title: Purification of micrococcal nuclease (MNase) for use in ribosome profiling of high-salinity extremophiles

status: Public on Jun 22 2024

city: Baltimore

last: Isada

state: Maryland

address: 3400 N Charles St Levi 235

department: Biology

institute: Johns Hopkins University

summary: Nucleases, i.e. enzymes that catalyze the hydrolysis of phosphodiester bonds in nucleic acids, are essential tools in molecular biology and biotechnology. Staphylococcus aureus nuclease (MNase) is particularly interesting due to its thermostability and Ca2+-dependence, making it the prime choice for applications where nuclease modulation is critical, such as ribosome profiling in bacteria and halophilic archaea. The latter poses a technical and economical challenge, as high salt reaction conditions negatively impact MNase activity, necessitating large amounts of nuclease to be used for achieving efficient cleavage. Here, we set out to generate an optimized production protocol for two forms of MNase — fully processed MNaseA and the 19 aa propeptide containing MNaseB — and to biochemically benchmark them against a commercial nuclease. Our results show that both MNases are highly active in normal reaction conditions, but MNaseA maintains higher enzymatic activity in high salt concentrations than MNaseB. MNaseA also retains >90% of its activity after multiple freeze-thaw cycles when stored at -80°C in a buffer containing 5% glycerol. Importantly, ribosome profiling experiments in Haloferax volcanii demonstrated that MNaseA produces ribosome footprints highly comparable to those obtained with the commercial nuclease, making it a suitable alternative for high-salt ribosome profiling applications. In conclusion, our method can be easily implemented for efficient MNaseA production, thereby providing access to an effective, robust, and cost-efficient alternative to commercial nucleases, as well as facilitating future translation studies into halophilic organisms.

overall_design: We have generated a construct design and optimized expression conditions and biochemical assaying to produce and characterize MNaseA (abbreviated MNA) for robust and reproducible utilization in molecular biology methods that require the presence of high salt concentrations. Furthermore, we compared purified MNA to a commercially available nuclease (Roche MNase, abbreviated MNR) and, as a proof-of-concept, applied it for Ribo-seq of halophilic model archaeon H. volcanii (Hv). In brief, we harvested Hv culture, cryogenically lysed the cells, and isolated ribosomes from lysate. Ribosomes were then footprinted with either MNA or MNR, then separated along a sucrose gradient. Ribosome footprints were isolated from gradient fractions corresponding to single ribosomes and used for library preparation. Our subsequent analysis of ribosome footprints produced by either MNA or MNR digestion shows that MNA matches and exceeds the performance of its commercial counterpart. Consequently, this work provides a simple protocol for the efficient production of highly active MNaseA, making future Ribo-seq studies of halophilic organisms more accessible.

first: Pavlina

last: Gregorova

first: Jocelyne

last: DiRuggiero

first: L

middle: P

last: Sarin


title: Uncovering human yolk sac-derived innate lymphoid-biased multipotent progenitors prior to hematopoietic stem cell formation

status: Public on Jun 15 2024

city: Beijing

first: Guoju

last: You

state: Beijing

address: Shuangqing Road

country: China

institute: Tsinghua University

type: Expression profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Genomic and phenotypic stability of fusion-driven pediatric sarcoma cell lines [Infinium genotyping data]

status: Public on Jun 30 2024

city: Heidelberg

first: Thomas

middle: Georg Philipp

last: Grunewald

address: Im Neuenheimer Feld 280

department: Division of Translational Pediatric Sarcoma Research

country: Germany

institute: German Cancer Research Center (DKFZ)

type: Genome variation profiling by genome tiling array

summary:
Human cancer cell lines are the mainstay of cancer research. Recent reports showed that highly mutated adult carcinoma cell lines (mainly HeLa and MCF-7) present striking diversity across laboratories and that long-term continuous culturing results in genomic/transcriptomic heterogeneity with strong phenotypical implications. However, to what extent these limitations in terms of reproducibility may be generalized to all cancer cell lines remained unexplored. Here, we hypothesized that oligomutated pediatric sarcoma cell lines mainly driven by a fusion transcription factor, such as Ewing sarcoma (EwS), were genetically and phenotypically more stable than the previously investigated (adult) carcinoma cell lines. A comprehensive molecular and phenotypic characterization of multiple EwS cell line strains, together with a simultaneous analysis during 12 months of continuous cell culture showed that fusion-driven pediatric sarcoma cell line strains are genomically more stable than adult carcinoma strains, display remarkably stable and homogenous transcriptomes, and exhibit uniform and stable drug response. Additionally, the analysis of multiple EwS cell lines subjected to long-term continuous culture revealed that variable degrees of genomic/transcriptomic/phenotypic changes among fusion-driven cell lines, further exemplifying that the potential for reproducibility of in vitro scientific results may be rather understood as a spectrum, even within the same tumor entity.
When flasks reached approximately 70% confluency, samples were lysed, and total DNA was extracted with the NucleoSpin Tissue kit (Macherey Nagel) following the manufacturer’s protocol. For each sample, 900 ng of DNA in one technical replicate was used as input material and was profiled on Illumina Infinium Global Screening array

overall_design: Human cell lines were cultured continously over a period of 12 months and subjected to genotyping analysis at three different time points (m0=ITP, m6=MTP, m12=FTP)

first: Merve

last: Kasan

first: Florencia

last: Cidre-Aranaz

middle: G

last: Grünewald


title: Uncovering human yolk sac-derived innate lymphoid-biased multipotent progenitors prior to hematopoietic stem cell formation (scRNA-Seq)

summary: Hematopoietic stem cell (HSC)-independent lymphopoiesis has been elucidated in murine embryos. However, our understanding regarding human embryonic counterparts remains limited. Here, we unveiled the presence of human yolk sac-derived lymphoid-biased progenitors (YSLPs) expressing CD34, IL7R, LTB and IRF8 at Carnegie Stage 10, much earlier than the first HSCs emergence. The number and lymphopoietic potential of these progenitors were both significantly higher in yolk sac than embryo proper at this early stage. Importantly, single-cell/bulk culture and CITE-seq have elucidated the tendency of YSLP differentiating into innate lymphoid cells and dendritic cells. Notably, lymphoid progenitors in fetal liver before and after HSCs seeding displayed distinct transcriptional features, with the former closely resembling those of YSLPs.

overall_design: 7-AAD– cells from one CS10 YS (in vivo) were sorted for scRNA-seq (10X Genomics). All generated CD45+ cells from cultured CS12b-YS were sorted for scRNA-seq (10X Genomics). The generated cells from cultured CS12a-YS, CS12a-EP, CS12a-YS (CD45+CD127+) and CS12a-YS (CD45+CD127–) were combined with the following CITE-Seq antibodies: CD56, CD117, CD11b, CD161, CD5 and CD1a. Cells were individually hashtagged per group and pooled together.

first: Chen

last: Liu

first: Yandong

last: Gong

first: Xiaoyu

last: Su


title: Uncovering human yolk sac-derived innate lymphoid-biased multipotent progenitors prior to hematopoietic stem cell formation (CITE-Seq)

overall_design: The generated cells from cultured CS12a-YS, CS12a-EP, CS12a-YS (CD45+CD127+) and CS12a-YS (CD45+CD127–) were combined with the following CITE-Seq antibodies: CD56, CD117, CD11b, CD161, CD5 and CD1a. Cells were individually hashtagged per group and pooled together.


title: Lipid-Associated Macrophages Reshape BAT Cell Identity in Obesity [DBDB_singlecell]

status: Public on Jun 14 2024

city: Rome

first: Andrea

last: Ninni

address: Via della ricerca scientifica 1

country: Italy

institute: University of Rome "Tor Vergata"

summary: Obesity and type 2 diabetes cause a loss in brown adipose tissue (BAT) activity in mice and human, but the molecular mechanisms that drive BAT cell remodeling remain largely. Using a multilayered approach, we comprehensively map a deep reorganization in the BAT cells. We uncovered a subset of macrophages as the lipid-associated macrophages (LAM), which were massively increased in genetic and dietary model of BAT expansion. LAM  participate in this scenario by capturing extracellular vesicles carrying damaged lipids and mitochondria released from metabolically-stressed brown adipocytes. CD36 scavenger receptor drives LAM phenotype and through * in vitro* and *in vivo* models, we demonstrated that CD36-deficient LAM increased brown fat genes. LAM release Tgfb1 that reduces brown adipocytes identity through Aldh1a1 induction. This study provides the first description of cell dynamics in BAT of obese models identifying LAM as responder to tissue-level metabolic stress and key driver to loss of BAT cell identity.

