title: Mediator Subunit 30 Directs Myc Oncogenesis [RNA-seq]

overall_design: RNA seq in Mia PaCa-2 was performed with 0.5ug/ml doxycycline-induced teton shRNA or cDNA overexpression, or pLKRO.1 shRNA knockdown directly. RNAseq in GSC3565 was performed in pLKO.1 shRNA knockdown. All shRNAs were packaged in lentivirus and infected into cells. RNA was extracted by Trizol (Life Technologies) RNA concentration and quality was quantified by Qubit and Agilent 2100 Bioanalyzer system, respectively. Libraries were generated using NEBNext Ultra II RNA Kits (NEB) and sequenced on the NovaSeq 6000 with 100 bp paired-end reads.


title: Novel Bioinformatic Analyses of Somatic Cell Contamination in Sperm Samples

status: Public on Nov 07 2023

city: Provo

first: Timothy

last: Jenkins

state: UT

address: LSB

department: Cell Biology

country: USA

institute: Brigham Young University

summary:
To establish contamination profiles, the sperm donors with normal sperm counts were analyzed using an Infinium HumanMethylation450 array. Somatic cell lysis, sperm isolation, DNA extraction, and bisulfite conversion were performed as described by Aston et al. The bisulfite converted sperm DNA was hybridized to Illumina Infinium HumanMethylation450K microarrays at the University of Utah and run as recommended by the manufacturer (Bibikova et al. 2011). Unpaired blood samples were extracted using Qiagen's DNeasy Blood and Tissue kit and bisulfite converted using Zymo's EZ DNA Methylation kit. All procedures were performed according to the instructions of the manufacturer.
Four permutations were run on each sample, including pure blood, half blood and half sperm by DNA concentration, half blood and half sperm by cell count, and pure sperm (n = 16). Concentration was normalized using a spectrophotometer. A Makler cell counting chamber was used to count white blood cells and sperm, which were then normalized in a 1:1 ratio.

overall_design: Bisulphite converted DNA from the 16 samples were hybridised to the Illumina Infinium450 Human Methylation Beadchip

first: Ki

last: Aston


title: A molecularly defined and spatially resolved cell atlas of the whole mouse brain [Bulk RNA sequencing]

city: Cambridge

first: Won

last: Jung

state: Massachusetts

address: 12 Oxford Street, NAITO 032D

department: CCB

institute: Harvard University

type: Expression profiling by high throughput sequencing

summary: In mammalian brains, millions to billions of cells form complex interaction networks to enable a wide range of functions. The enormous diversity and intricate organization of cells have impeded our understanding of the molecular and cellular basis of brain function. Recent advances in spatially resolved single-cell transcriptomics have enabled systematic mapping of the spatial organization of molecularly defined cell types in complex tissues. However, these approaches have only been applied to a few brain regions and a comprehensive cell atlas of the whole brain is still missing. Here, we imaged a panel of >1,100 genes in ~10 million cells across the entire adult mouse brain using multiplexed error-robust fluorescence in situ hybridization (MERFISH) and performed spatially resolved, single-cell expression profiling at the whole-transcriptome scale by integrating MERFISH and single-cell RNA-sequencing (scRNA-seq) data. Using this approach, we generated a comprehensive cell atlas of >5,000 transcriptionally distinct cell clusters, belonging to >300 major cell types, in the whole mouse brain with high molecular and spatial resolution. Registration of this atlas to the mouse brain common coordinate framework (CCF) allowed systematic quantifications of the cell-type composition and organization in individual brain regions. We further identified spatial modules characterized by distinct cell-type compositions and spatial gradients featuring gradual changes of cells. Finally, this high-resolution spatial map of cells, each with a transcriptome-wide expression profile, allowed us to infer cell-type-specific interactions between several hundred cell-type pairs and predict molecular (ligand-receptor) basis and functional implications of these cell-cell interactions. These results provide rich insights into the molecular and cellular architecture of the brain and a foundation for future functional investigations of neural circuits and their dysfunction in diseases.

overall_design: RNA expression levels of individual genes from three different whole mouse brains aged 56-62 days were extracted using RNAqueous Micro total RNA isolation kit. RNA quality was assessed using Agilent TapeStation and samples with an RNA integrity score >8 were kept for sequencing. RNA sequencing libraries were constructed using the Kapa mRNA HyperPrep Kits and were sequenced using the Illumina NextSeq500 platform performed by the Bauer Center Sequencing Core at Harvard University.

first: Meng

last: Zhang

first: Xingjie

last: Pan

first: Aaron

last: Halpern

first: Stephen

last: Eichhorn

first: Zhiyun

last: Lei

first: Limor

last: Cohen

first: Kimberly

last: Smith

first: Bosiljka

last: Tasic

first: Zizhen

last: Yao

first: Hongkui

last: Zeng

first: Xiaowei

last: Zhuang


title: Mediator Subunit 30 Directs Myc Oncogenesis [ChIP-seq]

status: Public on Nov 08 2023

city: La Jolla

first: Guofeng

last: Zhao

address: 9500 Gilman Dr.

department: Medicine

institute: UCSD

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: To map the binding sites of Mediator component MED17, MED30 and MYC, these three factors ChIP-seq were performed in Mia PaCa-2 cells in normal culture condition. To test the effect of MED30 deletion in Myc's binding. MYC ChIP-seq was perform with or without three days 0.5mg/ml Dox treatment in teton shMED30 Mia PaCa-2 cells. 1-5 million cells were dual cross-linked with 2 mM disuccinimidyl glutarate (DSG, ProteoChem, #C1104-1GM) for 20min at room temperature and followed by 1% formaldehyde for 10 min for Mediator’s ChIP. For transcription factors’ ChIP, cells were only fixed in 1% formaldehyde for 10 min. In both situations, the cross-linking was quenched with 0.125M glycine for 5 min. Chromatin was fragmented using a Bioruptor to get 200-500bp fragments. Subsequently, the soluble chromatin was incubated with 1-3 mg antibodies at 4°C overnight. Immunoprecipitated complexes were collected using 20 ul Protein G Dynabeads (Life Technologies) per reaction. After washing, the protein-DNA complexes were eluted and de-crosslinked overnight at 65°C. and DNA was purified with QIAquick PCR Purification Kit (Qiagen, # 28104).

first: Chunyu

last: Jin

first: Kenneth

last: Ohgi

first: Michael

middle: G

last: Rosenfeld


title: Effect of activated TAZ expression on mammary epithelial cells

city: Worcester

first: Jennifer

last: Benanti

state: MA

address: 364 Plantation St. LRB525

department: Molecular, Cell & Cancer Biology

institute: UMass Medical School

summary: TAZ is a transcriptional effector of hippo pathway. Expression of activated form of TAZ (4SA TAZ) results in significant alteration of gene expression as measured by RNA sequencing. In addition to the already established targets, our analysis identified several unique proteins, which are differentially regulated upon expression of activated TAZ.

overall_design: HMT-S1 cells were transfected with vector or 4SA-TAZ and RNA sequencing was performed. Samples were analyzed in duplicate.

first: HIRA

middle: L

last: GOEL

first: ARTHUR

middle: M

last: MERCURIO


title: Transcriptome analysis reveals key dysregulated transcriptional programmes in uveal melanoma

city: Wenzhou

first: Meng

last: Zhou

address: 270 Xueyuan Xi Road

country: China

institute: Wenzhou Medical University

summary: In this study, we conducted  whole-transcriptome sequencing in five primary Uveal Melanoma and four adjacent tissues to uncover key dysregulated the gene-regulatory circuits in Uveal Melanoma.

overall_design: The present study was performed according to the Declaration of Helsinki and approved by the Ethics Committee of the Eye Hospital of Wenzhou Medical University (ethics approval no. 2022-043-K-28-02). All patients provided written informed consent. All data were anonymously analyzed. Human UM tissues were collected from 7 patients diagnosed with UM during surgical resection in the Eye Hospital of Wenzhou Medical University. The patients did not receive preoperative treatments prior to collection.  Total RNA from UM tissues was isolated using TRIzol Reagent (Invitrogen; Thermo Fisher Scientific, Inc.). The OD260/OD280 ratio was used as an indicator of RNA purity. The ratio is required to be close to 2.0 for pure RNA (acceptable range, 1.8-2.1). RNA concentration and purity was measured using the NanoDrop 2000 Spectrophotometer (Thermo Fisher Scientific, Inc.).

first: Lanfang

last: Sun

first: Wencan

last: Wu


title: Effect of RFX1 on gene expression and the potential binding target genes in LPS-stimulated macrophages [RNA-seq]

city: Changsha

first: Shuang

last: Yang

address: No.139，Renmin Road Central Changsha-Hunan-China

institute: The Second Xiangya Hospital of central south university

summary:
To investigate the function of RFX1 in the regulation of macrophage polarization, we overexpressed RFX1 in mouse peritoneal macrophages(PMAs) or human CD14+ derived macrophage(hMDMs) by  lentivirus infections or reduce RFX1 expression in PMAs from mice with Rfx1 deficiency(CKO) .
We then performed gene expression profiling analysis using data obtained from RNA-seq of 4 different cells at two time points.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Comparative gene expression profiling analysis of RNA-seq data for PMAs or hMDMs cells and its  derivatives (pLV-RFX1, pLV-Rfx1, Rfx1 konckout) or chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for RFX1  in hMDMs cells.

first: Shaung

first: Pei

last: Du


title: ChIP-seq for RFX1

summary:
Current proteomic approaches disassemble and digest nucleosome particles, blurring readouts of the ‘histone code’. To preserve nucleosome-level information, we developed Nuc-MS, which displays the landscape of histone variants and their post-translational modifications (PTMs) in a single mass spectrum. Combined with immunoprecipitation, Nuc-MS quantified nucleosome co-occupancy of histone H3.3 with variant H2A.Z (sixfold over bulk) and the co-occurrence of oncogenic H3.3K27M with euchromatic marks (for example, a >15-fold enrichment of dimethylated H3K79me2). Nuc-MS is highly concordant with chromatin immunoprecipitation-sequencing (ChIP-seq) and offers a new readout of nucleosome-level biology.
Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for RFX1  in hMDMs cells.

overall_design: Shuang Yang


title: Loss of Nkd1 is dominant over loss of Axin2 in regulating Wnt signaling.

city: Guelph

first: Terence

last: Van Raay

address: 50 stone road east

department: MCB

country: Canada

institute: University of Guelph

summary: Wnt signaling is a crucial developmental pathway involved in early development as well as stem cell maintenance in adults and its misregulation leads to numerous diseases. Thus, understanding the regulation of this pathway becomes vitally important. Axin2 and Nkd1 are widely utilized negative feedback regulators in Wnt signaling where Axin2 functions to destabilize cytoplasmic β-catenin, and Nkd1 functions to inhibit the nuclear localization of β-catenin. Here, we set out to further understand how Axin2 and Nkd1 regulate Wnt signaling by creating axin2-/-, nkd1-/- single mutants and axin2-/-;nkd1-/- double mutant zebrafish using sgRNA/Cas9. All three Wnt regulator mutants were viable and had impaired heart looping, neuromast migration defects, and behavior abnormalities in common, but there were no signs of synergy in the axin2-/-;nkd1-/- double mutants. Further, Wnt target gene expression by qRT-PCR, and RNA-seq analysis and protein expression by mass spec demonstrated that the double axin2-/-;nkd1-/- mutant resembled the nkd1-/- phenotype demonstrating that Axin functions upstream of Nkd1 and that loss of Nkd1 is dominant over the loss of Axin2. In support of this, the data further demonstrates that Axin2 uniquely alters the properties of β-catenin-dependent transcription having novel readouts of Wnt activity compared to nkd1-/- or the axin2-/-;nkd1-/- double mutant. We also tested the sensitivity of the Wnt regulator mutants to exacerbated Wnt signaling, where the single mutants displayed characteristic heightened Wnt sensitivity, resulting in an eyeless phenotype. Surprisingly, this phenotype was rescued in the double mutant, where we speculate that cross-talk between Wnt/β-catenin and Wnt/Planar Cell Polarity pathways could lead to altered Wnt signaling in some scenarios. Collectively, the data emphasizes both the commonality and the complexity in the feedback regulation of Wnt signaling.

