies suggest significant genetic and epigenetic differences between black and non-Hispanic white women who are treatment naïve. This study aims to explore immunogenic differences and hypermethylation phenotype in black women that may contribute to poorer outcomes within this patient group. We collected primary tumor samples from 36 black and 31 non-Hispanic white (NHW) treatment-naïve patients via biopsy and performed methylomic and transcriptomic analyses. Our analysis revealed significant differences in gene expression and methylation between black and NHW patients. Among the differentially expressed genes, we found significant pathway enrichment within p53/apoptosis signaling pathway and cholesterol/lipid modulation. Additionally, we observed mild hypermethylation in black patients vs. the NHW cohort. Furthermore, we discovered differences in the proportion of important immune cell types between the two groups, with CD8+ T-cells and CD4+ memory-resting T-cells being significantly higher proportionally in black patients. In conclusion, our study identified significant differential expression between black and NHW patient groups, as well as mild hypermethylation and differences in the proportion of important immune cell types. These findings provide further insights into the biological mechanisms underlying the disparities in outcomes between black and NHW patients with EOC. Further exploration of the contribution of these differences to clinical outcomes and treatment response in black women is warranted.


title: Circadian dependent transcriptional oscillation in the olfactory and brain tissues of Aedes and Anopheles mosquitoes regulate perireceptor events and olfactory sensitivity

city: PUNE

first: Tanwee

last: Das De

state: MAHARASHTRA

address: Dr. Homi Bhaba Road, Pashan

department: Department of Biology

country: India

institute: Indian Institute of Science Education and Research

summary: Mosquitoes are the vectors of a variety of infectious diseases. Olfaction and circadian rhythm gate different behavioral events in mosquitoes. However, the mechanisms of odor detection and processing of chemical signals, which are essential for olfactory perception, is poorly understood, including how perception changes in response to zeitgeber time in different mosquito species. To this end, we performed a circadian-dependent RNA-Sequencing study of the brain and peripheral olfactory-tissues of female Anopheles and Aedes mosquitoes. Data analysis revealed a significant upregulation of genes encoding: (a) chemosensory proteins (CSPs), required for transportation of odorant molecules towards the olfactory receptors, and; (b) xenobiotic-metabolizing enzymes (XMEs) during the dusk-transition phase. While XMEs primarily function in the elimination of toxic xenobiotics, concurrent elevation of XMEs and CSPs are hypothesized to act cumulatively for the regulation of perireceptor events and odorant sensitivity. During navigation towards their vertebrate-host/floral-host/mate-partners, female mosquitoes are exposed to diverse environmental chemicals of varied release rates that require continuous degradation/biotransformation by XMEs to avoid receptor saturation in such a noisy environment. Finally, our electroantennographic (EAG) analysis with both Anopheles and Aedes mosquitoes against diverse active volatile odorants, combined with XMEs inhibitors and RNAi studies, establish the proof-of-concept that XMEs function in peri-receptor events during odorant perception and influence the odorant sensitivity in mosquitoes. Additionally, our RNA-Seq and RNAi data also revealed that daily temporal modulation of neuronal serine proteases may facilitate the reorganization of synaptic connections influencing synaptic plasticity and olfactory perception in mosquitoes. In summary, this study highlights the crucial function of XMEs in odorant reception, therefore, improved understanding of species-specific rhythmic expression may provide an important information for successful implementation of mosquito control methods.

sample_organism:
Aedes aegypti
Anopheles culicifacies

platform_organism:
Aedes aegypti
Anopheles culicifacies

overall_design: To understand the circadian dependent molecular rhythm in the olfactory and brain tissues of the diurnal Aedes and nocturnal Anopheles mosquitoes, that drive their different behavioral activities (day time biting behavior of Aedes mosquito and night-time biting behavior of Anopheles mosquito), we performed RNA-Seq study of both olfactory and brain tissues at 4 different circadian time, 6am, 12pm, 6am and 12am. In this study total 488 samples were processed that includes two different tissues (olafctory and brain) from two different mosquito species (Aedes aegypti and Anopheles culicifacies) at four different time points (6am, 12pm, 6pm, 12am) with three biological replicates

first: Krishanpal

last: Karmodiya

first: Rajnikant

last: Dixit

first: Om

middle: P

last: Singh


title: Mediator kinase inhibition suppresses hyperactive interferon signaling in Down syndrome I

city: Boulder

first: Meaghan

last: Courvan

state: CO

address: 596 UCB

department: Biofrontiers

institute: University of Colorado, Boulder

summary: Individuals with Down syndrome have a increased likelihood of suffering from inflammatory and autoimmune conditions, due to the presence of four interferon receptors on chromosome 21. Transcription of genes plays a key role in the interferon response, and the Mediator kinases, CDK8 and CDK19, are essential in potentiating this transcription. Given the role CDK8/19 play in the interferon response, and the fact that selective inhibitors of these kinases exist (such as Cortistatin A), it is possible that targetting Mediator kinase activity may alleviate symptoms of chronic inflammation in Down syndrome.

overall_design: Immortalized lymphoblastoid cells from a pair of brothers, one with Down syndrome and one without, were pre-treated for 30 minutes with either vehicle (10% DMSO) or 100 nM Cortistatin A. After pre-treatment, they were subsequently exposed to either vehicle (40 mM Tris pH 7.4) or 10 ng/mL IFNgamma. After 45 minutes of exposure to vehicle or IFN, nuclei were isolated from cells and nuclear run-on reactions containing biotinylated CTP were performed. Nascent RNA was captured using strepdavidin, and RNA from each condition was reverse-transcribed into cDNA and labeled with unique indices for sequencing.

first: Kira

middle: A

last: Cozzolino


title: Non-canonical hepatic androgen receptor mediates glucagon sensitivity in female mice through the PGC1α/ERRα/mitochondria axis

city: Hangzhou

first: Chen

last: Jie

state: Zhejiang

address: Dunyu Road No.300

department: School of Life Science

country: China

institute: Westlake University

summary: Glucagon has recently been found to modulate liver fat content, in addition to its role in regulating gluconeogenesis. However, the precise mechanisms by which glucagon signaling synchronizes glucose and lipid metabolism in the liver remain poorly understood. By employing chemical and genetic approaches, we demonstrate that inhibiting the androgen receptor (AR) impairs the ability of glucagon to stimulate gluconeogenesis and lipid catabolism in primary hepatocytes and female mice. Notably, AR expression in the liver of female mice is up to three times higher than that in their male littermates, accounting for the more pronounced response to glucagon in females. Mechanistically, hepatic AR promotes energy metabolism and enhances lipid breakdown for liver glucose production in response to glucagon treatment through the peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α)/estrogen-related receptor alpha (ERRα)-mitochondria axis. Overall, our findings highlight the crucial role of hepatic AR in mediating glucagon signaling and the sexual dimorphism in hepatic glucagon sensitivity.

overall_design: mRNA profile of mouse primary hepatocytes treated with shScr±glucagon/shAr±glucagon

first: Jie

last: Chen

first: Jia

last: You

first: Lianfeng

last: Wu


title: Interferons are the key cytokines acting on pancreatic islets in type 1 diabetes

city: Anderlecht

first: Florian

last: Szymczak

state: Brussels

address: Route de Lennik 808 - CP618

department: Faculty of Medicine

country: Belgium

institute: Université Libre de Bruxelles

summary: The pro-inflammatory cytokines IFNα, IFNγ, IL-1β and TNFα may contribute to innate and adaptive immune responses during islet inflammation (insulitis) in type 1 diabetes (T1D). We used deep RNA-sequencing analysis to characterize the response of human pancreatic beta cells to each cytokine individually and compared the signatures obtained with those present in islets of individuals affected by T1D. IFNα and IFNγ had a much greater impact on the beta cell transcriptome when compared to IL-1β and TNFα. The IFN-induced gene signatures have a strong correlation with those observed in beta cells from T1D patients, and the level of expression of specific IFN-stimulated genes is positively correlated with proteins present in islets of these individuals, regulating beta cell responses to “danger signals” such as viral infections. These data suggest that IFNα and IFNγ are the central cytokines at the islet level in T1D, contributing to the triggering and amplification of autoimmunity.

overall_design:
The raw RNA-seq data of EndoC-BH1 cells treated with IFNa, IL-1B, and TNFa [ 2 x 150-bp paired-end with a sequencing depth > 200 million reads (Eurofins Genomics, Germany)] were first subjected to a quality check using fastp to remove low quality reads and adaptor trimming. The passing reads were then subjected to quality control (QC) using FastQC (Babraham Bioinformatics, Cambridge, UK). For comparison purposes, these five paired RNA-seq replicates were re-analyzed as described below. Gene expression was quantified using Salmon v1.457 with additional parameters “-seqBias -gcBias -validateMappings” and expressed in TPM (transcripts per million). GENCODE v36 was used as the reference genome and indexed with default k-mer values.
The genes differentially expressed after exposure to cytokines were identified by DESeq2 v1.30.158. For each gene, a log2 fold change value was obtained based on the comparison performed (cytokine-treated condition versus non-treated condition). Each gene comparison has an associated Wald test statistic, p-value, and an adjusted p-value (Benjamini-Hochberg correction). Genes presenting an adjusted p-value < 0.05 were considered significantly differentially expressed.

last: Coomans de Brachène

first: Maria

last: Ines Alvelos

first: Décio

middle: L

last: Eizirik


title: Apical-basal polarity drives pancreatic alpha/beta cell fate allocation

city: Munich

first: Chenglei

last: Tian

address: Ingolstädter Landstraße 1

institute: Helmholtz Munich

summary: A central question in cell and developmental biology is how extracellular cues control the differentiation of multipotent progenitors in a dynamically changing niche. Here, we identify apical-basal polarity as the main regulator of the differentiation of multipotent pancreatic Neurogenin3+ endocrine progenitors (EPs) into the beta or alpha cell fates. We show that human EPs dynamically change their apical-basal polarity status. Whereas polarized EPs are predisposed to differentiate into beta cells rather than alpha cells, inhibiting apical-basal polarity selectively suppresses beta cell differentiation. Single-cell RNA sequencing and complementary mechanistic data demonstrate that apical-basal polarity in human EPs promotes beta cell specification via cyclic AMP (cAMP)/PKA-cAMP response element binding protein (CREB)-EGR1-mediated inhibition of ARX expression, while reduced cAMP levels in non-polarized human EPs maintain expression of ARX, leading to alpha cell differentiation. These findings identify the apical-basal polarity status of multipotent EPs as a critical epithelial feature that determines their fate into the alpha or beta cell lineages.