overall_design: Brown adipose tissue samples from 8-weeks and 16-weeks old WT, db/db and HFD mice with at least three biological replicates were used for expression profiling.

first: Daniele

last: Lettieri Barbato

first: Fabio

last: Zaccaria


title: Lipid-Associated Macrophages Reshape BAT Cell Identity in Obesity [DBDB_Bulk]

overall_design: Brown adipose tissue samples from 8-weeks WT and db mice with at least three biological replicates were used for expression profiling.


title: Lipid-Associated Macrophages Reshape BAT Cell Identity in Obesity [CD36_Bulk]

overall_design: Brown adipose tissue samples from 8-weeks WT and Cd36KO mice with at least three biological replicates were used for expression profiling.


title: Transcriptomic analysis between Red-Flesh and White-Flesh Mesocarps in Red-Flesh apple at 84 Days After Full Bloom (DAFB)

status: Public on Jun 11 2024

city: Beuacouzé

first: Jean-Marc

last: Celton

address: 42 ue Georges Morel

country: France

institute: IRHS

summary: To functionally characterise flesh pigmentation pattern, we performed transcriptomic comparisons between RF and WF mesocarps. The transition step between fully red fruits and heterogeneously pigmented fruit were targeted during fruit development (84 Days After Full Bloom - DAFB).

overall_design: Comparative gene expression analysis between white-flesh and red-flesh mesocarps in red-flesh apple

first: Pierre

last: Bouillon

first: Sandrine

last: Balzergue


title: Lipid-Associated Macrophages Reshape BAT Cell Identity in Obesity [ATGL_Bulk]

overall_design: Brown adipose tissue samples from 10-weeks WT and Ucp1Δ/Δ mice with at least three biological replicates were used for expression profiling.


title: CircRNA sequencing data of tumors and paired adjacent tissue from HCC patients

status: Public on Jun 10 2024

city: shanghai

first: xindong

last: wei

address: 1200 Cailun Road, Shanghai

institute: shanghai university of Chinese medicine

type: Non-coding RNA profiling by high throughput sequencing

summary: CircRNAs have emerged as crucial contributors to cancer progression. Nonetheless, the expression regulation, biological functions, and underlying mechanisms of circRNAs in mediating hepatocellular caicinoma(HCC) progression remain insuffuciently elucidated. In this study, we generated the circular RNA expression profile of HCC tissue together with the adjacent normal tissues by high-throughput RNA sequencing.

overall_design: Circular RNA sequencing data of 5 tumors and paired adjacent tissues from HCC patients.

first: Xindong

last: Wei

first: Anfeng

last: Si


title: Single-cell sequencing of full-length transcripts and T-cell receptors with automated high-throughput Smart-seq3 [HT SS3]

status: Public on Jul 02 2024

city: Foster City

first: Ruidong

last: Li

state: CA

address: 324 Lakeside Drive

department: Research Data Sciences

country: USA

institute: Gilead Sciences, Inc.

summary: We developed an automated high-throughput Smart-seq3 (HT Smart-seq3) workflow via robotic implementation and established best practices to consistently achieve high cell capture efficiency and data quality. In comparison with the 10X platform, HT Smart-seq3 analysis of primary CD4+ T-cells demonstrated superior sensitivity in gene detection and similar capability to capture major cellular heterogeneity upon sufficient scaling up. Notably, through T-cell receptor (TCR) reconstruction, HT Smart-seq3 identified more productive pairs of alpha and beta chains without additional primer design, enabling more comprehensive profiling of TCRs. Collectively, HT Smart-seq3 provides a cost-effective and scalable method for characterization of single-cell transcriptomes and immune repertoires.

overall_design: PPBMCs were isolated from two healthy donors and subsequently treated with either DMSO (vehicle) or PMA/Ionomycin. After a 3-hour incubation, FACS-sorted CD4+ T-cells were collected for single-cell library preparation using the HT Smart-seq3 protocol.

first: Hsiu-Chun

last: Chuang

first: Kai-Hui

last: Sun

first: Li


title: Effect of knock-down of circular RNA circUCK2 on gene expression in PLC/PRF/5 cell

summary: CircRNAs have emerged as crucial contributors to cancer progression. Nonetheless, the expression regulation, biological functions, and underlying mechanisms of circRNAs in mediating hepatocellular caicinoma(HCC) progression remain insuffuciently elucidated. Here we uncovered a novol circular RNA, circUCK2, which is highly expressed in HCC tissues and associated with recurrence-free survival in HCC patients. Knockdown of circUCK2 inhibits proliferation and metastasis of HCC cell lines.To investigate the function of circUCK2-silence in HCCs, we performed gene expression profiling analysis using data obtained from RNA-seq of PLC/PRF/5 cells.

overall_design: PLC/PRF/5 cells were treated with100 nM CircUCK2 siRNA and Scramble siRNA for 24 hours.  Total RNA was harvested, sequencing was performed in three biological replicates.


title: PI3K/mTOR dual inhibitor GSK458 and arsenic trioxide exert synergistic anti-tumor effects against ovarian clear cell carcinoma

status: Public on Jun 18 2024

city: SUITA

first: WANG

last: YAN

state: OSAKA

address: 2-2 YAMADAOKA

country: Japan

institute: Osaka University

summary: Ovarian carcinoma is a significant cause of death among all gynecologic malignancies. OCCC originates from endometriosis with recurrent somatic mutations, especially in phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA). To testify the key enrichment pathway of combination of GSK458 with As2O3 in OCCC therapy, we use RNA sequence of four therapy groups (Control group, As2O3 (1uM) monotherapy, GSK458 (0.1uM) monotherapy, As2O3 combined with GSK458 group for 48h) in TOV21G cell line.

overall_design: The groups were divided into four groups-Control group, As2O3 (1uM) monotherapy, GSK458 (0.1uM) monotherapy, As2O3 combined with GSK458 group for 48h. Total RNA was extracted using the Qiagen RNeasy Mini Kit (Qiagen, Venlo, Netherlands). RNA integrity was evaluated using the Agilent 2100 Bioanalyzer RNA Nano Chip (Agilent Technologies, Santa Clara, CA, USA). There is one sample for each group. Totally there are four samples from four therapy groups.

first: Yan

last: Wang

first: Yasuto

last: Kinose


title: Regulatory mechanism of Sox9 gene expression in chondrocytes [ChIP-seq]

city: Suita

first: Kenji

last: Hata

state: Osaka

address: 1-8 Yamadaoka

department: Biochemistry

institute: Osaka University graduate school of dentistry

type: Genome binding/occupancy profiling by high throughput sequencing

summary: The transcription factor Sox9 (Sry-Related HMG-Box gene 9) is essential in chondrogenesis. The expression Sox9 is regulated by 1.5Mb long range genomic region in temporospatial manner and genome translocation or deletion which affect Sox9 expression cause campomelic dysplasia (CD), characterized by skeletal malformations. Although functional mechanism of Sox9 are well studied so far, regulatory mechanism of Sox9 expression and its related enhancer in chondrocytes are largely remains to be elucidated. The purpose of this study is to identify Sox9 enhancers in chondrocytes

overall_design: ChIP-seq

first: Sachi

last: Kobayashi


title: CD22 modulation regulates neuroinflammation in microglia

city: Hong Kong

first: Jianlin

address: L 7-08, Department of Obstetrics and Gynecology  Laboratory block, 21th Sassoon Road, Hong Kong

country: Hong Kong

institute: The University of Hong Kong

summary: Expression data comparing the effect of soluble CD22 in monocyte-derived microlgia and in mouse.  Expression data on the effect of SM03 in Aβ-injected AD model in human CD22 transgenic mice.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

first: Chi

middle: H

last: Chong


title: Regulatory mechanism of Sox9 gene expression in chondrocytes [RNA-seq]

overall_design: RNA-seq


title: Regulatory mechanism of Sox9 gene expression in chondrocytes


title: Regulatory mechanism of Sox9 gene expression in chondrocytes [ATAC-seq]