overall_design: To investigate Axin2 and Nkd1 regulation of Wnt signaling, axin2-/- and nkd1-/- single knockout zebrafish were created using sgRNA/Cas9. These were crossed together to create axin2-/-;nkd1-/- double knockout zebrafish. The nkd1-/- zebrafish were made in the Tu genetic background and the axin2-/- were made in the TL genetic background. The RNA-seq samples were collected at 50% epiboly with biological replicates coming from the same parents but collected on different days. Gene expression analysis of the RNA-seq data was used to determine that the nkd1-/- single mutant was similar to axin2-/-;nkd1-/- double mutant. Furthermore, gene expression analysis was used to assess Wnt target gene expression in the mutants

first: Ian

last: Bell


title: PRMT1 inhibition activates the interferon pathway to potentiate antitumor immunity and enhance checkpoint blockade efficacy in melanoma

city: Hangzhou

first: Liyuan

state: 杭州市

address: 国科大杭州高等研究院

institute: University of Chinese Academy of Sciences

summary: Despite the immense success of immune checkpoint blockade (ICB) in cancer treatment, many tumors, including melanoma, exhibit innate or adaptive resistance. Tumor-intrinsic T-cell deficiency and T-cell dysfunction have been identified as essential factors in the emergence of ICB resistance. Here, we found that protein arginine methyl transferase 1 (PRMT1) expression was inversely correlated with the number and activity of CD8+ T cells within melanoma specimen. PRMT1 deficiency or inhibition with DCPT1061 significantly restrained refractory melanoma growth and increased intratumoral CD8+T cells in vivo. Moreover, PRMT1 deletion in melanoma cells facilitated formation of double-stranded RNA (dsRNA) derived from endogenous retroviral elements (ERVs) and stimulated an intracellular interferon response. Mechanistically, PRMT1 deficiency repressed the expression of DNA methyltransferase 1 (DNMT1) by attenuating modification of H4R3me2a and H3K27ac at enhancer regions of DNMT1, and DNMT1 downregulation consequently activated ERV transcription and the interferon signaling. Importantly, PRMT1 inhibition with DCPT1061 synergized with PD-1 blockade to suppress tumor progression and increase the proportion of CD8+T cells as well as IFNγ+CD8+T cells in vivo. Together, these results reveal an unrecognized role and mechanism of PRMT1 in regulating antitumor T-cell immunity, suggesting PRMT1 inhibition as a potent strategy to increase the efficacy of ICB.

overall_design: RNA-seq of control(CTRL), PRMT1-knockout B16-F10 cells(PRMT1 sg1) and control(shNC),PRMT1-konckdown A375 cells . ChIP-seq of H3K27 acetylation for control(CTRL) or  PRMT1-knockout(PRMT1 sg1) B16-F10 cells.

first: Hongru

last: Tao

first: Chen

first: Zhenzhong

last: Deng

first: Xiao

last: Li

first: Wenzhen

last: Dang

first: Shijie

last: Fan

first: Bing

first: Fei

last: Ye

first: JunYan

last: Lu

first: Xiangqian

last: Kong

first: Chuanpeng

last: Liu

first: Cheng

last: Luo

first: Yuanyuan

last: Zhang


title: Liver Transcriptomics from Male and Female HET3 Mice on Control, Low Amino Acid and Low Isoleucine Diets when started in Young and Middle Aged Mice

city: Madison

first: Cara

middle: Leanne

last: Green

state: Wisconsin

address: 2214 Kendall Ave

institute: University of Wisconsin-Madisin

summary: Mice were put on either a control, low amino acid or low isoleucine diet at 9 weeks of age or 6 months of age and their liver transcriptomic expression was measured after 14 weeks and 18 months respectively on diet after an overnight fast. This was to discover the impact of sex, protein and isoleucine level on the metabolism of the liver.

overall_design: To compare the metabolic effect of total amino acid and just isoleucine restriction in hetergeneous mice on different sexes and ages, 9-week-old or 6-month-old mice were placed on either a Control, Low Amino Acid or Low Isoleucine diets for 13 weeks. All mice were maintained at a temperature of approximately 22°C. Mice were euthanized by cervical dislocation and livers were harvested and snap frozen in liquid nitrogen following overnight (16-hr) fasting.


title: Bisulfite-sequencing of HBV 1.1 mer-transfected huh7 cells

city: Wuhan

first: Shuang

last: Ding

address: 262 Jinlong Street

institute: Wuhan Institute of Virology

type: Methylation profiling by high throughput sequencing

summary: RNA chemical modifications have been found to play important biological functions. Among which, the 5-methylcytosine (m5C) modification has been reported to participate in viral replication through affecting RNA processing, such as export, decay, translation and so on. In this study, we performed bisulfite sequencing (BS-seq) on HBV 1.1-mer-transfected huh7 cells to identify the m5C sites in HBV mRNA and their function in virus replication was verified. To investigate the mechanism by which m5C methyltransferase NSUN2 suppresses HBV replication, altered global m5C levels in host genes in HBV 1.1-mer-transfected cells were examined by BS-seq. We found that the m5C modification of genes associated with antiviral immunity changed significantly after viral infection. Our study provide new molecular insights into the mechanism of HBV-mediated IFN inhibition

overall_design: The changes of m5C modification in huh7 cells after HBV transfection were investigated by bisulfite sequencing.

first: Haibin

first: Fan

last: Zhu

first: Wuxiang

last: Guan


title: Mitochondrial DNA mutations drive aerobic glycolysis to enhance checkpoint blockade in melanoma

city: Glasgow

first: Payam

middle: A.

last: Gammage

address: Garscube Estate, Switchback Road

country: United Kingdom

institute: CRUK Beatson Institute

summary: The mitochondrial genome encodes essential machinery for respiration and metabolic homeostasis but is paradoxically among the most common targets of somatic mutation in the cancer genome, with truncating mutations in respiratory complex I genes being most over-represented1. While mitochondrial DNA (mtDNA) mutations have been associated with both improved and worsened prognoses in several tumour lineages1–3, whether these mutations are drivers or exert any functional effect on tumour biology remains controversial. Here we discovered that complex I-encoding mtDNA mutations are sufficient to remodel the tumour immune landscape and therapeutic resistance to immune checkpoint blockade. Using mtDNA base editing technology4 we engineered recurrent truncating mutations in the mtDNA-encoded complex I gene, Mt-Nd5, into murine models of melanoma. Mechanistically, these mutations promoted utilisation of pyruvate as a terminal electron acceptor and increased glycolytic flux driven by an over-reduced NAD pool and NADH shuttling between GAPDH and MDH1, mediating a Warburg-like metabolic shift. In turn, without modifying tumour growth, this altered cancer cell-intrinsic metabolism reshaped the tumour microenvironment promoting an anti-tumour immune response characterised by loss of resident neutrophils. This subsequently sensitised tumours bearing high mtDNA mutant heteroplasmy to immune checkpoint blockade, with phenocopy of key metabolic changes being sufficient to mediate this effect. Strikingly, patient lesions bearing >50% mtDNA mutation heteroplasmy also demonstrated a >2.5-fold improved response rate to checkpoint inhibitor blockade. Taken together these data nominate mtDNA mutations as functional regulators of cancer metabolism and tumour biology, with potential for therapeutic exploitation and treatment stratification.

overall_design: single cell RNA-seq

first: Mahnoor

last: Mahmood

first: Eric

first: Ed

last: Reznik

middle: A


overall_design: bulk RNA-Seq


title: Exosomal circRNA-MANBA mediates sorafenib resistance of hepatocellular carcinoma via miR-1290/CD109/p-STAT3 (Ser727) axis

city: Chengdu City

first: Dingzhong

last: Peng

state: Sichuan Province

address: No. 37, Guoxue Alley

institute: West China Hospital of Sichuan University

summary: Circular RNAs (circRNAs) are increasingly gaining importance and attention due to their diverse potential functions. Our study aims to explore the novel mechanisms by which exosome-contained circRNAs promote sorafenib resistance in hepatocellular carcinoma (HCC). Here we report that a circular RNA, circRNA-MANBA (a circular RNA upregulated in exosomes derived from sorafenib-resistant HCC cells), plays a significant role in sorafenib resistance in HCC. We identified that circRNA-MANBA is increased in sorafenib-resistant HCC cells, their exosomes, and tumor samples from sorafenib-treated HCC patients. Depletion of circRNA-MANBA substantially increases the cell-killing ability of sorafenib. Co-culture experiments revealed that exosomes from sorafenib-resistant HCC cells carrying large amounts of circRNA-MANBA could transmit drug resistant capacity to parental cells. Further studies revealed that circRNA-MANBA acted as ‘miRNAs sponge’ to absorb miR-1290, which prevented miR-1290 interaction with CD109 and thus upregulated STAT3 phosphorylation subsequently. Using different HCC mouse models, we demonstrated that silencing circRNA-MANBA by injection of siRNA could substantially overcome sorafenib resistance. Our study provides a proof-of-concept demonstration for a potential strategy to overcome sorafenib resistance in HCC patients by targeting circRNA-MANBA.

overall_design: Exosomes of parental HCC cells (Hep3B, PLC/PRF/5, Huh7) and sorafenib-resistant HCC cells (Hep3B-R, PLC/PRF/5-R, Huh7-R) were isolated by standard centrifugation steps. Total RNAs from exosomes of parental and sorafenib-resistant  HCC cells were  transcribed into fluorescent cRNA utilizing a random priming method (Arraystar Super RNA Labeling Kit; Arraystar). The labeled cRNAs were hybridized onto the Arraystar Human circRNA Array V2 (8x15K, Arraystar). After having washed the slides, the arrays were scanned by the Agilent Scanner G2505C.

first: Diongzhong

first: Jiong

first: Guilin

last: Nie

first: Ningyuan

last: Wen

first: Yaoqun

last: Wang

first: Xianze

last: Xiong

first: Bei

first: Nansheng

last: Cheng


title: The Enterotoxigenic Bacteroides fragilis (ETBF) in intestinal inflammation and colorectal carcinogenesis [exosomes]

city: Shanghai

first: Haoyan

last: Chen

state: Shanghai

address: 145 Shandong Road

institute: Renji Hospital

type: Non-coding RNA profiling by high throughput sequencing

summary: To elucidate whether  Enterotoxigenic Bacteroides fragilis (ETBF) plays a role in  intestinal inflammation and colorectal carcinogenesis, a RNA-seq analysis was performed to compare the microRNA profiles of  exosomes derived from ETBF-treated HCT116 colorectal cancer cell lines.

overall_design: miRNA-seq was performed in exosomes derived from HCT116 colorectal cancer cells after ETBF treatment.

first: Yingying

last: Cao


title: The Enterotoxigenic Bacteroides fragilis (ETBF) in intestinal inflammation and colorectal carcinogenesis