overall_design: We performed scRNA sequencing to understand how apical-basal polarity drives pancreatic alpha/beta cell fate allocation. We combined this technology to CITE-sequencing and HASH-sequencing in order to measure these data simultaneously in early/late and polarized/unpolarized endorine progenitors.

first: Ulf

last: Tiemann

first: Martin

last: Proks

first: Henrik

last: Semb


title: The inhibition of inner mitochondrial fusion in hepatocytes reduces non-alcoholic fatty liver and improves metabolic profile during obesity by modulating bile acid conjugation

city: Milano

first: Diletta

last: Dolfini

state: MI

address: Via Celoria, 26

department: Bioscience

country: Italy

institute: University of Milan

summary: Hepatocyte OPA1 deficiency protects mice from the development of metabolic syndrome by reducing lipid absorption from the diet. OPA1 deficiency affects bile acid conjugation as a consequence of reduced mitochondria-peroxisome-ER tethering.

overall_design: Comparative gene expression profiling analysis of RNA-seq data in CTRL and OPA1 hepatocyte KO mouse liver

first: Mirko

last: Ronzio

first: Danilo

last: Norata

first: Lorenzo

last: Da Dalt

first: Annalisa

last: Moregola


title: Gene expression signatures of spleen-derived and bone marrow-derived CD11b+ cells in high-fat diet (HFD)-fed mice

city: Shanghai

first: Xiangdong

last: Yang

state: China

address: 180 Fenglin Road, Xuhui District, Shanghai

institute: Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital, Fudan University

summary: To further search for potential gene targets which are involved in the pathological process of atherosclerosis, we have employed whole genome microarray expression profiling as a discovery platform to identify the expression of genes related to lipid metabolism. The CD11b+ cells were isolated from spleen and bone marrow of mice by magnetic beads, and four genotype mice of HDC-/-, STAT4-/-, STAT6-/- and WT were examined in this experiment. The mice were divided into 2 groups:  1. control group 2.  HFD group. Mice in control group were given a normal diet; mice in HFD group were fed with high-fat diet for 3 months. The data showed hundreds of up-regulated or down-regulated genes in HFD-fed mice, which were functionally connected with cholesterol synthesis and reverse cholesterol transport.

overall_design: The gene expression was measured in spleen-derived and bone marrow-derived CD11b+ cells of mice. Mice were divided into control and HFD groups. There were five mice in each group.

first: Baoling

last: Zhu


title: RNA Sequencing Facilitates Quantitative Analysis of Humanized Normal Livers, NASH Livers, and HCC Transcriptomes

city: Houston

first: Loning

last: Fu

state: Texas

address: One Baylor Plaza

department: Medicine

institute: Baylor College of Medicine

summary: RNAseq analysis that will help to define 1) the circadian dysfunction induced liver transcriptome that drives fatty liver disease induced hepatocarcinogenesis in a humanized mouse model; 2) the currently unknown circadian profile of NASH and HCC gene signatures specifically associated with human hepatocytes and liver or tumor microenvironment. These gene signatures will significantly advance our understanding of the mechanism of spontaneous HCC and have important preclinic values for developing novel therapeutic strategies for prevention and treatment of HCC in humans.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Control: mRNA prepared from normal livers, NASH livers, or HCCs of mice maintained in steady 12 hour light/12 hour dark (24 hrLD) cycles (n=4/ZT/group); CJ (chronic jet lag): mRNA prepared from livers or HCCs of mice treated with one 8-hr phase advance each Monday followd by an 8-hr phase delay on each Thursday (n=4/ZT in CJ NASH group and n=6/ZT in CJ HCC group). h-Liver: humanized liver (only hepatocytes in the livers are human origin); h-HCC: humanized HCC (only malignant hepatocytes in tumors are human origin).

first: Noha

last: Osman

first: David

last: Moore

first: Jennifer

last: Padilla


title: REPLACED BY GSE136256

city: Kansas City

first: Shiyuan

middle: (Cynthia)

last: Chen

state: MO

department: Computational Biology

institute: Stowers Institute for Medical Research

summary: The data for this Series have been removed by request of the submitter. The submitter states that the data had compromised quality and the replicates were inconsistent due to the biological replicates.

overall_design: Replaced by GSE136256.

first: Xi

middle: C

last: He

first: Linheng

last: Li


title: RNA sequencing of NHEK treated with polyIC and siRNaseL for 48 hours;

city: Baltimore

first: Luis

middle: Andres

last: Garza

state: MD

address: 1550 Orleans St. Suite 204 CRBII Koch

department: Dermatology

institute: Johns Hopkins School of Medicine

summary: Mammalian injury responses are characterized by fibrosis and scarring rather than functional regeneration. Limited regenerative capacity in mammals could reflect a loss of pro-regeneration programs or active suppression by genes functioning akin to tumor suppressors. To uncover programs governing regeneration in mammals, we performed comprehensive transcript screening in human subjects after laser rejuvenation treatment and cross-referenced these transcripts to those found in mice with enhanced Wound Induced Hair Neogenesis (WIHN), a rare example of mammalian organogenesis1,2. We find the anti-viral endoribonuclease RNase L to be a powerful suppressor of regeneration. Rnasel-/- mice exhibit remarkable regenerative capacity, with elevated WIHN through enhanced IL-36α. Consistent with the known role of RNase L to stimulate caspase-1, we find that pharmacologic inhibition of caspases promotes regeneration in a novel IL-36-dependent manner. Additionally, these responses are not limited to skin but extend to other organs, such as the colon, suggesting that suppression of regeneration is a fundamental characteristic of epithelial wound healing. Taken together, this work suggests that RNase L functions as a regeneration repressor gene in a functional tradeoff that prioritizes host antiviral abilities and is a target to enhance healing in multiple epithelial organs, perhaps even during viral infection.

overall_design: Four samples were analyzed, one for each condition.

first: Nasif

last: Islam

first: Charles

last: Kirby


title: Spatial transcriptomics and CUT&RUN of the aging mouse brain reveal origins of inflammation in the white matter

status: Public on Dec 31 2024

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH


title: Exosomal circ_0006896 promotes AML progression via interaction with HDAC1 and restriction of antitumor immunity

city: Jinan

first: Daoxin

last: Ma

state: shandong

address: No.107,West of Wenhua Road

institute: Qilu Hospital of Shandong University

summary: Drug resistance and immune escape continue to contribute to poor prognosis in AML. Increasing evidence suggests that exosomes play a crucial role in AML immune microenvironment. We aimed to find a functional exosomal circRNAs/lncRNA/mRNA correlating with the progression of AML patients and further analyze its underlying mechanism in AML cells and tumor microenvironment immune cells. We find a new crucial exosomal circRNA, circ_0006896, is upregulated in both AML cells and exosomes and correlates with the prognosis and relapse of AML. In vitro and in vivo studies suggest that circ_0006896 significantly promotes AML cell proliferation, reduces chemotherapy sensitivity, and more importantly, impairs the efficacy of adoptive T cell-transfer immunotherapy.

overall_design: The exosomes of the bone marrow serums from 6 AML patients and 4 healthy donors were analyzed for transcriptome sequencing using IlluminaNovaSeq6000 system. Transcriptional analysis of Molm-13 cells transfected with NC-shRNA or circ_0006896-shRNA.


title: STING-mediated proton channel activity, signaling, and autoinflammatory disease require ArfGAP2 [Jurkat]

city: Philadelphia

first: Samuel

last: Chauvin

state: PA

address: 518 Johnson Pavilion, 3610 Hamilton Walk

institute: University of Pennsylvania Perelman School of Medicine

summary: STING transmits signals downstream of the cytosolic DNA sensor cGAS, leading to transcriptional up-regulation of cytokines. However, components of the STING signaling pathway, such as IRF3 and IFNAR1, are not essential for autoinflammatory disease in STING gain-of-function (SAVI) mice. Recent discoveries revealed that STING also functions as a proton channel that deacidifies the Golgi. Since pH impacts Golgi enzyme activity, protein maturation, and trafficking, we hypothesized that STING proton channel activity influences multiple Golgi functions. Here, we show that STING-mediated proton efflux non-transcriptionally regulates Golgi trafficking of protein cargos. This process requires the Golgi-associated protein ArfGAP2, a cell type-specific dual regulator of STING-mediated proton efflux and signaling. Deletion of ArfGAP2 in hematopoietic and endothelial cells markedly reduces STING-mediated cytokine and chemokine secretion, immune cell activation, and autoinflammatory pathology in SAVI mice. Thus, ArfGAP2 facilitates STING-mediated signaling and cytokine release in hematopoietic cells, significantly contributing to autoinflammatory disease pathogenesis.

overall_design: RNA-seq profiling of control and ArfGAP2 knockout Jurkat T cells after 36 hour induction of either WT STING or STING N154S.

first: Subhajit

last: Poddar

middle: D

first: Wei

last: Qian

first: Elisha

last: Roberson

first: Jonathan

last: Miner


title: STING-mediated proton channel activity, signaling, and autoinflammatory disease require ArfGAP2 [BMDM]

overall_design: RNA-seq profiling of control and ArfGAP2 knockout bone marrow-derived macrophages 24 hours after stimulation with the STING agonist diABZI (or a vehicle control).