overall_design: ATAC-seq


title: PanethcellTNF-signalinginducesbacterialsepsis: PC transcriptome 15h ater TNF in IFNARKO mice

status: Public on Jun 03 2024

city: Gent

first: Steven

last: Timmermans

address: Technologiepark 71

country: Belgium

institute: VIB-UGent

summary: Paneth Cells (PCs) are secretory cells located in the crypts of Lieberkühn of the small intestine. They produce antimicrobial peptides (AMPs) and are thought to keep the microbiome under control. The cytokine TNF, which functions mainly via its TNF receptor 1 (TNFR1 or P55), is a driving force in many inflammatory bowel disease (IBD) patients, but its impact on PCs is poorly known. We have generated PC-specific P55 knockout mice (P55Paneth KO) and found that these mice are protected against lethal TNF-induced systemic inflammatory response syndrome (SIRS). The impact of TNF on PCs is characterized by morphological changes and by intestinal bacterial translocation into organs, such as liver, in which these bacteria induce expression of numerous genes, associated with typical sepsis signatures. The presence of these bacteria cause lethality of the mice. Based on bulk RNA-SEQ on pure PCs, P55 signaling stimulated mainly a type-I IFN response in the PCs, which drives the bacterial translocation and lethality via IFNAR1. Interestingly, based on ER stress reporter mice, PCs constitutively have high amounts of spliced XBP1 (XBP1s) protein, presumably to support their secretion of AMPs, and TNF eradicates this XBP1s production, leading to failure of the unfolded protein response (UPR). This causes reduced antimicrobial activity in PCs, in a P55 and IFNAR1-dependent way. Our data suggest a novel axis induced by TNF in PCs, leading to interferon-induced UPR failure causing lethal bacterial translocation to the liver (and other organs)

overall_design: To investigate the effect of TNF on the paneth cell (PC) transcriptome, PCs were isolated 15h after TNF or PBS (control) injection in mice using cell sorting. 3 PBS and 3 TNF samples were taken (biological replicates; each from a different mice) for RNA isolation and sequencing. All mice were KO for Ifnar1

first: Charlotte

last: Wallaeys

first: Natalia

last: Garcia-Gonzalez

first: Jolien

last: Vandewalle

first: Tineke

last: Vanderhaeghen

first: Hester

last: Dufoor

first: Melanie

last: Eggermont

first: Riet

last: De Rycke

first: Serge

last: Verbanck

first: George

last: Kollias

first: Richard

middle: S

last: Blumberg

first: Takao

last: Iwawaki

first: Claude

last: Libert


title: Neutrophils-derived itaconate facilitates viral infection via alkylating GDI2 II

status: Public on Jun 01 2024

city: Shanghai

first: Shulei

last: Yin

state: Shanghai

address: No.800 Xiangyin Road, Yangpu District

department: Department of Immunology

institute: Naval Medical University

summary: The objective was to investigate the disparity in gene expression between IRG1+/+ BMDMs and IRG1-/- BMDMs following VSV infection.

overall_design: IRG1+/+ and IRG1-/- BMDMs were infected with VSV for 8 hours, and then subjected to RNA-seq.

first: S

first: Y

last: Tao

last: Xu

last: Yu


title: LXRalpha Controls Metabolism Associated Steatotic Hepatitis (MASH)

status: Public on Jun 06 2024

city: CHARLOTTESVILLE

first: Ira

middle: Glenn

last: Schulman

state: Virginia

address: 1340 Jefferson Park Ave, Jordan Hall, room 5218

department: Pharmacology

institute: University of Virginia

summary: Liver x receptor alpha (LXRα, Nr1h3) functions as an important intracellular cholesterol sensor that regulates liver fat and cholesterol metabolism at the transcriptional level in response to the direct binding of cholesterol derivatives. We have generated mice with a mutation in LXRα that reduces activity in response to endogenous cholesterol derived LXR ligands while still allowing transcriptional activation by synthetic agonists. The mutant LXRα functions as a dominant negative that shuts down cholesterol sensing. When fed a high fat, high cholesterol diet LXRα mutant mice rapidly develop pathologies associated with Metabolic Dysfunction-Associated Steatohepatitis (MASH) including ballooning hepatocytes, liver inflammation, and fibrosis. Strikingly LXRα mutant mice have decreased liver triglycerides but increased liver cholesterol. Therefore, MASH-like phenotypes can arise in the absence of large increases in hepatic triglycerides. Reengaging LXR signaling by treatment with synthetic agonist reverses MASH suggesting that LXRα normally functions to impede the development of liver disease.

overall_design: CRISR was used to generate mice with with mutation of tryptophan (W) 441 of the LXRalpha (Nr1h2) to phenylalanine (F). Liver RNA was isolated from homozygous controls (W/W), heterozygous mutants (W/F), homozygous mutants (F/F) that were maintained on either a normal chow diet or high fat, high cholesterol MASH promoting diet for 4 week. Mice on the high cholesterol diet were treated with daily with the LXR agonist T0901317 or with vehicle for the final 2 week

first: Alexis

middle: T

last: Clark


title: Microglia modulate the cerebrovascular reactivity through ectonucleotidase CD39

city: Charlottesville

first: Anwaruddin

last: Mohammad

state: VA

address: Pinn Hall

department: Bioinformatics Core

summary: Microglia and the border-associated macrophages contribute to the modulation of cerebral blood flow, but the mechanisms have remained uncertain. Here, we show that microglia regulate the cerebral blood flow baseline and the responses to whisker stimulation or intra-cisternal magna  injection of adenosine triphosphate, but not intra-cisternal magna injection of adenosine in mice model. Notably, microglia repopulation corrects these cerebral blood flow anomalies. The microglial-dependent regulation of cerebral blood flow requires the adenosine triphosphate-sensing P2RY12 receptor and ectonucleotidase CD39 that initiates the dephosphorylation of extracellular adenosine triphosphate into adenosine in both male and female mice. Pharmacological inhibition or CX3CR1-CreER-mediated deletion of CD39 mimics the cerebral blood flow anomalies in microglia-deficient mice and reduces the upsurges of extracellular adenosine following whisker stimulation. Together, these results suggest that the microglial CD39-initiated breakdown of extracellular adenosine triphosphate co-transmitter is an important step in neurovascular coupling and the regulation of cerebrovascular reactivity.

overall_design: Three groups are included. Five mice were included in each group. The first group is control group, in which mice were treated with control food. The second group is PLX3397 food treated mice group. In this group microglia were depleted by CSF1r inhibitor. The third group is repopulation group. In this group, mice were first treated with PLX3397 to deplete microglia and then put back to control food for microglia to repopulate.

first: Zhongxiao

last: Fu

first: Mallikarjunarao

last: Ganesana

first: Philip

last: Hwang

first: Xiao

last: Tan

first: Melissa

middle: M

last: Kinkaid

first: Yu-Yo

last: Sun

first: Emily

last: Bian

first: Aden

last: Weybright

first: Katia

last: Sol-Church

first: Ukpong

middle: B

last: Eyo

first: Clare

last: Pridans

first: Simon

middle: C

last: Robson

first: Pankaj

last: Kumar

first: Francisco

middle: J

last: Quintana

first: B

last: Venton

first: Anne

last: Schaefer

first: Chia

middle: Y

last: Kuan


title: Artesunate overcomes the resistance of basal type of bladder cancer to RC48

city: Tianjin

first: Jingxian

last: Li

address: Pingjiang Road 23#, Hexi District, Tianjin

institute: Tianjin institute of urology

summary: ERBB2 expression mediates sensitivity of bladder cancer cells to RC48. JAK/STAT3 pathways compensatory activation after exposing RC48. A combination of STAT3 inhibitor-artesunate and RC48 could significantly inhibit basal bladder cancer growth. These findings provided a theoretical foundation for subsequent individualized therapy for MIBC based on the molecular types.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for SW780 and RC48-exposed SW780 cells.