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

overall_design: Refer to individual Series


title: Global transcriptomics of murine splenic B cells after treatment with BAFF 3mer and 60mer in presence of a control antibody or mBaffR-mFc

city: Greenville

first: Akshaya

middle: K

last: Meher

state: North Carolina

address: MICROBIOLOGY AND IMMUNOLOGY, BRODY SCHOOL OF MEDICINE

institute: East Carolina University

summary: Mouse splenic total B cells (6 million cells/sample) were pre-treated with the control Ab (1000ng/ml, IgG1κ, BioXCell, #BE0083) or mBaffR-mFc (1000ng/ml, a fusion protein consists of murine BR3 ecto-domain fused to murine IgG1 Fc fragment obtained from Biogen, US) for 30 min, and then treated with 100 ng/ml of recombinant human BAFF 3mer (AdipoGen, #AG-40B-0016), 60mer (AdipoGen, #AG-40B-0112) or left untreated for the next 4 hours. Each treatment was in triplicates. Total RNA was extracted from the samples using QIAGEN RNeasy Mini Kit and were sent to Novogene Corporation (Wilmingtom, DE, USA) to determine the gene expression levels.

overall_design: Examination of the effect of BAFF 3mer and 60mer in B cell transcriptomics in presence or absence of a reagent that blocks BR3 binding sire on BAFF


title: Remodelling of H3K4me3 in response to long term drug treatment or oncogenic stress

city: Woolloongabba

first: Helmut

last: Schaider

state: Queensland

address: level 7, 37 Kent Street

department: Diamantina Institute

country: Australia

institute: The University Of Queensland

summary: Accumulating evidence indicates adaptive cellular reprogramming is a major contributor to disease recurrence and acquired drug resistance in cancer. Here we show that chronic drug exposure results in a multi-state metabolic and epigenetic adaption that allows cancer cells to regain proliferative capacity and eventually develop permanent drug resistance. Specifically, the escape from the previously described slow cycling induced drug tolerant cell state is driven by transiently increased expression of O-GlcNAc transferase (OGT) linking metabolic to epigenetic changes by facilitating global H3K4me3 remodeling. This adaptive process can be blocked by genetic or pharmacological inhibition of OGT or concurrent activation of AMPK to prevent metabolic adaption, subsequent epigenetic remodeling and permanent acquired drug resistance. This novel metabolic and epigenetic remodeling process is also observed following oncogene induced stress and the OGT dependence of this adaptive process uncovered promising targets for combination therapy with current standard of care drugs that could significantly increase response duration and patient outcomes.

overall_design: Experiments compare the H3K4me3 profiles of the BRAF mutant melanoma cell line WM164 without treatment or under long term exposure to sub lethal doses of the BRAF inhibitor dabrafenib. Additionally, H3K4me3 profiles of the BRAF mutant melanoma cell line WM164 overexpressing either GFP or NRAS Q61K are compared. All samples were prepared in duplicates.

first: Heinz

last: Hammerlindl

first: Gregory

last: Gimenez

first: Mike

last: Eccles


title: RNAseq of proximal small intestine scrapings of mice fed a galactose or glucose containg diet during postweaning

city: Wageningen

first: Evert

middle: M.

last: van Schothorst

address: De Elst 1

country: Netherlands

institute: Wageningen University

summary:
Background: Prolonged lactation provides substantial health benefits, possibly because of galactose as part of milk sugar lactose. Isocaloric replacement of dietary glucose [16 energy%(en%)] with galactose within a normal diet (64en% carbohydrates) during a 3-wk postweaning period provided substantial benefits on short- and long-term physiologic and metabolic parameters at the whole-body level and liver in female mice, which might be attributable to intestinal function.

Objectives: This study aimed to investigate if partial dietary replacement of glucose with galactose alters intestinal metabolism underlying hepatic health effects.

Methods: Proximal intestinal mucosa gene profiles in female mice were analyzed using RNAseq technology, validated, and correlated with hepatic health parameters.

Results: Transcriptome analysis revealed that the presence of galactose primarily affected the pathways involved in energy metabolism. A consistently higher expression was observed in the subset of mitochondrial transcripts (78 of 80, all P.adj < 0.1). Oxidative phosphorylation (OXPHOS) represented the most upregulated process (all top 10 pathways) independent of the total mitochondrial mass (P = 0.75). Moreover, galactose consistently upregulated carbohydrate metabolism pathways, specifically glycolysis till acetyl-CoA production and fructose metabolism. Also, the expression of transcripts involved in these pathways was negatively correlated with circulating serum amyloid A3 protein, a marker of hepatic inflammation [R (-0.61, -0.5), P (0.002, 0.01)]. Accordingly, CD163+ cells were decreased in the liver. Additionally, the expression of key fructolytic enzymes in the small intestinal mucosa was negatively correlated with triglyceride accumulation in the liver [R (-0.45, -0.4), P (0.03, 0.05)].

Conclusions: To our knowledge, our results show for the first time the role of galactose as an OXPHOS activator in vivo. Moreover, the concept of intestinal cells acting as the body's metabolic gatekeeper is strongly supported, as they alter substrate availability and thereby contribute to the maintenance of metabolic homeostasis, protecting other organs, as evidenced by their potential ability to shield the liver from the potentially detrimental effects of fructose.

overall_design: Nutritional intervention study for 3 weeks in young (3 weeks old, weaned) mice.

first: Ferran

last: Fos Codoner

first: Jaap

last: Keijer


title: Genome-wide Regulation of Pol II, FACT, and Spt6 Occupancies by RSC in Saccharomyces cerevisiae

city: Rochester

first: Chhabi

last: Govind

state: mi

address: 333 Science and Engineering Building

department: Biological Sciences

institute: Oakland University

summary: RSC (remodels the structure of chromatin) is an essential ATP-dependent chromatin remodeling complex in Saccharomyces cerevisiae. The catalytic subunit of RSC, Sth1 uses its ATPase activity to slide or remove nucleosomes. RSC has been shown to regulate the width of the nucleosome-depleted regions (NDRs) by sliding the flanking nucleosomes away from NDRs. As such the nucleosomes encroach NDRs when RSC is depleted and leads to transcription initiation defects. In this study, we examined the effects of the catalytic-dead Sth1 on transcription and compared them to the effects observed during acute and rapid Sth1 depletion by auxin-induced degron strategy. We found that rapid depletion of Sth1 reduces recruitment of TBP and Pol II in highly transcribed genes, as would be expected considering its role in regulating chromatin structure at promoters.  In contrast, cells harboring the catalytic-dead Sth1 exhibited a severe reduction in TBP binding, but surprisingly, also displayed a substantial accumulation in Pol II occupancies within coding regions. After depleting endogenous Sth1 in the catalytic dead mutant, we observed a further increase in Pol II occupancies, suggesting that the inactive Sth1 contributed to the observed accumulation of Pol II in coding regions. Notwithstanding the Pol II increase, the ORF occupancies of histone chaperones FACT and Spt6 were significantly reduced in the mutant. These results suggest a potential role for RSC in recruiting/retaining these chaperones in coding regions. Pol II accumulation despite substantial reductions in TBP, FACT, and Spt6 occupancies in the catalytic-dead mutant could be indicative of severe transcription elongation and termination defects. Such defects would be consistent with studies showing that RSC is recruited to coding regions in a transcription-dependent manner. Thus, these findings imply a role for RSC in transcription elongation and termination processes, in addition to its established role in transcription initiation.

overall_design: The S. cerevisiae cultures spiked in with 10% of S. pombe cells were formaldehyde crosslinked and subjected to sonication to obtain chromatin. Thus obtained chromatin was subjected to chromatin immunoprecipitation using antibodies against RNA Pol II (Pol II), TATA-binding protein, and histone chaperone Spt6 and Spt16. The ChIPed DNA was subjected to paired-end sequencing.


title: Diabetes-associated breast cancer has a distinct transcriptome and metabolome and shows DNA repair deficiency

city: Bethesda

first: Stefan

last: Ambs

state: MD

address: Building 37, Room 3050B

department: CCR

institute: NCI


title: Characterizing the function of MCUb as an inducible activator of mitochondrial oxidative phosphorylation

city: Cincinnati

first: Jeffery

middle: D

last: Molkentin

state: Ohio

address: 240 Albert Sabin Way MLC 7020

department: Molecular Cardiovascular Biology

institute: Cincinnati Children's Hospital Medical Center

summary:
Influx of Ca2+ across the inner mitochondrial membrane via the mitochondrial Ca2+ uniporter (MCU) enhances the activity of several electron transport chain dehydrogenases and the F1F0 ATP synthase.  In this study, we investigated the role of MCUb, an MCU complex gene product that is a direct inhibitor of MCU mediated Ca2+ influx. We find MCUb expression to be induced by caloric restriction in heart, skeletal muscle, liver and kidney where it functions as a metabolic activator of mitochondrial fatty acid utilization. Mice selectively deficient for Mcub in skeletal muscle showed a metabolic signature of deficient fatty acid utilization in muscle, associated with progressive age-related obesity, glucose intolerance and metabolic syndrome. To define transcriptional changes underlying, and potentially contributing to this regulation of Ca2+-dependent mitochondrial fatty acid utilization, microarray analyses of tibialis anterior (TA) muscle from control and knockout mice was carried out.
We used microarrays to elucidate effects of MCU and MCUb ablation on transcriptional profiles of TA muscle

overall_design: Tibialis anterior (TA) muscle was harvested from 2-month old control and experimental mice on the C57/BL6 background. Total RNA was isolated using the Trizol protocol and submitted to Affymetrix Clariom S array pipeline after quality control analysis using Agilent 2100 Bioanalyzer.

first: Jiuzhou

last: Huo

first: Vikram

last: Prasad


title: Abalone peptide increases stress resilience and cost-free longevity via SKN-1-governed transcriptional metabolic reprogramming in C. elegans

status: Public on Nov 06 2023

city: Guangzhou

first: Qiangqiang

last: Wang

email: wqq@whu.edu.cn, fewqq@mail.scut.edu.cn

state: Guangdong

address: Waihuan East Road, No. 381

department: School of Food Science and Engineering

institute: South China University of Technology

summary: A major goal of healthy aging is to prevent declining resilience and increasing frailty, which are associated with many chronic diseases and deterioration of stress response. We have successfully used loss-or-gain model of stress survival to quantify organismal resilience. As a proof of concept, this is demonstrated in C. elegans exposed to increasing oxidative stress induced by exogenous paraquat and with proteotoxic stress caused by endogenous polyQ and Aβ aggregation, respectively. Based on this, we reveal that abalone peptide AbaPep#07 (SETYELRK) not only promotes survival resilience against paraquat-induced oxidative stress but also confers protection against polyQ- or Aβ-mediated behavioral dysfunction in C. elegans, indicating its capacity against stress and neurodegeneration. Importantly, we further show that the peptide increases physical fitness of aged C. elegans and extends life span without compromise of growth and reproduction, demonstrating its cost-free prolongevity activity. Moreover, we reveal that the peptide induces differential expression of a number of genes, including innate immune, lipid metabolism and metabolic detoxification genes, which are associated with the SKN-1/Nrf transcription factor. Collectively, our findings demonstrate that the novel abalone cryptide AbaPep#07 increases stress survival resilience and cost-free longevity via SKN-1/Nrf-governed transcriptional reprogramming and provide an insight into the neuroprotective and health-promoting potential of cryptides as proteostasis regulators.

overall_design: To identify genes responsive to the peptide, we performed global transcriptomic analysis using RNA-seq to compare aged C. elegans (day 10 of adulthood) with and without AbaPep#07 treatment from L4 stage.

first: Liangyi

first: Zebo

last: Huang


title: Allometry of cell types in planarians by single cell transcriptomics

city: Exeter

first: Jordi

last: Solana

address: Northcote House

department: Health and Life Sciences

institute: University of Exeter

summary: We used scRNA-seq to examine cell type allometry in asexual planarians. We profiled 28,738 single cell transcriptomes of planarian of 3 different sizes categories: small (S), medium (M) and large (L). We detected changes in cell type frequency and gene expression patterns in planarian of different sizes, capturing regulatory programs of distinct cell types in response to allometric scaling.