title: B cell stimulation changes the structure and higher-order organization of the inactive X chromosome

city: PHILADELPHIA

first: MONTSERRAT

last: ANGUERA

address: 380 S. University Ave, Room 390EB

department: Biomedical Sciences

institute: University of Pennsylvania

summary: X Chromosome Inactivation (XCI) equalizes X-linked gene expression between sexes. B cells exhibit dynamic XCI, with Xist RNA/heterochromatic marks absent on the inactive X (Xi) in naive B cells but returning following mitogenic stimulation. The impact of dynamic XCI on Xi structure and maintenance was previously unknown. Here, we find dosage compensation of the Xi with state-specific XCI escape genes in naive and in vitro activated B cells. Allele-specific OligoPaints indicate similar Xi and Xa territories in B cells that are less compact than in fibroblasts. Allele-specific Hi-C reveals a lack of TAD-like structures on the Xi of naive B cells, and stimulation-induced alterations in TAD-like boundary strength independent of gene expression. Notably, Xist deletion in B cells changes TAD boundaries and large-scale Xi compaction. Altogether, our results uncover B cell-specific Xi plasticity which could underlie sex-biased biological mechanisms.

overall_design: To determine how Xist deletion impacts transcriptional changes in splenic follicluar B cells, we sorted CD23+ splenic B cell subsets from female C57/BL6 WT and Xist cKO mice. We stimulated some follicular B cells with the TLR9 agonist CpG in vitro for 24 hours. We performed RNA sequencing naive and CpG-stimulated splenic follicular B cells with at least n=3 biological replicates per experimental group.

first: Isabel

last: Sierra

first: Natalie

middle: E

last: Toothacre

first: Robin

last: van de Weide

first: Claudia

last: Lovell

first: Son

last: Nguyen

first: R

last: Varnett

first: Ashley

last: Cook

first: Han-Seul

last: Ryu

first: Sarah

last: Pyfrom

first: Harrison

last: Wang

first: Daniel

last: Beiting

last: Philips-Cremins

first: Eric

middle: F

last: Joyce

first: Montserrat

last: Anguera


title: Fate Erasure Logic of Gene Networks Underlying Direct Neuronal Conversion of Somatic Cells by MicroRNAs III

city: Saint Louis

first: Kitra

last: Cates

address: 4523 Clayton Ave, MCD SCI 361

department: Developmental Biology

institute: Washington University in St. Louis

summary: Neurogenic microRNAs 9/9* and 124 (miR-9/9*-124) drive the direct reprogramming of human fibroblasts into neurons with the initiation of the fate erasure of fibroblasts. However, whether the miR-9/9*-124 fate erasure logic extends to neuronal conversion of other somatic cell types remains unknown. Here, we uncover that miR-9/9*-124 induce neuronal conversion of multiple cell types: dura fibroblasts, astrocytes, smooth muscle cells, and pericytes. We reveal the cell type-specific and pan-somatic gene network erasure induced by miR-9/9*-124, including cell cycle, morphology, and proteostasis gene networks. Leveraging these pan-somatic gene networks, we predict upstream regulators that may antagonize somatic fate erasure. Among the predicted regulators, we identify TP53 (p53) whose inhibition is sufficient to enhance neuronal conversion even in post-mitotic cells. This study extends miR-9/9*-124 reprogramming to alternate somatic cells, reveals the pan-somatic gene network fate erasure logic of miR-9/9*-124, and shows a neurogenic role for p53-inhibition in the miR-9/9*-124-signaling cascade.

overall_design: RNA-sequencing from a panel of human dermal fibroblasts, including lines AG13369, AG08379, GM23966, and F09-110, expressing pT-miR-9/9*-124 with rtTA-CTL or rtTA-p53DD for 5 or 15 days.

first: Andrew

middle: S

last: Yoo


title: Fate Erasure Logic of Gene Networks Underlying Direct Neuronal Conversion of Somatic Cells by MicroRNAs II

overall_design: RNA-sequencing from a panel of human dermal fibroblasts, including the following lines: AG13369 (NIA Aging Cell Culture Repository at the Coriell Institute for Medical Research), AG08379 (NIA Aging Cell Culture Repository at the Coriell Institute for Medical Research), AG06858 (NIA Aging Cell Culture Repository at the Coriell Institute for Medical Research), GM23974 (NIGMS Human Genetic Cell Repository at the Coriell Institute for Medical Research), GM23966 (NIGMS Human Genetic Cell Repository at the Coriell Institute for Medical Research), and F09-110 (deidentified fibroblast contributed by Pat Blanner). RNA was collected from each line expressing pT-miR-9/9*-124 for 20 days (Day 20) as compared to starting fibroblasts (Day 0).


title: tPOD-seq: a high throughput RNA-seq method to derive transcriptomic points of departure from cell lines

city: Sainte-Anne-de-Bellevue

first: Niladri

last: Basu

state: Quebec

address: 21,1111 Lakeshore Blvd, Apt #2

country: Canada

institute: McGill University

summary: There is growing scientific and regulatory interest in transcriptomic points of departure (tPOD) values from high-throughput in vitro experiments. To further help democratize tPOD research, here we outline ‘tPOD-seq’ which links microplate-based exposure methods involving cell lines for human (Caco-2, Hep G2) and environmental (rainbow trout RTgill-W1) health, with a commercially available RNA-seq kit, with a cloud-based bioinformatics tool (ExpressAnalyst.ca).  To demonstrate tPOD-seq’s utility, we applied it to derive tPODs for solvents (dimethyl sulfoxide, DMSO; methanol) and positive controls (3,4-dichloroaniline, DCA; hydrogen peroxide, H2O2) commonly used in toxicity testing. The majority of reads mapped to protein coding genes (~10.5k for fish cells; ~7k for human cells), and about 50% of differentially expressed genes were curve-fitted from which 90% yielded gene benchmark doses.  The most robust transcriptomic responses were caused by DMSO exposure, and tPOD values were 31-136.6 mM across the cell lines. OECD test guideline 249 (RTgill-W1 cells) recommends the use of DCA and here we calculated a tPOD of ~0.005 to ~0.076 mMmg/L.  Finally, exposure of the two human cell lines to H2O2 resulted in similar tPOD values that ranged from 0.68-1.1 in Caco-2 cells and  0.0048-0.03 in Hep G2 cells.  The methods outlined here are designed to be performed in laboratories with basic molecular and cell culture facilities, and the performance and scalability of the tPOD-seq approach can be determined with additional case studies.

sample_organism:
Oncorhynchus mykiss
Homo sapiens

platform_organism:
Oncorhynchus mykiss
Homo sapiens

overall_design: Caco-2, Hep G2 and RTgill-W1 cell lines were exposed to  solvents and positive controls commonly used  for in vitro assays at various concentrations. Endpoints examined were cytotoxicity and transcriptomics

first: Krittika

last: Mittal

first: Ke

last: Xu

first: Samuel

last: Rulli Jr


title: Fate Erasure Logic of Gene Networks Underlying Direct Neuronal Conversion of Somatic Cells by MicroRNAs IV

overall_design: Small RNA-sequencing for neonatal human dermal fibroblasts expressing either pT-miR-NS or pT-miR-9/9*-124 for 6 days.

first: Yangjian

last: Liu

first: Paul

last: Gontarz


title: Fate Erasure Logic of Gene Networks Underlying Direct Neuronal Conversion of Somatic Cells by MicroRNAs

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

overall_design: Refer to individual Series


title: Polymerase-Usage Sequencing Identifies Initiation Zones With Less Bias Throughout S phase in Mouse Embryonic Stem Cells

city: Okazaki

first: Tomomi

last: Tsubouchi

state: Aichi

address: Nishigonaka 38, Myodaiji

institute: National Institute for Basic Biology

summary: In mammalian cells, DNA replication can initiate from various locations even in different cycles of the same cell of origin. Various methods have been developed to map replication initiation zones (IZs) genome-wide, often finding IZs far less than expected considering the replication fork speed and genome size. In particular, IZs for later stages of S phase are under-represented, potentially due to reduced initiation events at this stage, but also likely to be due to variability in the population, requiring higher sensitivity of the methods. However, to what degree these factors contribute to the data, have not been investigated in detail. Mouse ES cells present similar density of replication initiation events throughout S phase, unlike non-ES cells. Here, we re-analyzed IZs with respect to replication timing within S phase, using data sets previously obtained for mouse ES cells. These data sets identified over 5-fold more early IZs compared to late IZs, suggesting that more than 80% of late IZs are missed. In addition, by newly establishing cell lines to carry out polymerase-usage sequencing (Pu-seq), we mapped IZs genome-wide in mouse ES cells. Pu-seq presented less bias towards early IZs, suggesting that better sensitivity for identifying IZs in late S phase.

overall_design: We mapped ribonucleotide-incorporation by the mutated DNA polymerase ε (Pole1-M630F) at single-nucleotide resolution in mouse embryonic stem cells and subsequent informatics analysis was performed to generate datasets listed here.

first: Akino

last: Matsumoto

first: Yasukazu

last: Daigaku


title: Fate Erasure Logic of Gene Networks Underlying Direct Neuronal Conversion of Somatic Cells by MicroRNAs I

overall_design: RNA-sequencing from a panel of human somatic cells, including dermal fibroblasts (hDF), dural fibroblasts (hDuF), astrocytes (hAst), brain vascular smooth muscle cells (hBVSMC), and brain vascular pericytes (hBVP),  expressing either pT-miR-NS or pT-miR-9/9*-124 for five days


title: Gene expression profile at single cell level of immune cells from naïve mouse bladder

city: Hefei

first: Lu

last: Li

address: #373, Huangshan Road

institute: University of Science and Technology of China

summary: We set out to study the immune landscape of the mouse bladder by single-cell RNA sequencing (scRNA-seq), especially the heterogeneity of bladder-resident macrophages. Identifying TRM subsets in the bladder and specifying the distinct function of each subpopulation would therefore be essential for a comprehensive understanding of the immune responses during UTIs.

overall_design: Bladder single cell was isolated from naïve adult mice, CD45+ immune cells were sorted (FACS) and analyzed using scRNAseq.