first: Kun

last: Shan

first: Jianqiang

last: Zhu

first: E

last: Du


title: Comparative single-cell multiome identifies evolutionary changes in neural progenitor cells during primate brain development

status: Public on Jun 09 2024

first: Yiming

address: NO.5, Yiheyuan Road, Haidian District

department: School of Life Sciences

institute: Peking University

summary: Understanding the cellular and genetic mechanisms driving human-specific features of cortical development remains a challenge. We generated a cell-type resolved atlas of transcriptome and chromatin accessibility in the developing macaque and mouse prefrontal cortex (PFC). Comparing with published human data, our findings demonstrate that although the cortex cellular composition is overall conserved across species, progenitor cells show significant evolutionary divergence in cellular properties. Specifically, human neural progenitors exhibit extensive transcriptional rewiring in growth factor and extracellular matrix (ECM) pathways. Expression of the human-specific progenitor marker ITGA2 in the fetal mouse cortex increases the progenitor proliferation and the proportion of upper-layer neurons. These transcriptional divergences are primarily driven by altered activity in the distal regulatory elements. The chromatin regions with human-gained accessibility are enriched with human-specific sequence changes and polymorphisms linked to intelligence and neuropsychiatric disorders. Our results identify evolutionary changes in neural progenitors and putative gene regulatory mechanisms shaping primate brain evolution.

sample_organism:
Macaca mulatta
Mus musculus

platform_organism:
Macaca mulatta
Mus musculus

overall_design: To investigate the cellular and genetic mechanism of the human specifications in cortex development, We generated a comprehensive cell atlas of transcriptome and chromatin accessibility for macaque and mouse prefrontal cortex (PFC) during mid-digestion using snRNA-seq, scATAC-seq and snMultiome-seq.

first: Yuting

last: Liu

first: Xin

last: Luo

first: Ting

last: Hu

first: Kaimin

last: Chen

first: Jiahao

last: Xu

first: Fengyun

last: Zhang

first: Xiaoyu

last: Meng

first: Xiang

first: Xiechao

last: He

first: Cheng

first: Bing

last: Su


title: Transcriptional heterogeneity of HCC1143 Triple Negative Breast Cancer perturbed with paclitaxel or soluble ligand

status: Public on Jun 04 2024

city: Portland

first: Laura

last: Heiser

state: Oregon

address: 2730 S. Moody Ave

department: BME

institute: OHSU

summary: Triple Negative Breast cancer is a heterogeneous disease that is capable of adapting to a variety of stimuli. To study the dynamics of both drug and ligand response, we performed scRNA-seq to deeply profiled the transcriptional response of the HCC1143 cell line to a variety of stimuli.

overall_design: HCC1143 cells were grown in vitro and treated for 24 or 72 hours with soluble perturbation.

first: Nicholas

middle: L

last: Calistri

first: Tiera

middle: A

last: Liby

first: Zhi

first: Hongmei

first: Mark

last: Dane

first: Sean

last: Gross


title: FGF7 as an essential mediator for the onset of ankylosing arthropathy related to psoriatic skin lesions

city: Sendai

first: Shin

last: Ebihara

address: 2-1 Seiryo, Aoba, Sendai, Miyagi 980-8575

department: 1)	Department of Pathology

institute: Tohoku University

summary: IL-17A plays an important role in the pathology of psoriasis and psoriatic arthritis (PsA). However, the pathogenic association between the skin and joint manifestations in PsA is not completely understood. In this study, we initially observed that IL-17A and fibroblast growth factor (FGF) 7 induced endochondral ossification in the mouse entheseal histoculture. Importantly, the responses of endochondral ossification by IL-17A stimulation were strongly inhibited by the treatment of a blocking antibody to FGF receptor 2IIIb (FGFR2IIIb), which is the receptor of FGF7, suggesting that FGF7 acts as a downstream factor of IL-17A in the endochondral ossification in the culture. Next, using the animal PsA model, the administration of anti-FGFR2IIIb antibody resulted in significant suppression of ankylosing enthesitis but not dermatitis. Collectively, our findings indicate that augmented IL-17A in PsA dermatitis induces the elevation of FGF7 levels in joint enthesis, and results in the development of ankylosing enthesitis via FGF7 signaling in PsA.

overall_design: Gene expression analysis of entheseal tissues of C57BL/6J mice. Organs were stimulated with or without IL-17A (10 ng/mL) for 72 h (n = 3 per sample).

first: Yuji

last: Owada

first: Masao

last: Ono


title: Genomic and phenotypic stability of fusion-driven pediatric sarcoma cell lines

status: Public on May 31 2024

first: Thomas

last: Grunewald

summary: Human cancer cell lines are the mainstay of cancer research. Recent reports showed that highly mutated adult carcinoma cell lines (mainly HeLa and MCF-7) present striking diversity across laboratories and that long-term continuous culturing results in genomic/transcriptomic heterogeneity with strong phenotypical implications. However, to what extent these limitations in terms of reproducibility may be generalized to all cancer cell lines remained unexplored. Here, we hypothesized that oligomutated pediatric sarcoma cell lines mainly driven by a fusion transcription factor, such as Ewing sarcoma (EwS), were genetically and phenotypically more stable than the previously investigated (adult) carcinoma cell lines. A comprehensive molecular and phenotypic characterization of multiple EwS cell line strains, together with a simultaneous analysis during 12 months of continuous cell culture showed that fusion-driven pediatric sarcoma cell line strains are genomically more stable than adult carcinoma strains, display remarkably stable and homogenous transcriptomes, and exhibit uniform and stable drug response. Additionally, the analysis of multiple EwS cell lines subjected to long-term continuous culture revealed that variable degrees of genomic/transcriptomic/phenotypic changes among fusion-driven cell lines, further exemplifying that the potential for reproducibility of in vitro scientific results may be rather understood as a spectrum, even within the same tumor entity.

overall_design:
When flasks reached approximately 70% confluency, total RNA was isolated using the NucleoSpin RNA kit (Macherey-Nagel, Germany) according to the manufacturer’s protocol. RNA quality was verified on a Nanodrop Spectrophotometer ND-1000 (Thermo Fischer) and quantity was measured on a Qubit instrument (Life Technologies). For each sample, 50–100 ng of RNA in three biological and two technical replicates were used as input material and were profiled on an Illumina NextSeq 500 system.
Human cell lines were cultured continously over a period of 12 months and subjected to RNA-seq analysis at three different time points (m0=ITP, m6=MTP, m12=FTP)

first: Merve

last: Kasan

first: Florencia

last: Cidre-Aranaz

last: Grünewald


title: Effects of Irbesartan on Gene Expression Profiles in Kidney Tissues of Diabetic db/db Mice

status: Public on May 29 2024

first: Haojun

address: 2 YingHua Dongjie, Chaoyang District

department: Institute of Clinical Medical Sciences

institute: China-Japan Friendship Hospital

summary:
Diabetic nephropathy(DN) is a common diabetic microvascular complication, Irbesartan is the first-line drug for clinical treatment of diabetic nephropathy, but its pharmacological mechanism and target are not yet fully clear.
We used microarrays to explore the global profile of gene expression for better understanding the action mechanism of irbesartan in alleviating renal injuries of diabetic db/db mice

overall_design: Ten-week-old male db/db mice were orally administrated with irbsartan or vehcle for 12 weeks,the kidneys were harvested for RNA extraction and hybridization on Affymetrix microarrays.

first: Hailing

last: Zhao

first: Ping

last: Li


title: Mis-splicing of mitotic regulators sensitizes SF3B1-mutated human HSCs to CHK1 inhibition [RNA-seq]

city: Pavia

first: Martina

last: Sarchi

address: Via Ferrata 9

department: Dept. of Molecular Medicine

institute: University of Pavia

summary: Splicing factor SF3B1 mutations are frequent somatic lesions in myeloid neoplasms that transform hematopoietic stem cells (HSCs) by inducing mis-splicing of target genes. However, the molecular and functional consequences of SF3B1 mutations in human HSCs remain unclear. Here, we identify the mis-splicing program in human HSCs as a targetable vulnerability by precise gene editing of SF3B1 K700E mutations in primary CD34+ cells. Mutant SF3B1 induced pervasive mis-splicing and reduced expression of genes regulating mitosis in single cell transcriptomics, critically BUBR1 and CDC27, leading to altered differentiation and delayed G2/M progression. Mutant SF3B1 mis-splicing or reduced expression of BUBR1 and CDC27 was sufficient to delay G2/M transit, leading to activation of CHK1, sensitizing cells to CHK1 inhibition. Clinical CHK1 inhibitor prexasertib selectively targeted SF3B1-mutant HSCs and abrogated engraftment in vivo. These findings identify mis-splicing of mitotic regulators in SF3B1-mutant HSCs as a targetable vulnerability engaged by pharmacological CHK1 inhibition.