overall_design: We obtained cell dissociations from adult populations of the asexual strain of Schmidtea mediterranea of different body sizes.

first: Elena

last: Emili

first: Alberto

last: Pérez-Posada

first: Maria

last: Christodoulou


title: FAIRE (Formaldehyde-Assisted Isolation of Regulatory Elements) analysis of chromatin accessibility in kidney bean under drought stress

city: WEST LAFAYETTE

first: Shaojun

last: Xie

state: Indiana

address: 3135 Decatur Street

institute: Purdue University

summary:
Purpose:  To identify nucleosome-depleted regions in response to drought in the kidney bean, a genome-wide chromatin profiling strategy was implemented. A better understanding of these OCRs (Open chromatin regions) will be useful in exploring potential genomic regions for crop improvement.
Results: Using a comprehensive FAIRE-Seq approach, nucleosome-free chromatin regions were explored in kidney bean (Phaseolus vulgaris. L) in response to both drought and rewatering stress. In all samples, FAIRE peaks (>50%) were enriched in the promoter regions. The drought samples showed peaks very distinct from control and rewatered samples, which showed peaks predominantly at the transcription start site (TSS). Drought and heat-responsive genes and transcription factors were identified within the flanking region of differential peaks. When drought was compared to control, it showed upregulated differential peaks primarily in the promoter and distal intergenic regions.
Conclusion: By analyzing FAIRE we are able to identify distinct genomic regions corresponding to droughts and controls. Upstream regions of genes play a significant role in drought stress response. Our data suggest that promoter regions of >2 kb, downstream regions of <= 300, and distant regulatory regions are predominant during the upregulation of drought stress. It would also be helpful to note that genomic regions vary in response to drought, and distal intergenic regions may also be involved in drought responses. Understanding plant genomes and epigenomics could help us develop varieties that can withstand extreme weather conditions.

overall_design: Phaseolus vulgaris L. (Red Hawk dark red kidney bean) was exposed to drought stress at the R2 stage, when 50% of the flowers had opened. Plants were not watered until wilting occurred. Following 10 days, young leaves from three drought and control plants were randomly collected. An additional set of samples was collected ten days after irrigation of control and drought-stressed plants. The FAIRE-Seq was carried out on three biological replicates of each sample.

first: Mayavan

last: Subramani

first: Miranda

last: Penny

first: Lily

middle: W

last: Lofton

first: Carlos

middle: U

last: Florez

first: Michael

last: Moore

first: Jyothi

last: Thimmapuram

first: Venu

last: Kalavacharla


title: CD8 Tissue Resident Memory (TRM) induction

first: David

state: Maryland

address: 9000 Rockville Pike

institute: National Cancer Institute

summary: We investigated the differentiation pattern induced by MAdCAM co stimulation on α4β7+ CD8+ T cells by analyzing their transcriptional signature.

overall_design: We stimulated freshly isolated bulk primary CD8+ T cells with combinations of anti-CD3 (OKT3), MAdCAM/retinoic acid (RA) with or whitout TGFb

first: Alexandre

last: Girard

first: Claudia

last: Cicala

first: James

last: Arthos


title: Transplanted Human Neural Stem Cells Rescue Phenotypes in zQ175 Huntington's Disease Mice and Innervate the Striatum

city: Irvine

first: Jie

last: Wu

state: CA

address: Sprague Hall 332

department: Biological Chemistry

institute: University of California, Irvine

summary: Long-term implantation of human NSCs in Huntington’s disease (HD) mice ameliorates behavioral deficits and reduces mutant Huntingtin accumulation. The human NSCs survive and differentiate into diverse neuronal populations, form connections, improve synaptic properties and show restoration of several transcriptional changes in HD mouse cells.

overall_design: Single nuclei RNAseq (snRNAseq) was performed on striatal tissue from female zQ175 mice implanted with hNSCs for 6 months to observe gene expression changes at a similar time frame as behavioral changes.

first: Ricardo

last: Miramontes

first: Ryan

last: Lim

first: Leslie

last: Thompson


title: DRG2 is required for cell surface localization of PD-L1 in melanoma cells and optimum response to anti-PD-1 immune checkpoint blockade

city: Seoul

first: Jeonghwan

last: Kim

address: 369 Sangdo-Ro, Entrepreneurship & Small Business Center 612

country: South Korea

institute: Soongsil University


title: Investigation of lncRNA expression profiles and analysis of immune-related lncRNA-miRNA-mRNA networks in neovascular age-related macular degeneration

city: Jinan

first: Gongqiang

last: Yuan

address: No. 372, Jingsi Road

institute: Eye Institute of Shandong First Medical University

summary: The aim of the present study was to investigate the alteration of lncRNAs and mRNAs in neovascular AMD. The differentially expressed mRNAs (DEmRNAs) and lncRNAs (DElncRNAs) were identified and the immune-related ceRNA network was characterized.

overall_design: Nine patients with neovascular AMD were included in the study. Control group comprised seven patients with epiretinal membranes. RNA sequencing was performed to obtain the differentially expressed mRNAs (DEmRNAs) and lncRNAs (DElncRNAs).

first: Liying

last: Qin

first: Xiang

last: Gao

first: Xiuhai

last: Lu

first: Wencai

last: Liu

first: Han

last: Jiang

first: Jingyi

last: Tian

first: Jingjing

last: Zhang


title: Dormant tumors circumvent tumor-specific adaptive immunity by establishing a Treg-dominated niche via DKK3

city: Durham

first: Zachary

last: Hartman

state: NC

address: Research Dr MSRBI

department: Surgery

institute: Duke University

summary: Approximately 30% of breast cancer survivors deemed ‘free of disease’ will experience locoregional or metastatic recurrence even up to 30 years post initial diagnosis, yet how residual/dormant tumor cells escape immunity elicited by the primary tumor remains unclear. We demonstrate that intrinsically dormant tumor cells are indeed recognized and lysed by antigen-specific T cells in vitro and elicit robust immune responses in vivo. However, despite close proximity to CD8+ killer T cells, dormant tumor cells themselves support early accumulation of protective FoxP3+ T regulatory cells (Tregs), which can be targeted to reduce tumor burden. These intrinsically dormant tumor cells maintain a hybrid epithelial/mesenchymal or stem-like state, which is associated with immune dysfunction, and we find the tumor-derived stem/basal gene Dickkopf WNT Signaling Pathway Inhibitor 3 (DKK3) is critical for Treg inhibition of CD8+ T cells. We also demonstrate that DKK3 promotes immune-mediated progression of proliferative tumors and is significantly associated with poor survival and immune suppression in human breast cancers. Together, these findings reveal that latent tumors can use fundamental mechanisms of tolerance to alter the T cell microenvironment and subvert immune detection. Thus, targeting these pathways, such as DKK3, may help render dormant tumors susceptible to immunotherapies.

overall_design: D2A1 or D2.1 mouse mammary tumor cells were implated into the mammary fat pad of female Balb/c or SCID-beige mice (10^6/mouse). Tumors were collected upon reaching ~500 for bulk RNAseq. Cultured cells were also collected in triplicate for RNAseq.

first: Trotter

last: Timothy

first: Hartman

last: Zachary


title: DRG2 is required for cell surface localization of PD-L1 in melanoma cells and optimum response to anti-PD-1 immune checkpoint blockade [bulk RNA-seq]

summary: Tumors expressing high level of programmed cell death-1 (PD-1) ligand 1 (PD-L1) are more likely to respond to immune checkpoint blockers (ICBs) targeting PD-1 or PD-L1. However, more than half of tumor patients with high PD-L1 expression does not respond to ICBs and the underlying mechanisms are yet to be clarified. Here we show that depletion of developmentally regulated GTP-binding protein 2 (DRG2) inhibited recycling of endosomal PD-L1 and reduced surface PD-L1 level in melanoma cells. DRG2-depleted cells showed decreased binding with recombinant PD-1. Although DRG2-depleted cells expressed high levels of PD-L1, anti-PD-1 ICB did not activate T cells within DRG2-depleted tumors and failed to improve the survival of DRG2-depleted tumor-bearing mice. Cohort analysis of melanoma patients under anti-PD-1 treatment revealed that patients bearing tumors with high DRG2 protein level were more sensitive to PD-1 anti-PD-1 ICBs. These findings identify DRG2 as a regulator of recycling of endosomal PD-L1 and a key determinant for response to anti-PD-1 ICB and provide insights into how to increase the correlation between PD-L1 expression and response to ICB.

overall_design: We performed RNA-Seq on total RNA samples (RIN above 8.5) from B16F10-pLKO and B16F10-shDRG2 cells stimulated with 5 ng/ml recombinant mouse IFN-γ (PeproTech, 315-05) for 24 h.

first: Junil


title: The role of ALDH1B1 in human SW480 colon cancer cells

city: New Haven

first: Vasilis

last: Vasiliou

address: 60 College St

institute: Yale university

summary: To investigate the cooperative function ALDH1B1 in the regulation of colon cancer cells, we established SW480 cell lines in which ALDH1B1 has been knocked down by shRNA and the control SCR SW480 cells.

overall_design: We then performed gene expression profiling analysis using data obtained from RNA-seq of two cells groups.

first: Yewei

last: Wang


title: RNA-seq results in the kidney of female Tet1 knockout mice treated with or without ischemia reperfusion

city: WuHan

first: Yu

last: Fan

address: luojiashan

department: college of life science

institute: Wuhan university

summary: To understand key pathways and regulatory genes in  the kidney of female Tet1 knockout mice treated with ischemia reperfusion

overall_design: Female Tet1 knockout mice and their age-, sex- matched control mice were treated with unilateral renal ischemia/reperfusion (45 min / 3 days)  . For each experimental group, two biological replicates of RNA samples were sent for RNA-seq. Each sample consists of a pool of RNAs extracted from kidney of 3 different mice.

first: Ling

last: Zheng

first: Kun

last: Huang


title: Gene expression profile at single cell level of Innate Lymphoid Cells (ILCs), Natural Killer (NK) cells , and T cells from human tonsils

city: Huddinge

first: Christopher

middle: Thomas

last: Stamper

address: Alfred Nobels Allè 8

department: Medicine, Huddinge

country: Sweden

institute: Karolinska Institutet

summary: We used single cell RNA sequencing to investigate the diversity of ILCs, NKs, and T cells in the human tonsil

overall_design: Tonsillar ILCs, NK cells, and T cells were isolated by Fluorescence-activated cell sorting (FACS) and analyzed using scRNAseq.

middle: T

first: Jenny

last: Mjösberg


title: Oncometabolite succinylacetone activates HIF-1α signaling and promotes angiogenesis in GSTZ1-deficient HCC

city: chongqing

first: tang

last: ni

address: yixueyuan Road NO.1

institute: Chongqing Medical University

summary: We describe a unique regulation of the succinylacetone/PHD2/HIF-1α axis by a metabolic enzyme GSTZ1. The total RNA was isolated from HepG2 cells stably transfected with or without GSTZ1-KO and analyzed using RNA-seq. Our work demonstrates that GSTZ1-deficient promotes HCC angiogenesis through stabilizing HIF-1α due to succinylacetone accumulation. In addition, the combination of targeting HIF-1α and PD-L1 could limit tumorigenesis and progression for Gstz1-/-mice, thus providing a potential target for HCC therapy.

overall_design: To characterize GSTZ1-dependent global changes in the transcriptome under hypoxia, we conducted a genome-wide RNA sequencing (RNA-seq) analysis. First, we established GSTZ1 knockout (GSTZ1-KO) HepG2 cell lines using the CRISPR/Cas9 system. Total RNA was isolated from GSTZ1-KO and parental HepG2 cells (control) maintained under 1% O2 concentration for 12 h and subjected to sequencing.