first: Qiancheng

last: Jiao

first: Zhutian

last: Zeng


title: Attempts to create transgenic mice carrying the Q3925E mutation in RyR2 Ca2+ binding site

city: Charleston

first: Allison

last: Trouten

state: SC

address: 173 Ashley Ave Room BSB 642

department: Regenerative Medicine and Cell Biology

institute: Medical University of South Carolina

summary: Over 200 point mutations in ryanodine receptor (RyR2) of cardiac sarcoplasmic reticulum (SR) are known to associate with cardiac arrhythmia. We have already reported on the calcium signaling phenotype of a point mutation in RyR2 Ca2+ binding site Q3925E expressed in human stem-cell derived cardiomyocytes (hiPSC-CMs) that was found to be lethal in a 9 year old girl. CRISPR/Cas9 gene edited mutant cardiomyocytes carrying RyR2-Q3925E mutation exhibited loss of calcium-induced calcium release (CICR) and caffeine-triggered calcium release, but continued to beat arrhythmically without generating significant SR Ca2+ release, consistent with remodeling of calcium signaling pathway. RNAseq heat-map confirmed significant changes of calcium associated genes, supporting the remodeling possibility. To determine the in-situ cardiac phenotype in animal model of this mutation, we generated a knock-in mouse model of RyR2-Q3924E+/– using CRISPR/Cas9 technique. We obtained 3 homozygous and one chimera mice, but they all died before 3 weeks of age preventing the establishment of germline mutation transmission in the offsprings. Histo-pathological analysis of the heart showed significant cardiac hypertrophy suggesting Q3924E-RyR2 mutation was lethal in the mouse.

overall_design: We sought to utilize hiPSC-CMs to recapitulate the previously identified Q3925E and E3848A mutations using CRISPR/Cas9 gene editing

first: Xiao-hua

last: Zhang

first: Fu-Lei

last: Tang

first: Martin

last: Morad


title: HalALMT1 mediates malate efflux in the cortex of mature cluster rootlets of Hakea laurina, occurring naturally in phosphorus-poor soil

city: Higashi-Hiroshima

first: Hirotsuna

last: Yamada

state: Hiroshima

address: 7-1 1-chome Kagamiyama

department: Graduate school of integrated science for life

institute: Hiroshima University

summary: Hakea laurina, a woody plant in Proteaceae family, naturally occurs in severely phosphorus (P)-impoverished environments in southwest Australia. It develops distinctive “cluster roots” that exhibit a high capacity for carboxylate exudation and acid phosphatase activity, contributing to P acquisition. However, the molecular mechanisms underlying these physiological functions remain poorly understood. We explored the cluster root transcriptome using de novo RNA-Seq, which detected homologous transcripts of an aluminum (Al)-activated malate transporter (ALMT), designated HalALMT1, and two multidrug and toxic compound extrusion (MATE) proteins specifically expressed in mature cluster roots. Deferentially-expressed genes highlighted multiple carboxylate-related processes at maturity. Electrophysiological assays and overexpressed transgenic experiments confirmed HalALMT1’s ability to release malate and that its activity is enhanced by exposure to Al3+. Notably, HalALMT1 was specifically expressed in the mature cortex cells of cluster rootlets, which do not develop a suberized exodermis. Furthermore, acid phosphatase activity was pronounced throughout the rootlets, whereas it was only present in the epidermis and stele of non-cluster roots, revealing its extensive production in the rootlets. These results suggest that malate release and acid phosphatase production in the dominant cortex cells in cluster rootlets increase the amount exuded. This study sheds light on an outstanding P-acquisition strategy of Proteaceae.

overall_design: After germinating the Hakea laurina seedlings, six seedlings were transferred to a 4 L hydroponic solution with low P, specifically in a basic 0 µM P hydroponic setup supplemented with 6.4 µM NaH2PO4 every two to three months, and cultivated in the glasshouse at Hiroshima University. Mature cluster roots (MCR) and tips of cluster-bearing lateral roots (TCR) produced by approximate-three-year-old seedlings were harvested (n = 3). Roots were promptly frozen in liquid nitrogen and stored at -80˚C for RNA extraction, which was subjected to de novo RNA-Seq analsis. We investigated the genes related in phosphorus absorption and up-regulated in MCR.

first: Jun

last: Wasaki

first: Sho

last: Nishida

first: Takuo

last: Enomoto

first: Akira

last: Tanaka


title: Passive Infusion of an S2-Stem Broadly Neutralizing Antibody protects against SARS-CoV-2 infection and lower airway inflammation

city: Atlanta

first: Gregory

middle: K

last: Tharp

state: GA

address: 954 Gatewood Dr

department: Developmental and Cognitive Neuroscience

institute: Yerkes National Primate Research Center

summary: The continued evolution of SARS-CoV-2 variants capable of subverting vaccine and infection-induced immunity suggests the advantage of a broadly protective vaccine against betacoronaviruses (β-CoVs). Recent studies have isolated monoclonal antibodies (mAbs) from SARS-CoV-2 recovered-vaccinated donors capable of neutralizing many variants of SARS-CoV-2 and other β-CoVs. Many of these mAbs target the conserved S2 stem region of the SARS-CoV-2 spike protein, rather the receptor binding domain contained within S1 primarily targeted by current SARS-CoV-2 vaccines. One of these S2-directed mAbs, CC40.8, has demonstrated protective efficacy in small animal models against SARS-CoV-2 challenge. As the next step in the pre-clinical testing of S2-directed antibodies as a strategy to protect from SARS-CoV-2 infection, we evaluated the in vivo efficacy of CC40.8 in a clinically relevant non-human primate model by conducting passive antibody transfer to rhesus macaques (RM) followed by SARS-CoV-2 challenge. CC40.8 mAb was intravenously infused at 10mg/kg, 1mg/kg, or 0.1 mg/kg into groups (n=6) of RM, alongside one group that received a control antibody (PGT121). Viral loads in the lower airway were significantly reduced in animals receiving higher doses of CC40.8. We observed a significant reduction in inflammatory cytokines and macrophages within the lower airway of animals infused with 10mg/kg and 1mg/kg doses of CC40.8. Viral genome sequencing demonstrated a lack of escape mutations in the CC40.8 epitope. Collectively, these data demonstrate the protective efficiency of broadly neutralizing S2-targeting antibodies against SARS-CoV-2 infection within the lower airway while providing critical preclinical work necessary for the development of pan–β-CoV vaccines.

overall_design: 24 RMs (6 females and 18 males; mean age of 5 years and 11 months old; range 5-6 years old) were infused intravenously 5 days pre infection with either a 0.1 mg/kg, 1mg/kg, or 10mg/kg concentration of experimental mAb CC40.8 or with control mAb PGT121, with each experimental group consisting of 6 animals (1 female and 5 males). Animals were screened for preexisting, SARS-CoV-2 spike specific antibodies prior to enrollment in this study. Preinfection baseline samples were collected 4 days prior to inoculation. At day 0, all RM were inoculated with 1mL intranasally and 1mL intratracheally with a combined total of 1.1 x 106 plaque forming units (PFU) of SARS-CoV-2 (2019-nCoV/USA-WA1/202). BAL and nasal swabs were collected from inoculated animals at 2dpi and at the time of necropsy (7 or 8dpi), with viral titers peaking in these tissues at 2dpi in the infected animals. PBMCs were isolated from the BAL of SARS-CoV-2 infected RM at 2 dpi and 7/8 dpi and captured for 10x scRNA-seq. Samples were multiplexed in groups of 3.

first: Christopher

middle: T

last: Edwards

first: Steven

last: Bosinger

first: Kirti

last: Karunakaran

first: Elijah

last: Garcia

first: Thomas

middle: R

last: Rogers

first: Kathryn

last: Pelligrini

first: Dennis

last: Burton

first: Raiees

last: Andrabi

first: Stacey

last: Lapp

first: Thang

last: Ton

first: Mark

last: Lin

first: Amanda

last: Metz


title: CH6824025, potent and selective DDR1 inhibitor, reduces kidney fibrosis in UUO mice

city: Totsuka-ku, Yokohama

first: Yukari

last: Yasui

state: kanagawa

address: Totsuka-cho 216

department: Research Div.

institute: Chugai Pharmaceutical Co., Ltd.

summary: Discoidin domain receptor 1 (DDR1) is a collagen receptor with tyrosine kinase activity, and its expression is enhanced in various disease conditions. Although previous research suggests that DDR1 contributes to renal disease progression, DDR1 inhibitors for renal fibrosis have yet to be developed. In this study, we used unilateral ureteral obstruction (UUO) mice to investigate whether CH6824025, a strong and selective DDR1 phosphorylation inhibitor, can improve renal fibrosis. Furthermore, to analyze its action in detail, we performed 10x Visium spatial transcriptomics (ST) analysis on the kidney. Orally administered CH6824025 suppressed the phosphorylation of DDR1 in renal tissue, and the amount of hydroxyproline, the Sirius red-positive area, and the mRNA expression of fibrosis and inflammation-related genes in the kidney were significantly decreased. 10x Visium ST analysis suggested that DDR1 is mainly expressed in distal nephrons under normal conditions, but that its expression appears to increase in the injured proximal tubules in UUO mice. Comparing mRNA expression in DDR1 positive spots in the vehicle and the CH6824025 administration group, oxidative phosphorylation and mitochondrial dysfunction were improved, and pathways involved in fibrosis inhibited in the CH6824025 administration group. Downstream analysis suggested that mRNA expression changes in the CH6824025 administration group contribute to the inhibition of cell movement. Taken together, our findings suggest that CH6824025 inhibited the migration of inflammatory cells to the injury site in UUO mice, reduced inflammation, and brought about the maintenance of cell homeostasis and inhibition of fibrosis. DDR1 inhibitors are expected to be a promising treatment for renal fibrosis.