overall_design: RNA-seq was performed on FACS-sorted BFP+CD34+ gene edited HSPCs and clonally derived K562 cells. For each cell type, transcriptome (gene expression and splicing) of SF3B1 mutant cells was compared to wild-type gene edited controls.

first: Courtnee

last: Clough

first: Edie

middle: I

last: Crosse

first: Jason

last: Kim

middle: D

last: Baquero Galvis

first: Nelli

last: Aydinyan

first: Rachel

last: Wellington

first: Anna

last: Gallì

first: J

last: Creamer

first: Sintra

last: Stewart

first: Robert

middle: K

last: Bradley

first: Luca

last: Malcovati

first: Sergei

last: Doulatov


title: Transcriptional analysis of cDC subsets in Irf8+32 -/- and Zeb2 -165(d1+2+3) mice

city: SAINT LOUIS

first: Suin

last: Jo

state: MO

address: 4939 Childrens PI

department: Division of Biology & Biomedical Sciences

institute: Washington University in St. Louis

summary: To determine if the residual cDC subsets in the Δ1+2+3 and Δ32 mice reflected their normal counterparts, we performed bulk RNA-sequencing analysis of cDC1 and cDC2 from WT, Δ1+2+3 and Δ32 mice. cDC1 from WT and Δ1+2+3 clustered together in principal component analysis (PCA) plot, whereas cDC2 from WT and Δ32 mice cluster together. Further, cDC1 and cDC2 from WT mice show a large number of differentially expressed genes (DEGs), as expected. However, there were few DEGs in cDC1 between WT and Δ1+2+3 mice, or in cDC2 between WT and Δ32 mice, indicating no substantive transcriptional differences between corresponding cDC subsets in WT, Δ32 and Δ1+2+3 mice.

overall_design: mRNA profiles of splenic cDC1 and cDC2 isolated from WT, Irf8 +32-/-, or Zeb2 -165(d1+2+3) mice

first: Tiantian

last: Liu

first: Theresa

last: Murphy

first: Kenneth


title: Effect of targeting NOS2 on gene expression of triple-negative breast cancer (metaplastic) SUM159 cell lines

status: Public on May 28 2024

city: HOUSTON

first: HONG

last: ZHAO

state: Texas

address: 6670 Bertner Ave

institute: HOUSTON METHODIST HOSPITAL

summary: To investigate the role of iNOS in tumor cell morphology and gene expression, we utilized CRISPR-Cas9 gene editing (NOS2 Double Nickase Plasmid, Santa Cruz) to knockout NOS2 in SUM159 cells. After puromycin selection, resistant clones were selected by immunoblotting for iNOS expression and nitrite/nitrate colorimetric assay. We then performed gene expression profiling analysis using data obtained from RNA-seq of parental SUM159 cells compared to NOS2KO SUM159 cells.

overall_design: RNA-seq of parental SUM159 cells compared to three clones of NOS2KO SUM159 cells.

first: Tejaswini

last: Reddy

first: Hong

last: Zhao


title: Human skeletal muscle possesses an epigenetic memory of high intensity interval training

status: Public on May 30 2024

city: Oslo

first: Adam

middle: P

last: Sharples

address: 220 Sognsveien

country: Norway

institute: Norwegian School of Sport Sciences

summary: Twenty healthy subjects (25±5yrs) completed two high-intensity interval training interventions (training and retraining) lasting 8 weeks separated by 12 weeks of detraining. Measurements at baseline and after training, detraining and retraining included maximal oxygen consumption (V̇O2max), along with vastus lateralis biopsy for genome wide DNA methylation using Illumina Epic arrays in 5 of the participants for all conditions (baseline, training, detraining and retraining).

overall_design:
MATERIALS AND METHODS
Subjects:
The study intervention was conducted at the University of Pavia, Italy. Twenty male (n = 11) and female (n = 9) adults were recruited from the local community. Inclusion criteria included: To have not been involved in structured training programs before; maximal oxygen consumption (V ̇O2max), determined by an initial screening test, lower than 40 and 45 ml·min−1·kg−1 for women and men, respectively. Exclusion criteria specified that all participants were free from major diseases at the pulmonary, cardiovascular and muscle level and they were not taking any drugs. All procedures were in accordance with the Declaration of Helsinki and the study was approved by the local ethics committee (Besta 64-19/07/2019) and participants gave their written informed consent after being informed about objectives, methods and risks of the study. Below are the participants anthropometrics and physiological characteristics measured during the initial screening test grouped according to sex. V ̇O2max, maximal oxygen uptake. All values are means ± SD.    Males (n = 11), Age (yrs) 27 ± 5, Weight (kg) 72.1 ± 10.7, Height (cm)177 ± 5, V ̇O2max (ml·min−1·kg−1) 36.8 ± 8.3, V ̇O2max (l·min−1) 2.601 ± 0.448    Females (n = 9), Age (yrs) 23 ± 3, Weight (kg) 58.7 ± 7.2, Height (cm)167 ± 4, V ̇O2max  (ml·min−1·kg−1), 34.1 ± 5.3, V ̇O2max (l·min−1) 1.985 ± 0.308

Study design:  The current study exposed the same individuals to two identical 8-week high-intensity interval training periods separated by a three-month washout period during which participants were instructed to return to their habitual life. Levels of physical activity were monitored before and throughout the entire period of the study by accelerometers (wGT3X-BT, ActiGraph, Florida, USA). Data collection was performed at four different time points: at baseline (BASELINE), after the first training period (TRAINING), following detraining (DETRAINING) and after retraining (RETRAINING). For each time point participants visited the laboratory on two separate occasions. During the first visit, anthropometric measurements were collected, and an incremental cycle ergometer exercise test up to voluntary exhaustion was performed. On the second visit, approximately 100 mg of skeletal muscle tissue was obtained from the vastus lateralis muscle under local anesthesia (1% lidocaine) for genome wide DNA methylation (methylome), targeted gene expression based on genes that demonstrated significantly differentially methylation positions (DMPs) or regions (DMRs) (described below) and protein abundance based on molecular pathways demonstrating retained epigenetic and or transcriptional retention. Participants were instructed to abstain from strenuous physical activity for at least 48 h prior to each testing session.

Training:  Exercise prescription consisted of two identical 8-week periods of high-intensity interval training (HIIT) based on individual results obtained from the exercise incremental tests. The first three training sessions were conducted under the supervision of a researcher, who oversaw giving adequate instructions about the training procedures. The remaining training sessions (3/week) were conducted at home on electronically braked cycle ergometer. Time, heart rate (HR) and power (W) from each session were recorded and uploaded on a web-based app (MyZone, England, UK) to allow the researchers to check the adherence to the training regimen. Every week the subjects were contacted by phone to monitor progression, provide feedback and encouragement, and to answer any questions. Exercise sessions were composed by 10 min of warm-up followed by high-intensity bouts of either short (1-2 min) or long (3-4 min) duration. Exercise prescription varied during the training period to facilitate participant motivation and compliance, as detailed previously (Robach et al., 2014). To avoid stagnation (Granata et al., 2016a) the training stimulus (power and repetitions) was progressively incremented. An identical training protocol was applied for both training and retraining.