first: HuaTing

last: Luo

first: Ni

last: Tang

first: Qiujie

first: Kai

first: Bo

last: Qin

first: Fan

last: Yang

first: Rui


title: Time series of motor neuronal translatomes over the course of neuroinflammation

city: Hamburg

first: Jan Broder

last: Engler

address: Falkenried 94

department: Institut für Neuroimmunologie und Multiple Sklerose

country: Germany

institute: Universitätsklinikum Hamburg-Eppendorf

summary: During the course of multiple sclerosis (MS), inflammatory insults drive neuro-axonal loss and disability progression. Here we leverage translating ribosome affinity purification (TRAP) to extract ribosome-bound mRNA from Chat-positive motor neurons of mice undergoing experimental autoimmune encephalomyelitis (EAE), the animal model of MS. This unique dataset allows to follow the temporal dynamics of neuronal responses to inflammation and enables the extractions of molecular targets for therapeutic intervention.

overall_design: mRNA of Chat+ spinal cord motor neurons derived from mice undergoing experimental autoimmune encephalomyelitis (EAE) and healthy control mice was enriched by translating ribosome affinity purification (TRAP) and sequenced by RNA-seq.

first: Iris

last: Winkler

first: Jan

middle: B

first: Manuel

last: Friese


title: CRISPR Phenotypic enrichment screen for epigenetic regulators of MEIS1 expression in acute myeloid leukemia (AML) [CRISPRScreens]

status: Public on Nov 10 2023

city: La Jolla

first: Karina

middle: Ofelia

last: Barbosa Guerra

address: 10901 North Torrey Pines Rd, Deshpande Lab RM6210B

department: Tumor Initiation & Maintenance Program

institute: Sanford Burnham Prebys Medical Discovery Institute

summary:
We perfomed CRISPR screens to investigate the upstream epigenetic regulators of MEIS1 expression, using a pooled CRISPR library targeting chromatin modifiers (~600 genes). We constructed the pooled library and used it to transduce U937 cells modified with a GFP knock-in in the MEIS1 locus. The cells were sampled at "Day 0" and kept in culture for 5 days, then sorted to harvest the top- and bottom- 20% fractions by GFP signal. Genomic DNA was then prepared from each fraction, as well as the Day 0 sample.
We then performed library preparation and next generation sequencing to compare the relative abundance of sgRNA sequences in the fractions of interest.

overall_design: Examination of sgRNA abundance in sorted fractions on the basis of GFP-MEIS1 expression in the U937 AML cell line.

last: Barbosa

first: Anagha

last: Deshpande

first: Ping

last: Xiang

first: Anna

last: Minkina

first: Fiorella

last: Schischlik

first: Adam

last: Brown

first: Neil

last: Robertson

first: John

last: Doench

first: Peter

last: Adams

first: R

last: Humphries

first: Eytan

last: Ruppin

first: Jay

last: Shendure

first: Prashant

last: Mali

first: Aniruddha

middle: J


title: DNA methylation and transcriptome dynamics of Flt3L induced ex vivo BMDC differentiation

city: Atlanta

first: Chris

last: Scharer

state: GA

address: 1510 Clifton Rd, Suite 3086A

department: Microbiology and Immunology

institute: Emory University

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

overall_design: Refer to individual Series


title: DNA methylation dynamics of Flt3L induced ex vivo BMDC differentiation

summary: To understand the DNA methylation dynamics during Flt3L induced Bone-marrow derived dendritic cell (BMDC) differentatiion we performed reduced representation bisulfite sequencing (RRBS).  Three distinct stages were chosen and included the initial bone marrow cells, the Common Myeloid Progenitor (CMP), and fully differentiated BMDC.

overall_design: RRBS on three distinct stages of ex vivo differentation of bone marrow derived dendritic cells.

first: C

first: P

last: Jain


title: RNA-seq of liver tissue extracted from mouse hepatectomy models (Control group vs. gelADSC group)

status: Public on Nov 05 2023

city: Suita

first: Hiroki

last: Imamura

state: Osaka

address: 2-2 Yamada-oka

country: Japan

institute: Osaka University

summary: To clarify the difference in the RNA expression between the control group (hepatectomized mice treated with fibrin gel only) and the gelADSC group (hepatectomized mice treated with ADSC embedded in fibrin gel), we extracted RNAs from the liver tissue.

overall_design: We investigated RNAs purified from the liver tissue of hepatectomy model mice. RNAs of the control group, which included hepatectomized mice treated with fibrin gel, were compared with those of the gelADSC group, which included hepatectomized mice treated with ADSC embedded in fibrin gel.


title: Gene expression dynamics of Flt3L induced ex vivo BMDC differentiation

summary: To understand the gene expression dynamics during Flt3L induced Bone-marrow derived dendritic cell (BMDC) differentatiion we performed RNA-seq.  Three distinct stages were chosen and included the initial bone marrow cells, the Common Myeloid Progenitor (CMP), and fully differentiated BMDC.

overall_design: RNA-seq on three distinct stages of ex vivo differentation of bone marrow derived dendritic cells.


title: Gene expression of PCSM-LPD

city: Kiel

first: Julia

last: Richter

address: Arnold-Heller Straße 3, Haus U33

department: Department of Pathology, Hematopathology Section

institute: University Hospital Schleswig-Holstein

summary: Primary cutaneous CD4+ small or medium T-cell lymphoproliferative disorder (PCSM-LPD) is a clonal T-cell proliferation disease confined to the skin. PCSM-LPD shares expression of T-follicular T helper cell (Tfh) markers with various mature T-cell lymphomas. However, the benign presentation of PCSM-LPD contrasts the clinical behavior of other Tfh-lymphomas. The aim of our study was to delineate the molecular similarities and differences between PCSM-LPD and other Tfh-derived lymphomas, to explain the clinical behavior and to unravel possible pathological mechanisms. We performed targeted next generation sequencing of 19 genes recurrently mutated in T-cell neoplasms in n=17 PCSM-LPD with high and in n=21 PCSM-LPD with low tumor cell content. Furthermore, gene expression profiling was used to identify genes potentially expressed in the PD1+ neoplastic cells. Expression of some of these genes was confirmed in situ using multi-stain immunofluorescence. We found that PCSM-LPD rarely harbors mutations recurrently detected in other T-cell neoplasms. PCSM-LPD is characterized by the invariable expression of the T-cell receptor associated LCK protein. CD70 and its ligand CD27 are co-expressed on PD1+ PCSM-LPD cells, suggestive of auto-activation of the CD70 pathway. In conclusion, PCSM-LPD differs from disseminated lymphomas of Tfh origin by their mutation profile. Activation of CD70 signaling also found in cutaneous T-cell lymphoma represents a potential driver of neoplastic proliferation of this benign neoplasia of Tfh.

overall_design: 10 PCSM-LPD primary samples and 5 patients diagnosed with pseudolymphomas were analyzed using NanoSting PanCancer Immune Profiling Panel (Human) covering 770 genes


title: DRG2 is required for cell surface localization of PD-L1 in melanoma cells and optimum response to anti-PD-1 immune checkpoint blockade [scRNA-seq]

first: Jeonghwan

last: Kim

overall_design: B16F10 tumors were collected at 3 days after 2nd administration of anti-PD-1 antibody. The tumor samples were first mechanically dissociated using a scalpel, then enzymatically dissociated in digestion medium (2mg/ml Collagenase P (Sigma Aldrich) and 0.2mg/ml DNAse I (Roche) in DMEM (Thermo Fisher Scientific)). Red blood cells were removed from the cell suspension using red blood cell lysis buffer (Roche), and the cells were filtered using a 40-µm Flowmi tip strainer (VWR). The number of living cells was determined using a LUNA automated cell counter (Logos Biosystems). We performed single-cell RNA-seq on the single-cell suspension using the Chromium Single Cell 3’ Solution from 10x Genomics according to the manufacturer’s instructions. Up to 5,000 cells were loaded onto a 10x Genomics cartridge for each sample. Cell-barcoded 5′ gene expression libraries were sequenced on an Illumina NextSeq and/or NovaSeq6000 system.

first: Junil


title: Vitamin D Receptor binding and chromatin accessibility in mouse duodenal crypt and villus and colon epithelium

city: Piscataway

first: Michael

middle: P

last: Verzi

state: NJ

address: 145 Bevier Road

department: Genetics

institute: Rutgers, the State University of New Jersey

summary: VDR is critical for Ca++ homeostasis. While its role in this process within the intestine has primarily been focused to the duodenal epithelium, we have uncovered a compensatory role for VDR in the colon in mediating Ca++ uptake. Here, we compare VDR genomic binding properties in the colon and duodenum using ChIP-seq. We also examine the chromatin accessibility between these intestinal regions.

overall_design: The C57BL/6J mice were fed with regular chow diet and exposed to a 12h-light, 12h-dark cycle. Food and water were given ad libitum. When the mice were between the age of 4-6 weeks old, 100uL of 2.5 ng/uLof 1,25(OH)2D3 (Caymon Chemical Company, Ann Arbor, MI) (IP 10ng/g body weight) were administered intraperitoneally for 1 hour and harvested. Some of the C57BL/6J originated from the Jackson Laboratory while others were generated and maintained at the animal care facility at Rutgers University according to the guidelines of the Animal Care and Use Committee at Rutgers University

first: Dennis

last: Aldea

first: Joseph

last: Hur

first: Oscar

last: Pellon-Cardenas

first: Lei

last: Chen


title: Endogenized hypervirulence-associated DNA virus BdDV-1 is primarily restricted to enzootic lineages of the amphibian pathogen Batrachochytrium dendrobatidis

city: Riverside

first: Jason

last: Stajich

state: California

address: 900 Univeristy Avenue

department: Microbiology and Plant Pathology

institute: University of California, Riverside

summary: The Global Panzootic Lineage (GPL) of the pathogenic fungus Batrachochytrium dendrobatidis (Bd) has caused severe amphibian population declines, yet the drivers underlying the high frequency of GPL in regions of amphibian decline are unclear. Using publicly available Bd genome sequences, we identified multiple non-GPL Bd isolates that contain a circular Rep-encoding single stranded DNA-like virus which we named BdDV-1. We further sequenced and constructed genome assemblies with long read sequences to find that the virus is integrated into the nuclear genome in some strains. Attempts to cure virus positive isolates were unsuccessful, however, phenotypic differences between naturally virus positive and virus negative Bd isolates suggested that BdDV-1 decreases the growth of its host in vitro but increases the virulence of its host in vivo. BdDV-1 is the first described CRESS DNA mycovirus associated with hypervirulence with a distribution inversely associated with the emergence of the panzootic lineage.

overall_design: To identify differentially expressed genes (DEGs) between v+ and v- Bd-BRAZIL we extracted and sequenced mRNA from three samples of the virus positive strain, CLFT067 and the virus negative strain, CLFT071. These strains were selected due to their close phylogenetic relationship and their shared geographic location

first: Mark

last: Yacoub

first: Rebecca

last: Clemons

first: Timoth

middle: Y

last: James

middle: E


title: Effects of different reoxygenation methods after acute hypoxic stress on the metabolic remodeling of golden pompano (Trachinotus blochii)

city: Haikou

first: tian

last: jiang

address: 58 of Ren min Road

institute: Hainan University

summary: Numerous studies have demonstrated that golden pompano (Trachinotus blochii) is sensititive to hypoxia, which causes a devastating blow to the golden pompano industry. And different methods of reoxygenation after hypoxia could bring differnt effects on metabolism for golden pompano.

overall_design:
Examination a total of 24 RNA-seq of liver in different duration of hypoxia and different methods of reoxygenation.
Please note that the identifiers in processed data (e.g. Trachinotus_GLEAN_10000042, etc.) are self-numbered in the transcriptome sequencing, and then these identifiers were annotated based on the genome as included in the Tra_gene.nr.xls annotation file.