overall_design: After the allocation of mice, the oral once daily administration of vehicle or CH6824025 was started. Seven days after the surgery of UUO, mice were subjected to analysis. FFPE blocks of kidneys from each group (Sham n=1, UUO Vehicle group n=3, UUO CH6824025 200 mg/kg group n=4) were used.

first: Takeshi

last: Murata

first: Yoshinori

last: Tsuboi

first: Naoshi

last: Horiba


title: Transcriptomic analysis of NAFLD model cells with the co-administration of pseudo-natural flavonol and copper

city: Wuhan

first: Xiang

last: Li

address: Youyi Road

institute: Hubei University

summary: Copper deficiency is closely associated with non-alcoholic fatty liver disease (NAFLD). We developed pseudo-natural flavonol (HQF) as novel ionophore for rapid intracellular copper delivery. The co-administration of HQF and copper has synergistic effects in restraining lipid droplet formation with unknown mechanism. Here, hepatic HepG2 cells were treated with palmitic acid (PA) to construct a NAFLD cell model, which was used to investigate the biological effects of HQF-mediated copper delivery on NAFLD intervention.

overall_design: We performed gene expression profiling analysis using data obtained from mRNA-seq of NAFLD model cells, with or without the co-administration 0.5 µM HQF and 5 µM CuCl2 for 24 h. For the construction of NAFLD cell model, HepG2 cells were treated with 750 μM palmitic acid for 24 h, and the palmitic acid solution was blocked by BSA. PA: NAFLD model cells, PA_CuCl2_HQF: NAFLD model cells treated with copper and HQF.

first: Si

last: Qin

first: Qin


title: Piezo1 positively regulates the differentiation of Interleukin-9-producing CD4+ T cells in cancer

city: beijing

first: qiuli

last: yang

address: beijing

institute: beijing normal university

summary: RNA-seq analysis of tumor-infiltrating CD4 T cells from B16 melanoma with and without CD4-specific Piezo1 genetic deletion

overall_design: To invastigate the mechanism of Piezo1 in Th9 cell differentiation, CD4 T cells were sorted by flow cytometry and RNA-seq was performed.

first: Liu

last: guangwei

first: Yang

last: qiuli


title: Natural genetic variation in wild-derived mice controls host survival and transcriptional responses during endotoxic shock

city: Burlington

first: Seth

last: Frietze

state: VT

address: 149 Beaumont Avenue

department: Biomedical and Health Sciences

institute: University of Vermont

summary: Innate immune cells sense microbial danger signals such as bacterial lipopolysaccharide (LPS) through pattern recognition receptors, resulting in dynamic transcriptional reprogramming and rapid inflammatory responses. If not properly regulated, such responses can be detrimental to the host, as is seen in septic shock. A better understanding of the genetic regulation of transcriptional responses during endotoxemia could provide potential therapeutic insights. However, the majority of animal model studies have been performed using classic inbred laboratory strains of mice, typically C57BL/6 (B6), capturing limited genetic diversity and representing a potential blind spot. Here, we compared classic inbred B6 mice with wild-derived and genetically divergent PWD/PhJ (PWD) mice using in vivo and in vitro models of endotoxemia, to determine the role of natural genetic variation in septic shock. Compared with B6 mice, PWD mice were markedly resistant to LPS-induced endotoxic shock. Using LPS stimulation of bone marrow derived dendritic cells (BMDC) and RNA sequencing, we demonstrate that B6 and PWD BMDCs exhibit partially overlapping, yet highly divergent transcriptional responses, with B6 skewed towards stereotypical proinflammatory pathway activation, and PWD engaging regulatory or developmental pathways. To dissect genetic regulation of inflammatory responses by allelic variants, we used BMDCs from a sub-consomic strain carrying a ~50 Mb PWD-derived portion of chromosome 11 on a B6 background. This identified a subset of cis regulated and a large number of trans regulated genes. Bioinformatic analyses identified candidate trans regulators encoded in the chromosome 11 locus as transcription factors Irf1, Ncor1, and Srebf1. Taken together, our results demonstrate that natural genetic variation exerts a pronounced effect on host survival and transcriptional reprogramming during endotoxemia, suggesting possibilities for prediction of sepsis risk and/or personalized therapeutic interventions.

overall_design: This study includes an analysis of bulk RNA-seq data from four biological replicates per genotype, representing independent BMDC cultures from individual mice. BMDCs were seeded at 1x106 cells/well in a 12 well plate and treated with LPS for 6 hours at 10ng/ml concentration.


title: Intraesophageally challenge of OVA to epicutaneously sensitized mice with loss of epithelial Ikkβ results in experimental EoE that is faithful to human EoE esophageal mucosal pathology.

city: Chicago

first: Marie-Pier

last: Tetreault

state: IL

address: 300 East Superior Street

department: Medicine/GI&Hep division

institute: Northwestern University

summary: We report a novel mouse model that can faithfully mimic human EoE

overall_design: Examinationof 20 EoE patients and 8 healthy donors by bulk RNA-seq, with proximal and distal esophageal biopsies from each.

first: Margarette

last: Clevenger

first: Cenfu

last: Wei


title: Huntingtin is an RNA binding protein and participates in NEAT1-mediated paraspeckles

city: Toronto

first: Tiantian

state: Ontario

address: 101 College St.

institute: University Health Network

summary: Huntingtin protein, mutated in Huntington disease, is implicated in nucleic acid- mediated processes, yet evidence for direct huntingtin-nucleic acid interaction is limited. Here we show wildtype and mutant huntingtin co-purify with nucleic acids, primarily RNA, and interact directly with G-rich RNAs in in vitro assays. Huntingtin RNA immunoprecipitation sequencing from patient-derived fibroblasts and neuronal progenitor cells expressing wildtype and mutant huntingtin revealed NEAT1 as a significantly enriched transcript. Altered NEAT1 levels were evident in Huntington’s disease cells and postmortem brain tissues, and huntingtin knockdown decreased NEAT1 levels. Huntingtin co-localized with NEAT1 in paraspeckles, and we identified a high-affinity RNA motif preferred by huntingtin. This study highlights NEAT1 as a novel huntingtin interactor, demonstrating huntingtin’s involvement in RNA-mediated functions and paraspeckle regulation.

overall_design: Differential gene expression analysis of RIP-seq data for Input and IP of huntingtin cells.

first: Manisha

last: Yadav

first: Rachel

last: Harding

first: Cheryl

middle: H

last: Arrowsmith

first: Housheng

last: He


title: Gene expression profile at single cell level of CD45+ cells that were isolated from the lung of anti-PD-1 Ab-treated young or aged mice

city: Kyoto

first: Hirotake

last: Tsukamoto

state: Sakyo-Ku

address: 53 Kawahara-cho Shogoin

department: Center for Cancer Immunotherapy and Immunobiology

institute: Kyoto University

summary: In tumor-bearing aged mice, abberant immune cells are expected to be accumulated in damaged lung of aged mice in response to anti-PD-1 therapy. To identify subpopulation responsible for the lung damage, we analyzed for differences in CD45+ immune cell components isolated from the lung of young and aged mice, using single-cell RNA sequencing (scRNA-seq).

overall_design: After 3 times of administration with anti-PD-1 Ab, CD45+ cells were isolated from the lung of MC38 tumor-bearing young or aged mice using magnetic sorting and dead cell removing.

first: Tsukamoto

last: Hirotake

first: Hasan

last: Sharif


title: Genome-wide CRISPR Screen Identifies DNTT as a Key Regulator of Inotuzumab Ozogamicin Response in B-cell Acute Lymphoblastic Leukemia

city: Memphis

first: Carolin

last: Escherich

state: Tennessee

address: 262 Danny Thomas Place

institute: St. Jude Children's Research Hospital

summary: Inotuzumab Ozogamicin (InO) is an antibody-drug conjugate with striking efficacy in B-cell acute lymphoblastic leukemia (B-ALL). However, there is wide inter-patient variability in treatment response, and the genetic basis of this variation remains largely unknown. Using a genome-wide CRISPR screen, we discovered the loss of DNTT as a top driver of InO resistance. Our results directly show the effects of DNTT expression on H2AX phosphorylation, DNA damage response signaling, cell cycle arrest and mitochondrial apoptotic priming, ultimately conferring leukemia resistance to InO. Screening 196 primary human B-ALL samples for InO sensitivity confirmed the impact of DNTT expression on InO response. Additionally, low DNTT expression was found in the residual leukemia blasts post InO treatment in patients enrolled in the COG trial AALL1621. Collectively, we demonstrate that loss of DNTT diminishes B-ALL sensitivity to InO in vitro and promotes leukemia cell survival during InO treatment in vivo.

overall_design: Total RNA was purified from human B-ALL cell lines (Nalm6, REH and RS4:11 with WT or DNTT knockout genotype) with or without InO treatment by using the Qiagen Rneasy Mini Kit and subsquently used for RNA-sequencing.

first: Jun

last: Yang


title: The GATA-3 dependent transcriptome and tumor microenvironment are regulated by eIF4E and XPO1 in T-cell lymphomas

city: Ann Arbor

first: Chenguang

last: Wang

state: Michigan

address: 1500E Medical Center Dr, Room 4411

department: Department of Internal Medicine

institute: University of Michigan

summary: The transcription factor GATA-3 and the transcriptional program it regulates have emerged as oncogenic drivers across diverse T-cell lymphomas (TCL), many of which are resistant to conventional chemotherapeutic agents and characterized by recurrent losses of key tumor suppressor genes, including TP53 and PTEN, both of which are clients of the nuclear export protein XPO1. Here we demonstrated that XPO1 is highly expressed by malignant T cells expressing GATA-3 and by lymphoma-associated macrophages (LAM) within their tumor microenvironment (TME). Using complementary genetically engineered mouse (GEM) models, we demonstrated that TP53 and/or PTEN deficient TCL, and LAM within their TME, are sensitive to the selective XPO1 antagonist selinexor. In an effort to identify TP53 and PTEN independent mechanisms, we used complementary and orthogonal approaches to investigate the role of eIF4e and XPO1-dependent mRNA nuclear export in these TCL. We identified a novel role for eIF4E/XPO1 in exporting GATA-3 and GATA-3-dependent transcripts from the nucleus in TCL, and in the export of therapeutically relevant transcripts, including CSF-1R, from LAM. Therefore, XPO1 antagonism, by impairing oncogenic transcriptional programs in TCL and depleting LAM from their TME, is a novel approach to target two independent dependencies in a group of therapeutically challenging TCL.