Incremental exercise:  Incremental exercise was performed on an electronically braked cycle ergometer (818E, Monark, Sweden) to determine maximal oxygen consumption (V ̇O2max), gas exchange threshold (GET), and respiratory compensation point (RCP). Pedaling frequency was digitally displayed to the subjects, and subjects were asked to keep a constant cadence throughout the tests between 70 and 80 rpm. Power was increased 10-15 W every minute, depending on the individual’s fitness aiming to allow the subjects to reach voluntary exhaustion in 10–15 min. Voluntary exhaustion was defined as the incapacity to maintain pedaling frequency for 5 s at the imposed work rate despite vigorous encouragement by the researchers. After 30 min of recovery, all participants performed a verification trial consisting of a constant work rate exercise at 90% of the highest power attained in the incremental test (Wmax) (Rossiter et al, 2006).    Pulmonary ventilation (VE, in BTPS [body temperature (37°C), ambient pressure and gas saturated with water vapor]), oxygen consumption (V ̇O2), and CO2 output (V ̇CO2), both in STPD (standard temperature [0°C or 273 K] and pressure [760 mmHg] and dry [no water vapor]), were determined breath-by-breath by a metabolic cart (Vyntus CPX, Vyaire Medical GmbH, Germany). Before each test, gas analyzers were calibrated with ambient air and a gas mixture of known concentration (O2: 16%, CO2: 4%) and the turbine flowmeter was calibrated with a 3-L syringe at three different flow rates. RER was calculated as V ̇CO2/V ̇O2. HR was recorded by using a HR chest band (HRM-Dual, Garmin, Kansas, USA). Rating of perceived exertion (RPE) was determined using Borg® 6–20 scale every 2 min through the test (Borg, 1982). At rest, and at 1, 3, and 5 min of recovery, 20 μL of capillary blood was obtained from a preheated earlobe for blood lactate concentration (Biosen C-line, EKF, Germany); the analyzer was frequently calibrated with a standard solution containing 12 mmol·L−1 of lactate. The highest 20-s averaged cardiopulmonary and metabolic data recorded during the whole incremental test (i.e. including the verification phase) were taken as maximal values (Martin-Rincon et al. 2019). GET was determined by two independent investigators by using the modified “V-slope” method (Beaver et al., 1986) and “secondary criteria”. RCP was determined as the point where end-tidal PCO2 began to fall after a period of isocapnic buffering (Whipp et al, 1989). This point was confirmed by examining V ̇E/V ̇CO2 plotted against V ̇O2 and by identifying the sec breakpoint in the V ̇E-to-V ̇O2 relation.    Muscle Biopsy  Fifteen participants out of 20 [male (n = 9) and female (n = 6)] agreed to undergo muscle biopsy for all the time points required by the experimental protocol. Resting muscle biopsies were taken from the vastus lateralis muscle using a 130 mm (6”) Weil-Blakesley rongeur (NDB-2, Fehling Instruments, GmbH&Co, Germany) under local anesthesia (1% lidocaine). After collection, muscle samples were cleaned of excess blood, fat, and connective tissue in ice-cold BioPS, a biopsy-preserving solution containing (in mM) 2.77 CaK2EGTA, 7.23 K2EGTA, 5.77 Na2ATP, 6.56 MgCl2, 20 taurine, 50 MES (2-(N-morpholino) ethanesulfonic acid), 15 Na2phosphocreatine, 20 imidazole, and 0.5 dithiothreitol adjusted to pH 7.1 (Doerrier et al., 2018). A portion of ~20 mg of muscle tissue was immediately submerged in Allprotect Tissue Reagent (Qiagen, Netherlands) following the manufacturer’s instructions to stabilize and protect cellular DNA and was stored at -20°C for subsequent DNA methylome analysis (described below). A specimen of ~20-30 mg was rapidly frozen by immersion in liquid nitrogen and stored at -80°C for RNA isolation and gene expression analysis (described below). Finally, a portion of ~20-30 mg from each muscle sample was rapidly frozen by immersion in liquid nitrogen and stored at -80°C for protein analysis. Fifteen out of 15 subjects were analyzed for RNA/gene expression, 9 subjects were analyzed for protein abundance and 5 subjects per time point (BASELINE, TRAINING, DETRAINING, RETRAINING) for DNA methylome analysis.
Methylation analysis:  Five (n = 5) subjects across all time points of BASELINE, TRAINING, DETRAINING, RETRAINING were analyzed in the present data set for DNA methylome analysis. Tissue Homogenization, DNA Isolation, and Bisulfite Conversion. Tissue samples were homogenized for 45 s at 6,000 rpm × 3 (5 min on ice in between intervals) in lysis buffer (180 μl buffer ATL with 20 μl proteinase K) provided in the DNeasy spin column kit (Qiagen, United Kingdom) using a Roche Magnalyser instrument and homogenization tubes containing ceramic beads (Roche, United Kingdom). The DNA was then bisulfite converted using the EZ DNA Methylation Kit (Zymo Research, CA, United States) as per manufacturer’s instructions.   Infinium Methylation EPIC Beadchip array. All DNA methylation experiments were performed in accordance with Illumina manufacturer instructions for the Infinium Methylation EPIC 850K BeadChip Array (Illumina, USA). Methods for the amplification, fragmentation, precipitation and resuspension of amplified DNA, hybridization to EPIC BeadChip, extension and staining of the bisulfite converted DNA was conducted as detailed in paper from Seaborne and colleagues (Seaborne et al., 2018). EPIC BeadChips were imaged using the Illumina iScan System (Illumina, United States). DNA methylation analysis, CpG enrichment analysis (GO and KEGG pathways), differentially methylated region (DMR) analysis and Self Organizing Map (SOM) temporal profiling. Following DNA methylation quantification via Methylation EPIC BeadChip array, raw.IDAT files were processed using Partek Genomics Suite V.7 (Partek Inc. Missouri, USA) and annotated using the MethylationEPIC_v-1-0_B4 manifest file. We first checked the average detection p-values for each sample across all probes. The mean detection p-value for all samples across all probes was 0.000295, and the highest for any given sample was 0.000597, which is well below the recommended 0.01 in the Oshlack workflow (Maksimovic et al., 2017). We also produced density plots of the raw intensities/signals of the probes per sample. These demonstrated that all methylated and unmethylated signals were over 11.5 (mean median signal for methylated probes was 11.56 and unmethylated probes 11.69), and the mean difference between the median methylation and median unmethylated signal was 0.13, well below the recommended difference of less than 0.5 (Maksimovic et al., 2017). Upon import of the data into Partek Genomics Suite we removed probes that spanned X and Y chromosomes from the analysis due to having both males and females in the study design, and although the average detection p-value for each sample was very low (no higher than 0.000597) we also excluded any individual probes with a detection p-value that was above 0.01 as recommended (Maksimovic et al., 2017). Out of a total of 865,859 probes removing those on the X & Y chromosome (19,627 probes) and with a detection p-value above 0.01 (4,264 probes) reduced the total probe number to 843,355 (note some X&Y probes also had detection p-values of above 0.01). We also filtered out probes associated with single-nucleotide polymorphisms (SNPs) and any known cross-reactive probes using previously defined SNP and cross-reactive probe lists from EPIC BeadChip 850K validation studies (Pidsley et al., 2016). This resulted in a final list of 791,084 probes to be analysed. Following this, background normalization was performed via functional normalization (with noob background correction) as previously described (Maksimovic et al., 2012). Following functional normalization, we also undertook quality control procedures via Principle Component Analysis (PCA), density plots by lines as well as box and whisker plots of the normalized data for all samples. Any outlier samples were detected using Principle Component Analysis (PCA) and the normal distribution of β-values. Outliers were detected if they fell outside 2 standard deviations (SDs) of the ellipsoids and/or if they demonstrated different distribution patterns to the samples of the same condition. We confirmed that no samples demonstrated large variation [variation defined as any sample above 2 standard deviations (SDs) – depicted by ellipsoids in the PCA plots and/or demonstrating any differential distribution to other samples, depicted in the signal frequency by lines plots. Therefore, no outliers were detected in this sample set. Following normalization and quality control procedures, we undertook differentially methylated position (DMP) analysis by converting β-values to M-values [M-value = log2(β/(1 − ββ)], as M-values show distributions that are more statistically valid for the differential analysis of methylation levels (Du et al, 2010). We undertook a one-way ANOVA for comparisons of baseline, training, detraining and retraining muscle tissue. Any differentially methylated CpG position (DMP) with an unadjusted p-value of ≤ 0.01 was used as the statistical cut off for the discovery of DMPs. We then undertook CpG enrichment analysis on these differentially methylated CpG lists within gene ontology (GO) and KEGG pathways (Kanehisa & Goto, 2000; Kanehisa et al., 2016, 2017) using Partek Genomics Suite and Partek Pathway. Differentially methylated region (DMR) analysis, that identifies where several CpGs are consistently differentially methylated within a short chromosomal location/region, was undertaken using the Bioconductor package DMRcate (DOI: 10.18129/B9.bioc.DMRcate). Finally, to plot and visualize temporal changes in methylation across the timepoints we implemented Self Organizing Map (SOM) profiling of the change in m-value within each condition using Partek Genomics Suite.