first: Tian

last: Jiang

first: Jun

middle: L

last: Sun


title: Disease-specific transcriptomic changes induced by NFκB inhibition in idiopathic pulmonary fibrosis

city: Clayton

first: Nicholas

middle: C

last: Wong

state: Victoria

address: Wellington Road

department: Monash Bioinformatics Platform, Central Clinical School

institute: Monash University

summary: Idiopathic pulmonary fibrosis (IPF) is a progressive fibrotic lung disease of unknown etiology and poor prognosis. In IPF, aberrant extracellular matrix production by activated, hyperproliferative fibroblasts drives disease progression but the exact mechanisms by which this occurs remains undefined. We previously showed that inhibition of transcription factor nuclear factor kappa-B (NFκB) with ACT001 reduced pro-fibrotic fibroblast behaviors such as fibroblast-to-myofibroblast transition. The aim of this follow-up study was to characterize the transcriptional profile of primary fibroblasts treated with ACT001 and identify differences between cells from patients with IPF and non-diseased control donors. Primary lung fibroblasts derived from eight patients with IPF and eight age-matched non-diseased controls (NDC) were treated with 0, 3 or 10 µM ACT001 in the presence/absence of pro-fibrotic cytokine transforming growth factor (TGF)-β1 for 24 hours after which whole genome RNA sequencing was performed. Gene set enrichment analysis (GSEA) of the Hallmark gene sets was also completed. Although the majority of genes differentially expressed after ACT001 treatment were the same in NDC and IPF cells, several genes were uniquely changed in IPF. GSEA indicated that ACT001 treatment most upregulates the reactive oxygen species pathway.  ACT001 is a water-soluble compound with a stable half-life in plasma, thus making it an attractive candidate for further investigation as a therapeutic in IPF. This study adds to the growing body of literature that demonstrates anti-fibrotic activity of NF-ĸB inhibition in the context of IPF.

overall_design: Primary cultered human fibroblasts from 16 individuals, 8 with idiopathic pulmonary fibrosis (IPF) and 8 age-matched controls

first: Jade

last: Jaffar

first: Ian

last: Glaspole

first: Karen

last: Symons

first: Glen

last: Westall

first: Nick


title: RNA-sequencing assay to Identify changed gene expression in stomach adenocarcinoma  (STAD) cells with ACAT2 knockdown

status: Public on Nov 04 2023

city: Tianjin

first: Mengmeng

last: Zhang

address: Huanhuxi Street

department: Department of Gastric Surgery, Tianjin Medical University Cancer Institute & Hospital

summary: Purpose: To elucidate how ACAT2 influenced the expression profile in STAD cells, ACAT2 expression was reduced using one shRNA in STAD HGC-27 cells. Then, transcriptional profiling of cells with ACAT2 knockdown (shACAT2) and the control cells (shNC) was performed to characterize differentially expressed genes (DEG).Methods: When shNC and shACAT2 cells grew to approximately 80% confluence, these cells were washed twice using pre-chilled PBS. Total RNAs were harvested using Trizol reagents (Thermo Fisher Scientific).  Poly(A) mRNA was isolated and enriched using NEBNext Poly(A) mRNA Magnetic Isolation Module. The mRNA was then recovered for library generation with NEBNext ® Ultra™ Directional RNA Library Prep Kit for Illumina (NEB, E7420S) following the manufacturer’s instructions. The cDNA libraries were sequenced at WuXiNextCODE (China). FastQC was used to examine the quality of raw reads. Read alignment was conducted using STAR (v2.5.1b),  and R package edgeR (v3.8.5) was used to determine relative transcript abundances and differentially expressed genes (DEGs) between sample pairs. Results: Generally, 241 genes  were increased and 117 genes decreased in HGC-27 shACAT2 cells, compared with shNC cells (FDR<0.05, Fold Change>2). GO analysis showed that these changed genes were significantly enriched in signal pathways related to cancer. Conclusions: Reduced ACAT2 expression alters multiple cancer-related signal pathways.

overall_design: Three biological replicates for each group shNC and shACAT2 - were subjected to RNA-sequencing analysis respectively, among which shNC acted as the reference sample for shACAT2.

first: Jingyu

last: Deng


title: Pan-cancer single-nucleus total RNA sequencing using HH-seq

city: Hangzhou

first: Haide

address: 866 Yuhangtang road

institute: Zhejiang University

summary: Tumor heterogeneity and its drivers impair tumor progression and cancer therapy. Single-cell RNA sequencing has been used to investigate the heterogeneity of tumor ecosystems. However, most methods of scRNA-seq amplify the termini of polyadenylated transcripts, making it challenging to perform total RNA analysis and somatic mutation analysis during tumor processing. Additionally, frozen tumor samples constitute a vast and valuable material bank for cancer research. Therefore, we developed a high-throughput and high-sensitivity method called HH-seq, which combines random primers and a pre-index strategy in the droplet microfluidic platform. This innovative method allows for the detection of total RNA in single nuclei from clinically frozen samples. We also established a robust pipeline to facilitate the analysis of full-length RNA-seq data. We applied HH-seq to more than 730,000 single nuclei from 32 patients with various tumor types. The pan-cancer study enables us to comprehensively profile data on the tumor transcriptome, including expression levels, mutations, splicing patterns and clone dynamic, etc. We identified new malignant cell subclusters and explored their specific function across cancers. Furthermore, we investigated the malignant status of epithelial cells among different cancer types with respect to mutation and splicing patterns. The ability to detect full-length RNA at the single-nucleus level provides a powerful tool for studying complex biological systems and has broad implications for understanding tumor pathology.

overall_design:
Single nuclei RNA-sequencing of cell lines and mouse tissues using HH-seq, benchmark analysis was performed.
7 mouse brain samples were analyzed in four conditions of snHH-seq: different reverse transcription primer (No_index, RT primer without cell barcode 2; Index, RT primer with cell barcode 2), different sample storage (frozen, nuclei isolated from frozen sample; fresh, nuclei isolated from fresh sample).
The MIX samples (human-mouse mix experiment) show the effect of cell concentration on contamination rate.

first: Guoji

last: Guo

first: Xiunan

last: Fang

first: Jikai

last: Shao


title: Expression data from monocyte-derived macrophages co-cultured in the presence of multiple myeloma (MM) cell line and/or MM-bone marrow stromal cell (BMSC)

city: Pamplona

first: Aintzane

last: Zabaleta

state: Navarra

address: Avda. Pío XII 55

country: Spain

institute: FOUNDATION FOR APPLIED MEDICAL RESEARCH

summary:
Bone marrow (BM) microenvironment of multiple myeloma (MM) is populated by tumor associated monocytes and macrophages (TAMs), where they protect clonal plasma cells (PC). It is well known that MM cells can modulate the transcriptome of macrophages, thus, we sought to investigate if a key component of the tumor microenvironment such as MM-BMSCs have the potential to modulate the transcriptome of macrophages as MM cells do.
We used microarrays to analyze the potential of MM-BMSC to polarize the transcriptome of TAMs in the presence and absence of MM cells

overall_design: MACS sorted CD14+ monocytes from healthy donors were cultured in 12 well plates in the presence of 10 ng/ml of M-CSF for 7 days to differentiate into macrophages (MDM). On the seventh day cells were harvested and co-cultured in the presence of MM cells, MM-BMSC or both for 24 hrs. Afterwards macrophages were FACSsorted and stored in RLT Plus buffer till RNA extraction.

first: Bruno

last: Paiva


title: Chromatin binding of RUNX1 in T-cell acute lymphoblastic leukemia (T-ALL) cells with depletion cBAF complex [ChIP-seq]

city: Kyoto

first: Kosuke

last: Yusa

state: Kyoto

address: 53 Shogoin Kawaharacho, Sakyo-ku

department: Institue for Life and Medical Sciences

institute: Kyoto University

summary: Using ChIP-seq of RUNX1 in the cells with cBAF-targeting gRNA, we identified RUNX1 binding chromain loci which is regulated by cBAF complex

overall_design: Cas9-expressing cells were lentivirally transduced with gRNA, selected with puromycin for 4 days, and then harvested on day 7 after gRNA transduction.

first: Kazunari

last: Aoki

first: Yotaro

last: Ochi

first: Yusuke

last: Tarumoto


title: Deletion of NuRD component Mta2 in nephron progenitor cells causes developmentally programmed FSGS [2month.ChIPseq]

city: St. Louis

first: Jeannine

middle: M

last: Basta

state: MO

department: Internal Medicine/Nephrology

institute: Washington University in St. Louis

summary: Low nephron endowment at birth is a risk factor for chronic kidney disease. The prevalence of this condition is increasing due to higher survival rates of preterm infants and children with multi- organ birth defect syndromes that affect the kidney and urinary tract. We created a mouse model of congenital low nephron number due to deletion of Mta2 in nephron progenitor cells. Mta2 is a core component of the Nucleosome Remodeling and Deacetylase (NuRD) chromatin remodeling complex. These mice developed albuminuria at 4 weeks of age followed by focal segmental glomerulosclerosis (FSGS) at 8 weeks, with progressive kidney injury and fibrosis. Our studies reveal that altered mitochondrial metabolism in the post-natal period leads to accumulation of neutral lipids in glomeruli at 4 weeks of age followed by reduced mitochondrial oxygen consumption. We found that NuRD cooperated with Zbtb7a/7b to regulate a large number of metabolic genes required for fatty acid oxidation and oxidative phosphorylation. Analysis of human kidney tissue also supported a role for reduced mitochondrial lipid metabolism and ZBTB7A/7B in FSGS and CKD. We propose that an inability to meet the physiological and metabolic demands of post-natal somatic growth of the kidney promotes the transition to CKD in the setting of glomerular hypertrophy due to low nephron endowment.

overall_design: Binding of Mta2, Mbd3, and Zbtb7a to chromatin was measured by comparing specific chromatin binding to Input chromatin in 2 month old mouse kidney cortex.

first: Michelle

last: Brennan

first: Michael

last: Rauchman


title: Deletion of NuRD component Mta2 in nephron progenitor cells causes developmentally programmed FSGS


title: Implications of Prenatal Exposure to Hyperandrogen for Hippocampal Neurodevelopment and Autism-like Behavior in Offspring

city: Shijiazhuang

first: Dan

last: Qiao

state: Hbbei Province

address: 361 Zhongshan East Road, Chang 'an District, Shijiazhuang City, Hebei Province

institute: Hebei Medical University

summary: Autism spectrum disorder (ASD) is a highly heterogeneous neurodevelopmental disorder that significantly jeopardizes the physical and mental well-being of children. Autism spectrum disorder results from a combination of environmental and genetic factors. Hyperandrogenic exposure during pregnancy increases their risk of developing autism. Nevertheless, the prenatal exposure to androgens affects offspring neurodevelopment and the underlying mechanisms have not been fully elucidated. In the present study, administration of excessive dihydrotestosterone (DHT) to pregnant mice was found to impair neuronal development and dendritic spine formation in offspring, inducing autism-like behaviors. Furthermore, through mRNA transcriptome sequencing technology, the key molecule Nr4a2 was identified during this process of change. Overexpression of Nr4a2 and treatment with amodiaquine (AQ) significantly improved the abnormal phenotypes in offspring caused by prenatal exposure to androgens. Overall, Nr4a2 emerges as a crucial molecule involved in the impairment of offspring neurodevelopment due to prenatal androgen exposure, which provides a new perspective for the in-depth study of the influencing factors and underlying mechanisms.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for offspring of DHT and C57 mice.