overall_design: RIP on one cutaneous T cell lymphoma cell line and one T-cell acute lymphblastic leukemia cell line, as well as monocyte-derived macrophages.

first: Ryan

last: Wilcox


title: Melanocyte and nerve fiber cross-talk enhances UV-B-induced melanogenesis in human epidermis and is facilitated by semaphorin-4A

city: Muenster

first: Markus

last: Fehrholz

address: Mendelstr. 17

institute: Monasterium Laboratory GmbH

summary: Epidermal melanocytes form synaptic-like contacts with cutaneous nerve fibers but the functional outcome of this connection remains elusive. In this pilot study, we used our fully humanized re-innervated skin organ culture model to investigate whether melanocyte-nerve fibers interaction play a role in UV-B-induced melanogenesis. UV-B-irradiation significantly enhanced melanin content and tyrosinase activity in re-innervated skin ex vivo compared to non-innervated controls, indicating that neuronal presence is essential for exacerbating pigmentation upon UV-B irradiation in long-term culture. Comparative transcriptomic analysis between laser-capture-microdissected melanocytes from freshly embedded human skin and published microarray data on in vitro primary melanocytes identified Semaphorin-4A (SEMA4A) as possible mediator of melanocyte-nerve fibers interactions. SEMA4A protein levels in Gp100+ epidermal melanocytes were significantly higher in re-innervated skin than in skin alone, and reduced by UV-B treatment. Analysis of melanocytes in vitro showed reduced SEMA4A protein expression 24h after UV-B-irradiation while SEMA4A secretion into the medium was increased. In sensory neurons, conditioned media (CM) from UV-B irradiated melanocytes stimulated tubulin expression and axon growth. Re-transfer of CM from neurons that received CM from melanocytes back to melanocytes resulted into significant increased melanin content only when the CM derived from neurons previously receiving CM from UV-B irradiated melanocytes. These findings highlight the importance of melanocyte-neuron interactions for UV-B-induced melanogenesis and suggest that secreted proteins (e.g. SEMA4A) can function as a novel target in the treatment of hypo- and hyperpigmentation disorders.

overall_design: We have used a fully re-innervated human skin explant model, with extended skin survival, which overcomes the fast degeneration of nerve fibers upon skin extraction, in which sensory nerve fibers differentiated from human pluripotent stem cell (iPSC)-derived neural progenitor cells efficiently re-innervate human skin, to examine the cross-talk between cutaneous sensory nerve fibers and epidermal melanocytes during melanogenesis upon UV-B irradiation. Re-innervated skin was compared to non-innervated skin, where the endogenous nerve fibers had degraded during long-time culture periods of 10 days. Comparative transcriptomic analysis between laser-capture-microdissected melanocytes from freshly embedded human skin and published microarray data on in vitro primary melanocytes was performed was also performed.

first: Onur

last: Egriboz

first: Moe

last: Tsutsumi

first: Marta

last: Sousa

first: Jeremy

last: Cheret

first: Wolfgang

last: Funk

first: Maximilian

last: Kückelhaus

first: Ralf

last: Paus

first: Kentaro

last: Kajiya

first: Ilaria

last: Piccini

last: Bertolini


title: Iron-induced oxidative stress alters the transcriptome of bronchoalveolar lavage cells in silicosis

city: Bundoora

first: Christian

last: Aloe

state: VIC

address: Plenty Road

country: Australia

institute: RMIT University

summary: Elevated iron levels are observed in silicosis patients, leading alveolar macrophages to increase ferritin production in response to this cellular stress. Our study employed a multi-omic approach to investigate how excess iron/ferritin influences the molecular profile of bronchoalveolar lavage (BAL) cells. Thirty-one participants were included, with RNA sequencing conducted on BAL cells from stone benchtop industry workers diagnosed with simple or complicated silicosis, compared to a cohort with no disease. BAL fluid (BALF) was analysed using gas chromatography-mass spectrometry (GC-MS) to screen for 498 volatile organic compounds (VOCs). Compared to no disease, simple silicosis showed 172 differentially expressed genes (DEGs), while complicated silicosis displayed 531 DEGs. Spearman correlation analysis revealed 1072 genes significantly positively association with BALF ferritin. Gene set variation analysis revealed upregulation in glutathione metabolism pathways in BAL cells, including ferroptosis repressor genes (GCLC/GCLM, GSR, SLC7A11, TXNRD1, and AIFM2). In the BALF, 116 VOCs were detected, with 4 showing reduced levels in silicosis patients, independent of smoking status. Notably, the abundance of these VOCs exhibited an inverse association with ferritin levels. In summary, antioxidant defence pathways, incorporating genes that encode proteins to prevent ferroptosis, an iron-dependent non-apoptotic form of cell death, are upregulated in BAL from silicosis patients. Excess iron/ferritin was also associated with a reduction in multiple VOCs present in BALF, which may reflect their oxidative metabolism. We propose that BAL macrophages loaded with iron experience oxidative stress, leading to the activation of an anti-ferroptotic gene network that prolongs the survival of this dysfunctional BAL cell population.

overall_design: To investigates how excess iron and ferritin levels affect the molecular profile of bronchoalveolar lavage (BAL) immune cells isolated from stone benchtop industry workers.

first: Steven

last: Bozinovski

first: Nicole

last: Goh

first: Tracy

last: Leong

first: Daniel

last: Chambers

last: Hoy


title: Host-derived Interleukin 1α induces immune suppressive tumor microenvironment via regulation of monocyte-to-macrophage differentiation

city: COLUMBIA

first: HAO

last: JI

address: 715 SUMTER ST CLS RM 109

department: College of Pharmacy

institute: University of South Carolina

summary: Tumor-associated macrophages exhibit high heterogeneity and contribute to the establishment of an immune-suppressive tumor microenvironment (TME). Although numerous studies have demonstrated that extracellular factors promote macrophage proliferation and polarization, the regulatory mechanisms governing the differentiation process to generate phenotypically, and functionally diverse macrophage subpopulations remain largely unexplored. In this study, we examined the influence of interleukin 1α (IL-1α) on the development of an immune-suppressive TME using syngeneic transplantation murine models of breast cancer. Deletion of IL-1α led to the rejection of syngeneic tumor challenges. Single-cell sequencing analysis revealed that Cx3cr1+ macrophage cells were the primary sources of IL-1α in the TME. The absence of IL-1α reprogrammed the monocyte-to-macrophage differentiation process within the TME, characterized by a notable decrease in the subset of Cx3cr+ ductal-like macrophages and an increase in iNos-expressing stress-responsive cells. Comparative analysis of gene signatures in both human and mouse macrophage subsets suggested that IL-1α deficiency shifted macrophage polarization from M2 to M1 phenotypes, leading to enhanced cytotoxic T lymphocyte activity in the TME. Importantly, elevated levels of IL-1α in human cancers were associated with improved prognosis following immunotherapy. These findings underscore the pivotal role of IL-1α in shaping an immune-suppressive TME through the regulation of macrophage differentiation and activity, highlighting IL-1α as a potential target for breast cancer immunotherapy.

overall_design: Freshly harvested HER2+ H605 tumors from transplanted WT and IL1α-/- mice, two pairs in total, were disaggregated into single cells two weeks after transplantation and subsequently subjected to single-cell sequencing.

first: Manikanda

last: Keerthi Raja

first: Hao

last: Ji

first: Hexin

last: Chen


title: Dynamic nitrosation - palmitoylation of CD36 regulates serum lipids

city: Charlottesville

first: Brant

last: Isakson

state: Virginia

department: Cardiovascular Research Center

institute: University of Virginia

summary: Regulation of endothelial cell (EC) lipid content is crucial for cell and organ function. During obesity, ECs become lipid laden leading to lipotoxicity and endothelial dysfunction which further contribute to metabolic syndrome progression. Here, we demonstrate a novel pathway by which the endothelium, via eNOS-dependent nitrosation, inhibits excess lipid accumulation during hyperlipidemic conditions in obesity. In the vasculature, nitric oxide has been reported as a potent vasodilator. However, we highlight a new role for nitric oxide as a modulator of serum lipids. We show this occurs as a result of the downregulation of Cav1, a potent negative regulator of endothelial nitric oxide synthase, increasing EC endogenous nitric oxide synthesis. Using EC-specific Cav1 knockout mice, we are able to increase nitric oxide in vivo. This increased nitric oxide leads to nitrosation of cysteines 3 and 466 on the cytoplasmic tails of CD36, a fatty acid translocase, disrupting palmitoylation of these residues and subsequently inhibiting trafficking of CD36 to the plasma membrane. Together, this work suggests that CD36 nitrosation occurs as a protective mechanism to prevent lipotoxicity and EC dysfunction during the progression of metabolic syndrome.

overall_design: Sixteen hours prior to assessment HAMECs were treated with 1mM L-NAME and subsequently four hours prior to assessment HAMECs were treated with 50 μM lipids as well as 50 μM DETA NONOate. RNA was isolated using Quick-RNA MiniPrep Kit (Genesee Scientific: 11-328). When necessary, samples were further concentrated and purified with the RNeasy MinElute Cleanup Kit (Qiagen: 74804). PE150 reads were generated with the Illumina NovoSeq platform. The paired-end reads were mapped to the hg38 reference genome using STAR (v 2.7.9a). Gene counts were generated with FeatureCounts in the Rsubread (v. 2.8.8) package. DESeq2 (v. 1.34.0) was used to calculate log fold change and adjusted p-values. The adjusted p-values were then adjusted for multiple comparisons between groups by the Benjamini-Hochberg procedure using p.adjust(). Ensemble IDs were converted to gene symbols using the EnsDb.Hsapiens.v75 and org.Hs.eg.db packages.