title: UV-induced reorganization of 3D genome mediates DNA damage response [RNA-Seq]

status: Public on May 27 2024

city: Istanbul

first: Ogun

last: Adebali

address: Orta Mahalle, Üniversite Caddesi No:27 Tuzla, Sabancı Üniversitesi

department: Mol. Bio. Genetics & Bioeng.

country: Turkey

institute: Sabanci University

summary: While it is well-established that UV radiation threatens genomic integrity, the precise mechanisms by which cells orchestrate DNA damage response and repair within the context of 3D genome architecture remain unclear. Here, we address this gap by investigating the UV-induced reorganization of the 3D genome and its critical role in mediating damage response. Employing temporal maps of contact matrices and transcriptional profiles, we illustrate the immediate and holistic changes in genome architecture post-irradiation, emphasizing the significance of this reconfiguration for effective DNA repair processes. We demonstrate that UV radiation triggers a comprehensive restructuring of the 3D genome structure at all levels, including loops, topologically associating domains and compartments. Through the analysis of DNA damage and excision repair maps, we uncover a correlation between genome folding, gene regulation, damage formation probability, and repair efficacy. We show that adaptive reorganization of the 3D genome is a key mediator of the damage response, providing new insights into the complex interplay of genomic structure and cellular defense mechanisms against UV-induced damage, thereby advancing our understanding of cellular resilience.

overall_design: The goal is to understand the effects of UV-exposure on the 3D genome architecture and transcriptional regulation. Accordingly, HeLa-S3 cells were cultured and exposed to 254-nm UV (1 J/m2/s) for 20 seconds. At the recovery time points of 12, 30 and 60 minutes post-UV, cells were harvested. A non-UV control group has been processed as well. Hi-C, and RNA-Seq techniques have been applied.

first: Veysel

middle: O

last: Kaya


title: AMH protects the ovary from doxorubicin by regulating cell fate and the response to DNA damage

status: Public on May 24 2024

city: Boston

first: Ngoc Minh Phuong

last: Nguyen

state: MA

address: 55 Fruit st

department: Pediatric Surgical Research Laboratory

institute: Massachusetts General Hospital andHarvard Medical School

summary: we aim to explore the single-cell mechanisms behind ovarian damage in prepubertal mice following doxorubicin (DOX) treatment, and how Anti-Müllerian hormone (AMH) prevents chemotherapy-induced pathological damage, using single-cell RNA sequencing as well as quantitative and morphometric histological evaluation.

overall_design: we treated mice (n=5) at the prepubertal age (postnatal day 25) with two weekly doses of DOX (3mg/kg). Additionally, we co-treated another group of mice with recombinant human AMH (rhAMH) (0.75mg/kg), the dose that was proven previously to protect the ovarian reserve to assess the protective effects of AMH. For comparison, we also included a group that received only rhAMH treatment. Ovaries were collected 4 hours after the second dose of DOX.

first: Dadi

last: Gao

first: David

last: Pépin


title: Discrimination of direct HIV-1 Tat effects and compensatory reactions of cultured B cells by RNA-seq analysis

city: Moscow

first: Anna

last: Valyaeva

address: Leninskiye Gory, MSU, 1-73

department: Faculty of Bioengineering and Bioinformatics

country: Russia

institute: Lomonosov Moscow State University

summary: We constructed B cell lines with inducible and stable HIV-1 Tat expression and performed RNA sequencing to study the acute and chronic effects of Tat protein.

overall_design: RNA-seq of lymphoblastoid B cell line RPMI8866 with inducible or stable HIV-1 Tat protein expression. Mock RPMI8866 cells and RPMI(Tat-ind) cells before doxycycline treatment were used as controls. Three biological replicates in each group.

first: Maria

last: Tikhomirova

first: Anastasia

last: Zharikova

first: Alexey

last: Penin

first: Daria

last: Potashnikova

first: Yana

middle: R

last: Musinova

first: Andrey

last: Mironov

first: Yegor

last: Vassetzky

first: Eugene

middle: V

last: Sheval


title: UV-induced reorganization of 3D genome mediates DNA damage response [Hi-C]


title: Integrative Multiomics in the Lung Reveals a Protective Role of Asporin in Pulmonary Arterial Hypertension.

status: Public on May 22 2024

city: Los Angeles

first: Jason

last: Hong

address: 10833 Le Conte Ave, 43-229 CHS

institute: UCLA

summary: Integrative multiomics can elucidate pulmonary arterial hypertension (PAH) pathobiology but procuring human PAH lung samples is rare. Here, we leverage transcriptomic profiling and deep phenotyping of the largest multicenter PAH lung biobank to date (96 disease and 52 control) by integration with clinicopathologic data, genome-wide association studies, Bayesian regulatory networks, single-cell transcriptomics, and pharmacotranscriptomics. We identify two potentially protective gene network modules associated with vascular cells, and we validated ASPN, coding for asporin, as a key hub gene that is upregulated as a compensatory response to counteract PAH. Specifically, we find that asporin is upregulated in lungs and plasma of multiple independent PAH cohorts and correlates with reduced PAH severity. We show asporin inhibits proliferation and TGF-β/pSMAD2/3 signaling in pulmonary artery smooth muscle cells from PAH lungs. Finally, we demonstrate in Sugen-hypoxia rats that ASPN knockdown exacerbated PAH while recombinant asporin attenuated PAH. Our integrative systems biology approach to dissect the PAH lung transcriptome uncovered asporin as a novel protective target with therapeutic potential in PAH.

overall_design: Comparative gene expression profiling analysis of data obtained from RNA-seq of explant lung tissue from patients with pulmonary hypertension undergoing lung transplantation and failed donor lungs.

first: Mark

last: Geraci

first: Robert

last: Stearman

first: Micheala

last: Aldred


title: Transcriptional control of central T cell tolerance by NR4A family transcription factors

status: Public on May 20 2024

city: Atlanta

first: Chris

last: Scharer

state: GA

address: 1510 Clifton Rd, Suite 3086A

department: Microbiology and Immunology

institute: Emory University

summary: The diverse T cell receptor (TCR) repertoire – generated randomly through VDJ recombination early in development – is inherently self-reactive. Clonal deletion and diversion to regulatory T cell (Treg) fate are key central tolerance mechanisms engaged during thymic selection. The NR4A family of transcription factors has been implicated in both, but the extent of its contribution to clonal deletion and the transcriptional mechanisms at play remain unknown. Here, we investigated both NR4A-dependent and NR4A-independent transcriptional changes induced in developing thymocytes in response to model self-antigen presented by medullary thymic epithelial cells (mTECs). To do so, we took advantage of the MHCII-restricted TCR transgene OTII and model self-antigen (membrane-bound ovalbumin expressed under the control of the rat insulin promoter encoded by the RIP-mOVA transgene). Our results suggest that a surprisingly broad transcriptional program is enacted upon high affinity self-antigen encounter in the thymus. Furthermore, by investigating the expression profile of OTII Nr4a1-/- Nr4a3-/- double knock-out (OTII-DKO) thymocytes that receive high affinity antigen-dependent signal but escape both clonal deletion and Treg diversion, we reveal evidence for deletional and non-deletional central tolerance mechanisms.

overall_design: We performed comparative gene expression profiling using bulk RNA sequencing from OTII Nr4a1-/-Nr4a3-/- double knock-out (OTII-DKO) and control OTII Nr4a1+/+ Nr4a3+/+ (OTII-ctl) transgenic thymocytes under conditions of positive and negative thymic selection (i.e. clonal deletion). Two or three biological replicates were analyzed for each selection type.

first: Christopher

first: Julie

last: Zikherman

first: James

last: Mueller

first: Hailyn

last: Nielsen


title: Cellular senescence induced by down-regulation of PTBP1 correlates with exon skipping of mitochondrial related gene NDUFV3

status: Public on May 18 2024

first: haimei

last: wen

address: SongHu road 2005

department: Collaborative Innovation Center of Genetics and Development,

institute: Fudan university

summary: As the most prevalent type of alternative splicing in animal cells, exon skipping plays an important role in expanding the diversity of transcriptome and proteome, thereby participating the regulation of diverse physiological and pathological processes such as development, aging and cancer. Cellular senescence serves as an anti-cancer mechanism could also contribute to individual aging. Although the dynamic changes of exon skipping during cellular senescence were revealed, its biological consequence and upstream regulator remain poorly understood. Here, by using human foreskin fibroblasts (HFF) replicative senescence as a model, we discovered that splicing factor PTBP1 was an important contributor for global exon skipping events during senescence. Down-regulated expression of PTBP1 induced senescence-associated phenotypes and related mitochondrial functional changes. Mechanistically, PTBP1 binds to the third exon of mitochondrial complex I subunit coding gene NDUFV3 and protects the exon from skipping. We further confirmed that exon skipping of NDUFV3 correlates with and partially contributes to cellular senescence and related mitochondrial functional changes upon PTBP1 knockdown. Together, we revealed for the first time that mitochondrial related gene NDUFV3 is a new downstream target for PTBP1-regulated exon skipping to mediate cellular senescence and mitochondrial functional changes.