first: Chenyu

last: Mu

first: Huan

last: Chen

first: Di

last: Wen

first: Zhao

last: Wang

first: Chang

first: Yingjing

last: Lu

first: Yingling

last: Ge

first: Xudi

last: Jiang

first: Dexian

last: Li

first: Zixuan

first: Huixian

last: Cui

first: Sha


title: Characterization of cBAF complex in T-cell acute lymphoblastic leukemia (T-ALL)

first: Kosuke

last: Yusa


title: Chromatin accessibility of T-cell acute lymphoblastic leukemia (T-ALL) cells with depletion or inhbiiton of cBAF complex [ATAC-seq]

summary: Using ATAC-seq of the cells with cBAF-targeting gRNA or inhibitor, we identified chromatin loci whose accessibility was regulated by cBAF complex

overall_design: For gene depletion, Cas9-expressing cells were lentivirally transduced with gRNA and selected with puromycin for 4 days. Cells were harvested on day 10 after gRNA transduction. For inhibitor treatment, cells were treated with 1 uM BRM014 for 6 hours or 8 days.

first: Kazunari

last: Aoki

first: Yotaro

first: Yusuke


title: Multi-omics datasets for finer-scale dissection of chromatin architecture alterations in radiation induced malignant transformation of bronchial epithelial cells [RNA-seq]

city: Beijing

first: Yi

last: Hu

address: No. 28 Fuxing Rd， Haidian district，Beijing

institute: Chinese PLA General Hospital

summary: we generated ATAC-seq, RNA-seq and high-resolution Hi-C datasets for finer-scale dissection of chromatin architecture alterations in radiation induced malignant transformation of bronchial epithelial cells.

overall_design: The mRNA profiles of normal human bronchial epithelial cells and radiation induced lung cancer cells

first: Zhou

last: pingkun

first: Liu

last: qiaowei


title: Multi-omics datasets for finer-scale dissection of chromatin architecture alterations in radiation induced malignant transformation of bronchial epithelial cells [Hi-C]

overall_design: The Hi-C profiles of normal human bronchial epithelial cells and radiation induced lung cancer cells


title: Deletion of NuRD component Mta2 in nephron progenitor cells causes developmentally programmed FSGS [2month.RNAseq]

first: Jeannine

last: Basta

overall_design: We measured RNA expression by RNA-seq in control Mta2flx/flx and mutant Six2Cre Mta2flx/flx 2 month old kidneys.

first: Michelle

last: Brennan

first: Michael


title: Gene Expression of A375 Cells After Fluvastatin Treatment

city: Cleveland

first: Ernest

middle: R

last: Chan

state: OH

address: 10900 Euclid Ave, Robbins E211

department: PQHS

institute: CWRU

summary: Despite advances in melanoma treatment, more than 70% of patients with distant metastasis die within 5 years. Proactive treatment of early melanoma to prevent metastasis could save lives and reduce overall healthcare costs. Currently, there are no treatments specifically designed to prevent early melanoma from progressing to metastasis.  We used the Connectivity Map (cMap) to conduct an in silico drug screen and identified HMGCR inhibitors (statins) as a drug class that might prevent melanoma metastasis. To confirm the in vitro effect of statins, RNA-sequencing was completed on A375 cells after treatment with fluvastatin to describe changes in the melanoma transcriptome. Statins induced differential expression in genes associated with metastasis and used in commercially available prognostic tests for melanoma metastasis.

overall_design: We used RNA-sequencing to measure gene expression of A375 melanoma cells before and after treatment with fluvastatin. The A375 cell line was specifically chosen since it has moderate metastatic potential and has been used in previous mechanistic studies of statins. Fluvastatin was chosen because of its lipophilicity (allowing extrahepatic distribution), benign side effect profile, and excellent bioavailability. We found 2615 differentially expressed genes

first: Wesley

last: Yu

first: John

last: Pink

first: Alexander

last: Miron

first: E


title: Identification of a miRNA multi-targeting therapeutic strategy in glioblastoma

status: Public on Nov 03 2023

city: Lyon

first: Erika

last: COSSET

state: Rhône

address: 28 Rue Laennec

country: France

institute: Centre de Recherche en Cancérologie de Lyon

summary: Glioblastoma (GBM) is a deadly and the most common primary brain tumor in adults. Due to their regulation of a high number ofmRNA transcripts, microRNAs (miRNAs) are key molecules in the control of biological processes and are thereby promisingtherapeutic targets for GBM patients. In this regard, we recently reported miRNAs as strong modulators of GBM aggressiveness.Here, using an integrative and comprehensive analysis of the TCGA database and the transcriptome of GBM biopsies, we identifiedthree critical and clinically relevant miRNAs for GBM, miR-17-3p, miR-222, and miR-340. In addition, we showed that thecombinatorial modulation of three of these miRNAs efficiently inhibited several biological processes in patient-derived GBM cells ofall these three GBM subtypes (Mesenchymal, Proneural, Classical), induced cell death, and delayed tumor growth in a mouse tumormodel. Finally, in a doxycycline-inducible model, we observed a significant inhibition of GBM stem cell viability and a significantdelay of orthotopic tumor growth. Collectively, our results reveal, for the first time, the potential of miR-17-3p, miR-222 and miR-340multi-targeting as a promising therapeutic strategy for GBM patients.Cell Death and Disease (2023) 14:630 ; https://doi.org/10.1038/s41419-023-06117-z

overall_design: In this study, by harnessing the wealth of public data stored in the Cancer Genome Atlas Research Network (TCGA) database, we identified miR-17-3p, miR-222, and miR-340 as three clinically relevant miRNAs that can be used for a GBM multitargeting therapeutic strategy. Thus, to get some insight into the potential regulated genes, we performed RNASeq on GBM cell line transfected with the identified miRNAs.

first: Arthur

last: Bassot

first: Helena

last: Dragic

first: Sarah

last: Al Haddad

first: Laurine

last: Moindrot

first: Soline

last: Odouard

first: Francesca

last: Corlazzoli

first: Eliana

last: Marinari

first: Alexandra

last: Bomane

first: Augustin

last: Brassens

first: Antoine

last: Marteyn

first: Youssef

last: Hibaoui

first: Tom

last: Petty

first: Mounira

last: Chalabi-Dchar

first: Louis

last: Larrouquere

first: Evgeny

last: Zdobnov

first: Noémie

last: Legrand

first: Jérôme

last: Tamburini

first: Hubert

last: Lincet

first: Marie

last: Castets

first: Mayra

last: Yebra

first: Denis

last: Migliorini

first: Valérie

last: Dutoit

first: Paul

last: Walker

first: Olivier

last: Preynat-Seauve

first: Pierre-Yves

last: Dietrich

first: Érika

last: Cosset


title: Differential gene expression in CHO cells cultivated in shake flasks and bioreactors

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

institute: Wageningen University

summary: Chinese Hamster Ovary cell lines are currently the primary host for production of therapeutic glycoproteins. Fast process development resulting in robust and scalable processes is a critical success factor in the highly competitive market for biosimilars. In process development screening of hundreds of clones and selection of process conditions are routinely performed in uncontrolled cultivation systems like shake flasks. A handful of potential candidate clones is nominated to be evaluated more intensively in well controlled small-scale bioreactors. Cell performance in the uncontrolled systems and to a lesser extent in the small-scale bioreactors may, however, be different from that in the final production reactor, which may result in failures during scale-up and thus extra development time. In this work, the focus is on better understanding the differences in cell performance between controlled and uncontrolled systems, which can be used to make process development faster and more robust in terms of scale-up. For this, we evaluated differences in gene expression profiles between shake flask and bioreactor cultures at three different time points during the exponential and stationary phase of a batch culture using commercially available Affymetrix GeneChip CHO Gene 2.1 ST arrays and multivariate data analysis on the outcomes. The outcomes were correlated with differences in glycosylation patterns and other culture parameters. Results showed large differences in gene expression over time and much smaller differences between the two cultivation systems. Furthermore, our study identified differentially expressed genes and corresponding metabolic and mechanistic pathways between the two systems, which were directly related to the degree of the control of the systems.

overall_design: CHO cells, that originated from the same pre-culture, were grown in uncontrolled shake flasks and controlled bioreactors, and gene expression profiles were determined after 2, 4 and 5 days of culture.

first: Abdulaziz

middle: A

last: Alsayyari

first: Jos

last: Hageman

first: Rene

middle: H

last: Wijffels

first: Dirk

last: Martens


title: Multi-omics datasets for finer-scale dissection of chromatin architecture alterations in radiation induced malignant transformation of bronchial epithelial cells [ATAC-seq]

first: Yi

last: Hu

overall_design: ATAC-Seq profiles of normal human bronchial epithelial cells and radiation induced lung cancer cells

first: Zhou

last: pingkun

first: Liu

last: qiaowei


title: Expression profiles of T-cell acute lymphoblastic leukemia (T-ALL) cell lines with depletion or inhbiiton of cBAF complex [RNA-seq]

first: Kosuke

last: Yusa

summary: From comparative transcriptome analysis using RNA-seq of the cells with cBAF-targeting gRNA or inhibitor, we identified the genes regulated by  cBAF complex in T-ALL cell lines

overall_design: For gene depletion, Cas9-expressing cells were lentivirally transduced with gRNA and selected with puromycin for 4 days. RNA was isolated from the knockout cells on day 10 after gRNA transduction. For inhibitor treatment, RNA was isolated from the cells treated with 1 uM BRM014 for 6 hours.

first: Kazunari

last: Aoki

first: Yotaro

last: Ochi

first: Yusuke

last: Tarumoto


title: tRF & tiRNA sequencing of intestinal biopsies from IBS-D patients

city: Zhengzhou

first: Yuna

last: Chai

state: Henan

address: Jianshe East Road

department: Department of Pharmacy

institute: The First Affiliated Hospital of Zhengzhou University

summary:
Purposes: To investigate the epigenetic mechanism of IBS-D(Irritable Bowel Syndrome with Diarrhea) by tRF & tiRNA sequencing in intestinal biopsies of IBS-D patients and healthy volunteers
Methods: Five IBS-D and five healthy volunteers were screened, and biopsies were taken under colonoscopy. Small RNA sequencing was performed on Illumina NexSeq instrument
Results:If P < 0.05, fold change > 1.5 as the cutoff, there were 14 up-regulated tRFs & tiRNAs and 14 down-regulated tRFs & tiRNAs.
Conclusions:There were 14 up-regulated tRFs & tiRNAs and 14 down-regulated tRFs & tiRNAs in intestinal tissues of IBS-D .