first: Luse

last: Melissa

first: Dunaway

last: Luke

first: Isakson

last: Brant


title: DDIT3 deficiency ameliorates systemic lupus erythematosus by regulating B-cell activation and differentiation

city: Taiyuan City

first: Yukai

last: Jing

state: Shanxi

address: 99 Longcheng Street

department: Department of Clinical Laboratory

institute: Third Hospital of Shanxi Medical University

summary: Systemic lupus erythematosus (SLE) is characterized by the excessive production of autoantibodies, and B cells are considered to be the primary cells involved in the development of SLE. Recent studies have shown that DNA damage responses play a crucial role in B cell activity in SLE, and DNA damage induced transcript 3 (DDIT3) is involved in the innate immune response. However, the exact role of DDIT3 in humoral immune response and the pathogenesis of SLE is still unknown. In this study, we aim to investigate the regulatory role of DDIT3 in B-cell homeostasis and lupus pathogenesis. We observed an increased expression of DDIT3 in B cells from SLE patients, and this expression was positively correlated with disease activity. In DDIT3 deficient mice, we observed disturbances in B cell development and differentiation, as well as inhibition of B cell activation, actin cytoskeleton reorganization, and BCR signaling. Furthermore, DDIT3 deficiency resulted in a reduction in T-cell dependent humoral immune response and germinal center reaction. Mechanistically, we found that DDIT3 promotes the transcription and expression of Itgad, which enhances PI3K signaling and B cell activation. Additionally, in the BM12-induced lupus model, we found that DDIT3 deficiency attenuated lupus symptoms and decreased germinal center responses, levels of anti-dsDNA antibodies, plasmablast cell differentiation, and renal damage. In conclusion, our study reveals for the first time the regulatory role of DDIT3 in B cell homeostasis, B cell activation, BCR signaling, and B cell function. These findings provide a new potential target for intervention in the treatment of SLE.

overall_design: To investigate the transcriptional regulation of DDIT3 in B cells, we used RNA-sequencing to detect the expression of DDIT3 regulatory genes in B cells isolated from the spleen of WT and KO mice.

first: Xin

last: Dai


title: Pharmacological inhibition of SIRT1 limits the growth of ovarian granulosa cell tumors in vitro and in preclinical mouse models

city: Paris Cedex 13

first: Celine

last: Guigon

address: 4 avenue de l'Observatoire

country: France

institute: Université Paris-Cité, CNRS, Inserm, Unité de Biologie Fonctionnelle et Adaptative

summary: RNA-sequencing was used to analyze the impact of EX-527 treatment (SIRT1 inhibitor) on gene expression and molecular pathways

overall_design: RNA profiles of three granulosa cell tumor cell lines treated with vehicle or EX-527

first: Céline

first: Eloïse

last: Airaud

first: Victoria

last: Cluzet


title: Mouse extrahepatic cholangiocarcinoma study

city: Rochester

first: Jessica

last: Hohenstein

state: MN

address: 200 1st St SW

institute: Mayo Clinic

summary: We examined the transcriptome from bile duct tissue in mice with extrahepatic cholangiocarcinoma

overall_design: RNA samples were collected from mouse bile duct

first: Baoan

first: Chen

last: Wang

last: O'Brien

first: Chantal

last: McCabe


title: Exploiting the therapeutic vulnerability of IDH-mutant gliomas with zotiraciclib

city: BETHESDA

first: Zachary

last: Sergi

address: 37 CONVENT DR

department: Neuro-Oncology Branch

institute: National Cancer Institute

summary: Isocitrate dehydrogenase (IDH)-mutant gliomas have distinctive metabolic and biological traits that may render them susceptible to targeted treatments. Here, by conducting a high-throughput drug screen, we pinpointed a specific susceptibility of IDH-mutant gliomas to zotiraciclib (ZTR).  ZTR exhibited selective growth inhibition across multiple IDH-mutant glioma in vitro and in vivo models. Mechanistically, ZTR at low doses suppressed CDK9 and RNA Pol II phosphorylation in IDH-mutant cells, disrupting mitochondrial function and NAD+ production, causing oxidative stress.,. Integrated biochemical profiling of ZTR kinase targets and transcriptomics unveiled that ZTR-induced bioenergetic failure was linked to the suppression of PIM kinase activity. We posit that the combination of mitochondrial dysfunction and an inability to adapt to oxidative stress resulted in significant cell death upon ZTR treatment, ultimately increasing the therapeutic vulnerability of IDH-mutant gliomas. These findings prompted a clinical trial evaluating ZTR in IDH-mutant gliomas towards precision medicine (NCT05588141).

overall_design: Comparative gene expression profiling analysis of RNA-seq data for GSC923 & TS603 cells treated with zotiraciclib or untreated at 24 or 48 hours. Each group was comprised of 3 biological replicates.

first: Zach


title: Aberrant lipid metabolism in macrophages is associated with granuloma formation in sarcoidosis

city: Vienna

first: Georg

last: Stary

address: Spitalgasse 23

country: Austria

institute: Medical University of Vienna

summary: Rationale: Chronic sarcoidosis is a complex granulomatous disease with limited treatment options that can progress over time. Understanding the molecular pathways contributing to disease would aid in new therapeutic development.  Objectives: To understand if macrophages from non-resolving chronic sarcoidosis patients are predisposed to macrophage aggregation and granuloma formation, and if modulation of the underlying molecular pathways influence sarcoidosis granuloma formation.  Methods: Macrophages were cultivated in vitro from isolated peripheral blood CD14+ monocytes and evaluated for spontaneous aggregation. Transcriptomics analyses, phenotypic and drug inhibitory experiments were performed on these monocyte-derived macrophages. Human skin biopsies from sarcoidosis patients and a myeloid Tsc2-specific sarcoidosis mouse model were analyzed for validatory experiments. Measurements and Main Results: Monocyte-derived macrophages from chronic sarcoidosis patients spontaneously formed extensive granulomas in vitro compared to healthy controls. Transcriptomic analyses separated healthy and sarcoidosis macrophages and identified an enrichment in lipid metabolic processes. In vitro patient granulomas, sarcoidosis mouse model granulomas, and those directly analyzed from lesional patient skin expressed an aberrant lipid metabolism profile and contained increased neutral lipids. Conversely, a combination of statins and cholesterol-reducing agents reduced granuloma formation both in vitro and in vivo in a sarcoidosis mouse model.  Conclusions: Together, our findings show that altered lipid metabolism in sarcoidosis macrophages is associated with its predisposition to granuloma formation and suggest cholesterol-reducing therapies as a treatment option in patients.

overall_design: CD14-isolated monocytes from 5 chronic sarcoidosis patients and 4 healthy controls were differentiated with 500 IU/mL rhGM-CSF for 3 days. Up to 1.2 x106 CD14+ monocytes or monocyte-derived macrophages from patients and healthy controls were carefully washed, harvested and lysed in 700 ul Qiazol (Qiagen) and stored at -80°C until RNA isolation. CD14+ cells were lysed on the day of sample collection, whereas macrophages were harvested after 3 days of differentiation with GM-CSF. RNA was isolated from the aforementioned samples using the miRNeasy Kit (Qiagen) and concentration and purity assessed using a Nanodrop 2000c Spectrophotometer (Thermo Scientific). RNA quality of the samples was further assessed on the Agilent Bioanalyzer RNA 6000 Nano Electropherogram, and library preparation performed on samples with RIN quality between 8-10. 100ng of total RNA was used for library preparation using the NEBNext Ultra II directional RNA library preparation kit with poly-A tail enrichment. Thereafter, stranded mRNA 75 bp single read RNA sequencing was performed on the Illumina NextSeq 500 sequencer.

first: Ram

middle: V

last: Pandey

first: Clarice

last: Lim

first: Anna

last: Redl

first: Thomas

last: Weichhart


title: First-in-class humanized antibody against alternatively spliced tissue factor augments anti-metastatic efficacy of chemotherapy in a preclinical model of pancreatic ductal adenocarcinoma

city: Cincinnati

first: Xiang

last: Zhang

state: OH

address: 160 Panzeca Way, Kettering Room 336

department: Environmental and Public Health Sciences

institute: University of Cincinnati

summary: Alternatively spliced tissue factor (asTF) promotes progression of pancreatic ductal adenocarcinoma (PDAC) by activating beta1-integrins on PDAC cell surfaces. hRabMab1, a first-in-class, humanized, inhibitory, anti-asTF antibody we recently developed, can suppress PDAC primary tumor growth as a single agent. Whether hRabMab1 has the potential to suppress metastases in PDAC is unknown. Study results. When administered alone, hRabMab1 achieved only marginal penetration of experimental tumor tissue; however, hRabMab1 was abundant in tumor tissue when co-administered with gem/PTX which resulted in a significant decrease in tumor cell proliferation; leukocyte infiltration; and neovascularization. Gem/PTX alone reduced primary tumor volume, but not metastatic spread; only the combination of hRabMab1 and gem/PTX significantly reduced metastatic spread. RNAseq analysis of primary tumors showed that the addition of hRabMab1 to gem/paclitaxel enhanced the downregulation of tubulin binding and microtubule motor activity. In the liver, hRabMab1 reduced liver metastasis as a single agent. Only the combination of hRabMab1 and gem/PTX eliminated tumor cell-induced leukocytosis. Conclusions. This study demonstrates that hRabMab1 may help suppress metastasis in PDAC.

overall_design: We used KRAS mutant human PDAC cell line PaCa44 to generate aggressive primary orthotopic tumors. Experimental design featured orthotopic tumors formed by implantation of luciferase-labeled PaCa44 cells into the pancreata of mice. Four groups of mice were treated with vehicle; hRabMab1 alone; gemcitabine/paclitaxel (gem/PTX) alone; and the combination of hRabMab1 and gem/PTX).

first: Clayton

last: Lewis

first: Vladimir

last: Bogdanov


title: YAP1 regulates thrombopoiesis by binding to MYH9 in immune thrombocytopenia [scRNA-seq]

city: Suzhou

first: Shuhong

last: Hu

address: No.188 Shizi Street

institute: The First Affiliated Hospital of Soochow University

summary: Immune thrombocytopenia (ITP) is a complicated bleeding disease characterized by sharp platelet reduction. As a dominating element involved in ITP, megakaryocytes (MKs) are responsible for thrombopoiesis. However, the mechanism behind the dysregulation of thrombopoiesis in ITP remains undetermined. In this study, we examined the role of YAP1, Yes-associated protein 1, in thrombopoiesis during ITP. We observed a reduced YAP1 expression with cytoskeletal actin misalignment in MKs of ITP patients. Using experimental ITP mice, we showed that YAP1 reduction induced aberrant MK distribution, reduced the percentage of late MKs in total MKs, and caused submaximal platelet recovery. Mechanistically, YAP1 upregulation by GATA1 binding to its promoter promoted MK maturation. Phosphorylated YAP1 facilitated cytoskeletal activation by binding of its WW2 domain to MYH9, facilitating thrombopoiesis. Targeting YAP1 by its activator XMU-MP-1 was sufficient to rescue cytoskeleton defects and thrombopoiesis in YAP1+/- ITP mice and ITP patients. Taken together, these results demonstrate a crucial role for YAP1 in thrombopoiesis, providing a potential for the development of diagnostic markers and therapeutic options for ITP.

overall_design: Compare percentage of cells and gene expression profiling analysis using single-cell RNA sequencing data obtained from BM of WT, YAP1 mice.

first: Yifei

last: liu

first: Depei

last: Wu

first: Yang

last: Xu


title: EBV infection alters DNA methylation in primary human colon cells: a path to inflammation and carcinogenesis?

city: Rome

first: Probiomics

state: RM

address: via Montpellier 1

institute: Probiomics s.r.l

type: Methylation profiling by high throughput sequencing

summary: EBV is associated to several types of human cancers and changes in DNA methylation are reported to contribute to viral-driven carcinogenesis, particularly in cancers of epithelial origin. In previous study, we have shown that EBV infects human primary colonic cells (HCoEpC) and replicates in these cells, inducing pro-inflammatory and pro-tumorigenic effects, mostly prevented by inhibiting viral replication by PAA. Interestingly, the EBV-induced effects correlated with DNMT1 upregulation and were counteracted by pretreating cells by 5-AZA, suggesting a role for DNA hypermethylation. Based on this background, in this study, we investigated the methylation changes induced by EBV infection in HCoEpC, in the presence or in the absence of PAA or ERK1/2 and STAT3 inhibitors, pathways known to be activated by EBV and involved in dysregulating methylation in tumor cells. The genome wide-methylation analysis performed in this study allowed us to identify several biological processes and genes affected by this epigenetic change, as a possible underlying mechanism leading to the pathologic effects induced by EBV previously observed. In particular, we found that the virus induced intense methylation changes, that hypermethylation was prevalent over hypomethylation, and that several genes involved in embryogenesis and carcinogenesis and inflammation were affected.

overall_design: Primary human colonic epithelial cells (HCoEpC) were infected with EBV virions isolated by B95–8 cells (V), pretreated with PD0325901 and AG490 before infection (PD-AG-V), pretreated with phosphonoacetic acid (PAA-V), and treated with these compound in the absence of infection (PD-AG and PAA) or left untreated and uninfected as negative control (CT). Genomic DNA was purified using the PureLink genomic DNA mini kit and sheared to ~6 kb fragment size using Covaris G tubes. Nanopore sequencing libraries were prepared using the native library prep kit SQK-LSK110 and sequenced with R9.4.1 MinION flow cells (FLO-MIN106).

first: Mara

last: Cirore

first: Roberta

last: Santarelli

first: Giuseppe

last: Pascucci


title: Examining differentially expressed genes and pathways within the striatum using a rodent model of DYT-TOR1A dystonia

city: Würzburg

first: Chi Wang

last: Ip

address: Josef-Schneider-Str. 11

department: Department of Neurology

institute: University Hospital Würzburg

summary: DYT-TOR1A (DYT1) is the most common monogenetic form of dystonia characterized by a reduced disease penetrance of 30-40% only. Environmental factors are suspected to play a major role in eliciting dystonia in DYT1 gene carriers. In line with this observation a sciatic nerve crush injury as a traumatic environmental stimulus was previously shown to induce dystonia-like movements (DLM) in the asymptomatic DYT-TOR1A rat model (∆ETorA). The primary objective of this study was to assess the underlying pathophysiological mechanisms associated with DYT1 dystonia using this “second hit” DYT1 rodent model. The striatum, as one of the pathophysiologic key structures of dystonia, was analysed twelve weeks after nerve injury by unbiased RNA-sequencing in order to identify differentially expressed genes (DEG) and pathways.

overall_design: Unbiased RNA-sequencing was conducted in the striatum of DYT-TOR1A rats and wt rats with the aim of discovering differentially expressed genes (DEGs) and pathways

first: Susanne

last: Knorr

first: Chi

middle: W


title: Integrated HPV18 acts as enhancer-like RNA and form phase-separated condensates to promote oncogenic genes expression.

city: Yokohama

first: Hidefumi

last: Suzuki

address: Fukuura 3-9

institute: Yokohama City University

summary: Most of the cervical cancers are caused by human papillomavirus (HPV) infection. It has been known that, in HeLa cells, HPV18 viral genome is integrated at chromosome 8q24.21 and activates transcription of proto-oncogene c-Myc. However, the mechanism of how the integrated HPV fragment exhibits transcription activation function has not been fully elucidated. In this study, we found that HPV18 transcripts themselves have an enhancer RNA-like function to activate their proximal genes including CCAT1-5L and c-Myc. We showed that the human genome-integrated HPV18 genes are activated by transcription coregulators BRD4, Mediator. The transcribed HPV18 RNAs form a liquid-like condensate at chromosome 8q24.21 locus, which in turn accumulates RNA polymerase II. Moreover, we identified uncharacterized transcript from Upstream Region of CCAT1-5L, named URC. The URC RNA is transcribed as a chimera RNA with HPV18 and is composed of 3'-untranslated region of HPV18 transcript. We found that the URC contributes to stabilization of HPV18 RNAs by supplying a poly-adenylation site for HPV18 transcript. Our findings suggest that HPV18 integration at 8q24.21 locus causes HPV18-URC chimera RNA and promotes tumorigenesis through RNA-based liquid droplet formation.

overall_design: Experiment 1: 4sU-labeled RNA-seq prepaerd from HeLa cells upon treatment with 1,6-hexanediol and non-treat using D. melanogastor S2 cells as spike-in control. Experiment 2: 4sU-labeled RNA-seq in control and URC LNA-transfected HeLa cells.

first: Kazuki

last: Furugori

first: Ryota

last: Abe

first: Hidehisa

last: Takahashi

first: Atsushi

last: Toyoda


title: Transgelin 2 guards T cell lipid metabolism and anti-tumor function

city: New York

first: SUNGMIN

last: HWANG

state: New York

address: 1300 York Ave

institute: Weill Cornell Medicine

summary:
Mounting effective immunity against pathogens and tumors relies on the successful metabolic programming of T cells by extracellular fatty acids. Fatty acid-binding protein 5 (FABP5) plays a key role in this process by coordinating the efficient import and trafficking of lipids that fuel mitochondrial respiration to sustain the bioenergetic requirements of protective CD8+ T cells. Importantly, however, the mechanisms governing this immunometabolic axis remain unexplored. Here, we report that the cytoskeletal organizer Transgelin 2 (TAGLN2) is necessary for optimal fatty acid uptake, mitochondrial respiration, and anti-cancer function in CD8+ T cells. We found that TAGLN2 interacts with FABP5, enabling its cell surface localization and function in activated CD8+ T cells. Analysis of ovarian cancer
specimens revealed that endoplasmic reticulum (ER) stress responses elicited by the tumor microenvironment repress TAGLN2 in infiltrating CD8+ T cells, thereby enforcing their dysfunctional state. Restoring TAGLN2 expression in ER-stressed CD8+ T cells bolstered their lipid uptake, mitochondrial respiration, and cytotoxic capacity. Accordingly, chimeric antigen receptor T cells overexpressing TAGLN2 bypassed the detrimental effects of tumor-induced ER stress and demonstrated superior therapeutic efficacy in mice with metastatic ovarian cancer. Our study unveils the role of cytoskeletal TAGLN2 in T cell lipid metabolism and highlights the potential to enhance cellular immunotherapy in solid malignancies by preserving the TAGLN2-FABP5 axis.

overall_design: Profiling of single-cell transcriptome of total tumor-infiltrating CD45+CD3+ T cells sorted from Xbp1fl/fl or Xbp1fl/flCd4Cre mice bearing metastatic OvCa.

first: Sung-Min

last: Hwang

first: Deepika

last: Awasthi

first: Jieun

last: Jeong

first: Tito

last: Sandoval

first: Chang-Suk

last: Chae

first: Yusibeska

last: Ramos

first: Chen

last: Tan

first: Matías

middle: M

last: Falco

first: Camilla

last: Salvagno

first: Alexander

last: Emmanuelli

first: Ian

last: McBain

first: Bikash

last: Mishra

first: Lionel

middle: B

last: Ivashkiv

first: Dmitriy

last: Zamarin

first: Evelyn

last: Cantillo

first: Eloise

last: Chapman-Davis

first: Kevin

last: Holcomb

first: Diana

last: Morales

first: Xiaoqing

last: Yu

first: Paulo

last: Rodriguez

first: Jose

last: Conejo-Garcia

first: Martin

last: Kaczocha

first: Anna

last: Vähärautio

first: Minkyung

last: Song

first: Juan

last: Cubillos-Ruiz