overall_design: tRF & tiRNA sequencing of intestinal biopsies from five IBS-D patients and five healthy volunteers

first: Yaoyao

last: Lu


title: Multipotent Progenitors Instruct Ontogeny of the Superior Colliculus – MADM-CloneSeq E12

city: Klosterneuburg

first: Florian

last: Pauler

address: Am Campus 1

country: Austria

institute: Institute of Science and Technology Austria (IST Austria)

summary: The superior colliculus (SC) in the mammalian midbrain is essential for multisensory integration and is composed of a rich diversity of excitatory and inhibitory neurons and glia. The developmental principles directing the generation of SC cell-type diversity are however not understood. Here we pursued systematic cell lineage tracing in silico and in vivo, preserving full spatial information, using genetic MADM (Mosaic Analysis with Double Markers)-based clonal analysis with single-cell sequencing (MADM-CloneSeq). The analysis of clonally-related cell lineages revealed that radial glial progenitor (RGP) cells in SC are exceptionally multipotent. Individual resident RGPs have the capacity to produce all excitatory and inhibitory SC neuron types, even at the stage of terminal division. While individual clonal units show no pre-defined cellular composition, the establishment of appropriate relative proportions of distinct neuronal types occurs in a PTEN-dependent manner. Collectively, our findings provide an inaugural framework at single RGP/cell level of the mammalian SC ontogeny

overall_design: scRNA-Seq of neurons originating from a single radial glia progenitor (clone) labeled at E12 using MADM and Fz10-CreER or Sox2-CreER

first: Giselle

last: Cheung

first: Simon

last: Hippenmeyer


title: Multipotent Progenitors Instruct Ontogeny of the Superior Colliculus – Pten

overall_design: scRNA-Seq of Pten KO or control cells from the dorsal midbrain


title: Multipotent Progenitors Instruct Ontogeny of the Superior Colliculus - pooled embryonic Fzd10 lineage

overall_design: scRNA-Seq of cell from the Fzd10 lineage


title: Transcriptomic analysis of SLAN+ vs SLAN- non-classical monocytes

first: Aintzane

last: Zabaleta

summary:
Monocytes can be classified into four different subsets: classic, intermediate and SLAN- and SLAN+ non-classical monocytes. SLAN+ monocytes are significantly reduced in MM patients compare to the frequency found in healthy adults (HA). Thus, we sought to analyze the transcriptome of SLAN- and SLAN+ cells to unveil the differences among both populations
We used microarrays to detail the global programme of gene expression underlying the differences among both cell subsets
We developed a 12-color combination of mAbs to evaluate the contribution of each individual marker for identifying and subsetting BM TAMs in patients with MM (N = 5): CD36-UV405, CD16-UV525, CD163-BV450, CD45-BV525, CD206-BV610, CD86-BV660, CD62L-BV763, cyCD68-FITC, SLAN-PE, HLADR-PerCPCy5.5, CD300e-APC, CD14-APCH7. This panel allowed us to identify the aforementioned four monocyte subsets. Intermediate and non-classical monocytes from healthy donors were FACS sorted based on the 7-color mAb combination - HLADR-PacB, CD45-OC515, CD36-FITC, SLAN-PE, CD16-PECy7, CD300e-APC, CD14APCH7 -

overall_design: Gene expression data from CD36+ SLAN- monocytes, CD36- SLAN- monocytes, and CD36- SLAN+ monocytes.

first: Bruno

last: Paiva


title: Multipotent Progenitors Instruct Ontogeny of the Superior Colliculus

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

overall_design: Refer to individual Series


title: A recombination-resistant genome for live attenuated and stable PEDV vaccines by engineering the transcriptional regulatory sequences

city: Shenzhen

first: SHAO

middle: MIN

last: YANG

state: guangdong

address: Nanshan

department: Shenzhen Nanshan People's Hospital and the 6th Affiliated Hospital of Shenzhen University Health Science Center

institute: Department of Pain Medicine and Shenzhen Municipal Key Laboratory for Pain Medicine

summary: Porcine epidemic diarrhea virus (PEDV) is a deadly coronavirus for neonatal piglets and no effective vaccines are available. Transcriptional regulatory sequences (TRSs) are critical in regulating coronavirus discontinuous transcription. Also, TRSs contribute to a high recombination rate of coronaviruses, leading to difficulty in developing safe live vaccines. We hypothesize that recoding the TRS core sequences (TRS-CS) of PEDV can make the recombination impossible between the engineered vaccine virus and field strains or wildtype viruses. We used an infectious clone-derived reporter PEDV, dORF3-EGFP, as the backbone to generate a remodeled TRS (RMT) mutant that carries the recoded leader and body TRS-CSs. The RMT and dORF3-EGFP showed comparable replication efficiency in Vero cells. However, the incompatibility between the rewired and wildtype TRS-CSs led to few EGFP in RMT-infected cells. Furthermore, RMT and dORF3-EGFP had a similar attenuated phenotype, replication efficiency, and protective immunogenicity in neonatal pigs. RNA sequencing analysis indicated that EGFP transcription directed by the heterogenous TRS-CSs was significantly reduced to an extremely low level. Meanwhile, recombinant viruses were not detected in Vero cells and in pigs that were co-infected with RMT and a PEDV S-INDEL strain, Iowa106. In vitro and in vivo passaging of the RMT did not result in reversion mutations in the rewired TRS-CSs, introduced gaps, and disrupted wildtype TRSs. In summary, the RMT mutant was resistant to recombination and genetically stable and can be further optimized (e.g., deletion of the EGFP) to serve as a platform to develop safe PEDV live attenuated vaccines.

overall_design: The PEDV mutants were generated from the five-plasmid (A, B, C, D, and E) reverse genetic system for a highly virulent PEDV strain PC22A (28). For dORF3-EGFP, the accessory gene, ORF3, was removed and a reporter gene, EGFP, was inserted into plasmid E under the control of the original TRS for ORF3, designated as pE-dORF3. As for the RMT mutant, the wildtype leader TRS-CS (5’-CUAAAC-3’) in plasmid A was recoded (5’-GUGAAC-3’) using a NEB Q5® Site-Directed Mutagenesis Kit (NEB, Ipswich, MA). And the pE-dORF3 were modified accordingly to rewire all body TRS-CS to 5’-GUGAAC-3’, except the one for EGFP gene. Then the sequencing confirmed plasmids were digested with restriction enzymes for purification of DNA fragments that were assembled to form the full-length cDNA of PEDV mutants by in vitro ligation with T4 ligase (NEB, Ipswich, MA) at 4 °C overnight in an equal molar ratio. After purification with chloroform extraction, the cDNA was used as templates for in vitro transcription to generate the PEDV mutant mRNA genome using a mMessage mMachine T7 transcription kit (Ambion, Austin, CA). Meanwhile, a polyadenylated PEDV N gene transcript was generated to be co-electroporated with the mRNA genome into Vero cells at 450 V and 50 μF using a Gene Pulser II electroporator (Bio-Rad, Hercules, CA). At 6 hours post electroporation, cells were washed with PBS and cultured in the maintenance medium in the presence of 5 μg/mL of trypsin. Cells were monitored for CPE daily and harvested when CPE was observed for about 50% cells. This rescued virus was designed as P0 virus. The P0 virus was subjected to plaque purification and one clone was propagated. After the whole genome was confirmed by sanger sequencing, it was designated as the P1 stock of the mutants.

first: Qiuhong

last: Wang


title: Whole genome expression microarray of cancer stem cells isolated from HT29 and HCT116 Human colorectal cancer cell lines

city: Delhi

first: Janvie

last: Manhas

state: Delhi

address: Ansari Nagar

department: Biochemistry

country: India

institute: All India Institute of Medical Sciences

summary:
In the present study we have isolated and characterized cancer stem cells and non-cancer stem cells (bulk tumor cells) from high grade human colorectal cancer cell line HCT116 and low grade human colorectal cancer cell line HT29.
For this study, cancer stem cells and non-cancer stem cells (bulk tumor cells) were isolated from HCT116 and HT29 human colorectal cancer cell line. For isolating cancer stem cells by FACS, CD44 and CD166 tagged with V450 and PE respectively were used. CD44+CD166+ was the cancer stem cell population and CD44-CD166- was designated as the non-cancer stem cell (bulk tumor cells) population for this study. RNA was isolated from the isolated cell populations and microarray was done to study the whole genome transcriptomic changes.

overall_design: 4 x 44k Whole genome expression arrays of FACS sorted HT29 cancer stem cells (n=2) and HT29 non-cancer stem cells (n=2) using total RNA obtained from FACS sorted cells. 4 x 44k Whole genome expression arrays of FACS sorted HCT116 cancer stem cells (n=2) and HCT116 non-cancer stem cells (n=2) using total RNA obtained from FACS sorted cells.

first: Muzaffer

last: Ahmed Bhat

first: Sudip

last: Sen


title: Multipotent Progenitors Instruct Ontogeny of the Superior Colliculus – MADM-CloneSeq E10

overall_design: scRNA-Seq of neurons originating from a single radial glia progenitor (clone) labeled at E10 using MADM and Fz10-CreER or Sox2-CreER


title: Single nucleus gene expression profile of the rodent brains response to systemic innate immune system challenges

city: Vienna

first: Daniel

last: Bormann

state: Vienna

address: Waehringer Guertel 18-20

department: Department of Thoracic Surgery

institute: Medical University of Vienna

summary: The mammalian brain consists of heterogeneous cell populations with distinct responses to innate immune system challenges. Here we used single nucleus RNA sequencing (snRNAseq) to analyze cell type and stimulus dependent transcriptional responses of the rodent brain to peripheral LPS and Poly(I:C) challenges.

overall_design: Innate immune system challenges were induced by the intra peritoneal (i.p.) application of lipopolysaccharide (LPS), polyinosinic:polycytidylic acid (poly(I:C)), or saline (sham control) treatment. Four animals were pooled per treatment-group, generating 3 single nucleus RNA sequenced (snRNAseq) datasets: Saline treated = RN_brain_Saline, LPS treated = RN_brain_LPS, Poly(I:C) treated = RN_brain_PIC.  Seven hours after i.p. injection of LPS (5mg/kg bodyweight), poly(I:C) (12 mg/kg bodyweight), or saline brains were harvested for nuclei purification. Nuclei were isolated using nuclei extraction buffer (Miltenyi) and standard discontinuous iodixanol gradient separation, followed by library preparation using standard 10x Genomics workflows and snRNAseq (NovaSeq 6000, Illumina). Details on the study design and sample processing can be found in: Bormann, D., D. Copic, K. Klas, M. Direder, C. J. Riedl, G. Testa, H. Kühtreiber, E. Poreba, S. Hametner, B. Golabi, M. Salek, C. Haider, V. Endmayr, L. E. Shaw, R. Höftberger, H. J. Ankersmit and M. Mildner (2023). "Exploring the heterogeneous transcriptional response of the CNS to systemic LPS and Poly(I:C)." Neurobiology of Disease 188: 106339. https://doi.org/10.1016/j.nbd.2023.106339.

first: Dragan

last: Copic

first: Katharina

last: Klas

first: Martin

last: Direder

first: Christian

last: Riedl

first: Giulia

last: Testa

first: Hannes

last: Kühtreiber

first: Emilia

last: Poreba

last: Hametner

first: Bahar

last: Golabi

first: Melanie

last: Salek

first: Carmen

last: Haider

first: Verena

last: Endmayr

first: Lisa

last: Shaw

first: Romana

last: Höftberger

first: Hendrik

last: Ankersmit

first: Michael

last: Mildner


title: Single-cell Transcriptome analysis reveals cellular heterogeneity and highlights Fstl1-regulated alveolar myofibroblasts in mouse lung at birth

first: Wen

last: Ning

address: 94 Weijin Rd

institute: Nankai univercity

summary: The matricellular protein follistatin-like 1 (FSTL1) regulates lung development and saccular formation. Here, we employed single-cell RNA sequencing (scRNA-seq) to construct a transcriptomic atlas of 22,774 individual cells in wild-type (WT) and Fstl1–/– lung (E18.5) samples and identified 27 cell subtypes. We reveal abnormal population sizes and gene expression profiles in diverse cell subtypes in Fstl1–/– lung samples. We identify Pdgfra and Tgfbi as genetic markers specifically expressed in postnatal myofibroblasts (MyoFBs). Fstl1 deletion decreases the number of MyoFB cells and downregulates their roles in ECM organization and muscle tissue/vasculature development, in part, through the TGF-β1/BMP4 signaling pathway. Our data provide a single-cell view of the cellular heterogeneity and the molecular mechanisms underlying abnormal saccular formation and atelectatic lungs in Fstl1–/– mice.

overall_design: sc-RNA seq analysis of WT and Fstl1–/– lung mice at birth.

first: Si

last: Zhang

first: Xiuxue

last: Mo

first: Yueyue

last: Jin

first: Zhuan

last: Niu

first: Yue

first: Lian

last: Li

first: Gang

last: Hu