overall_design: Total RNA were isolated with TRIzol reagent according to manufacturer's instructions (Invitrogen, 15596026). Then, RNA was reversely transcribed into cDNA with oligo-(dT)23 VN primer and (or) random hexamers using HiScript II 1st Strand cDNA Synthesis Kit (+gDNA wiper) (Vazyme, R212). Gene expression at the RNA level was quantified by qRT-PCR using 2× SYBR mix (Vazyme, Q711). GAPDH’s RNA expression level served as an internal control.

first: Yu

last: Yang

first: Haimei

last: Wen


title: Identification of relapse mechanisms for pediatric T-ALL (T-cell acute lymphoblastic leukemia) through scRNAseq analysis

status: Public on May 17 2024

city: Valbonne - Sophia-Antipolis

first: Kevin

last: Lebrigand

address: 660 route des lucioles

institute: IPMC/CNRS

summary: If relapses occur in only 20% of T-ALL children, 70% will have a dismal outcome, justifying the need for new therapeutic options. Transcriptomic plasticity is a property of leukemic cells to adapt their functions and resist to treatments. The project aims at analyzing, at a single cell resolution, the variations of the gene expression, between diagnostic and relapse. This will allow identification of relapse-associated genes as potential new targets for future innovative treatments.

overall_design:
For 3 patients, we obtained paired samples at diagnostic and relapse that will allow us to study the evolution of the leukemia during acquisition of a resistance to treatments. Comparative gene expression profiling of scRNAseq data from paired diagnostic-relapse samples.
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Authors state "this is human sample so the raw data are currently being processed at EGA".

first: Jean-Francois

last: Peyron


title: ATAC-seq of frozen H1 Human Embryonic Stem Cells (hESC)

status: Public on May 12 2024

city: Stanford

first: William

middle: James

last: Greenleaf

address: Beckman Center B-279, Campus Drive West

department: Genetics

institute: Stanford University

summary: We performed Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) to profile genome-wide chromatin accessibility in the human H1 embryonic stem cell (ESC) line. We used this data to train a deep learning model called ChromBPNet which can accurately predict base-resolution accessibility profiles as a function of DNA sequence, while accounting for and correcting biases due the sequence preferences of the Tn5 transposase used in ATAC-seq. We interpreted the models to identify globally predictive transcription factor (TF) motifs, individual predictive motif instances in all accessible regions and Tn5-bias corrected canonical footprints of TFs at these predictive motifs.

overall_design:
Assay for Transposase-Accessible Chromatin using sequencing (ATAC-seq) was used to profile genome-wide chromatin accessibility in the human H1 embryonic stem cell (ESC) line.
We then used this data to develop ChromBPNet, a deep learning model capable of predicting DNA sequence-based base-resolution accessibility profiles, correcting biases from Tn5 transposase in ATAC-seq, and revealing globally predictive transcription factor motifs and their specific instances, along with Tn5-bias-corrected canonical footprints for TFs at these motifs.

first: Arwa

last: Kathiria

first: Viviana

last: Risca

first: Will

first: Anusri

last: Pampari

first: Anna

last: Shcherbina

first: Anshul

last: Kundaje


title: The effect of metal cations on the capture of Pol II-derived nascent transcripts in NET-seq experiments

status: Public on May 13 2024

city: Birmingham

first: Abigail

last: Huffines

address: 720 20th St S

institute: University of Alabama at Birmingham

summary: The goal of this study was to test the effect of metal cations (CaCl2 and MnCl2) during Pol II NET-seq. These metals are readily used in the majority of studies that use NET-seq to analyze Pol II occupancy, but their effect on nascent transcript capture has not been analyzed. Our results suggest that the inclusion of these metals in Pol II NET-seq experiments does not cause a significant change in Pol II occupancy in the untreated control vs. metal-treated (CaCl2 or MnCl2) samples.

overall_design: Pol II NET-seq was performed in triplicate for each of three treatment conditions: untreated, CaCl2-treated, and MnCl2-treated.

first: David

last: Schneider


title: Gene Regulatory Roles of Growth and Differentiation Factors in Retinal Development

status: Public on May 16 2024

city: Palo Alto

first: Ziming

last: Luo

address: 1651 Pagemill Rd.

department: Ophthalmology

summary: RGC differentiation is tightly controlled by extrinsic and intrinsic factors. Growth and differentiation factor 15 (GDF-15) promotes RGC differentiation, opposite to GDF-11 which inhibits RGC differentiation, both in the mouse retina and in human stem cells. To deepen our understanding of how these two closely related molecules confer opposing effects on retinal development, here we assess the transcriptional profiles of mouse retinal progenitors exposed to exogenous GDF-11 or -15. We find a dichotomous effect of GDF-15 on RGC differentiation, decreasing RGCs expressing residual pro-proliferative genes and increasing RGCs expressing non-proliferative genes, suggestive of greater RGC maturation. Furthermore, GDF-11 promoted the differentiation of photoreceptors and amacrine cells. These data enhance our understanding of the mechanisms underlying the differentiation of RGCs and photoreceptors from retinal progenitors and suggest new approaches to the optimization of protocols for the differentiation of these cell types.

overall_design: E14 retinal tissues were dissociated and cultured in vitro. Three conditions were included, in which cells were treated with PBS, GDF11 or GDF15  for 5 days for scRNA Sequencing.

first: Sahil

last: Shah

first: Bogdan

last: Tanasa

first: Kun-Che

last: Chang

first: Jeffrey

last: Goldberg


title: Transcriptional characterization of adjuvantly radio(chemo)therapy treated head and neck squamous cell carcinoma (HNSCC).

city: Munich

first: Kristian

last: Unger

address: Marchioninistr. 15

department: Department of radiation oncology

institute: LMU University Hospital

summary: We characterized n=134 HNSCC by Quantseq 3´-RNA-sequencing for their transcriptional profiles in order to analyze differential expression of clinically relevant subgroups, define transcriptionally defined molecular subgroups and to characterize the tumor microenvironment by gene expression devonvolution approaches.

overall_design: n=134 HNSCC with known HPV-status were characterized. The metabolic pathway subtypes were defined.

first: Julia

last: Heß

first: Martin

last: Selmansberger

first: Benedek

last: Danko

first: Horst

last: Zitzelsberger

first: Claus

last: Belka


title: ZNF143 binds DNA and stimulates transcription initiation to activate and repress direct target genes (ATAC-seq)

status: Public on May 07 2024

city: Farmington

first: Jinhong

last: Dong

state: CT

address: 400 Farmington Ave

department: Center for Cell Analysis and Modeling

institute: University of Connecticut

summary: Transcription factors bind to sequence motifs and act as activators or repressors. Transcription factors interface with a constellation of accessory cofactors to regulate distinct mechanistic steps to regulate transcription. We rapidly degraded the essential and ubiquitously expressed transcription factor ZNF143 to determine its function in the transcription cycle. ZNF143 facilitates RNA Polymerase initiation and activates gene expression. ZNF143 binds the promoter of nearly all its activated target genes. Paradoxically, ZNF143 binds near the site of genic transcription initiation to directly repress a subset of genes. Although ZNF143 stimulates initiation at ZNF143-repressed genes (i.e. those that increase expression upon ZNF143 depletion), the molecular context of binding leads to cis repression. ZNF143 competes with other more efficient activators for promoter access, physically occludes transcription initiation sites, and acts as a molecular roadblock to RNA Polymerase to repress genes in cis. The term "context specific" is often invoked to describe transcription factors that have both activation and repression functions. Here, we define the context and molecular mechanisms of ZNF143-mediated cis activation and repression.

overall_design: HEK293T cells with ZNF143 tagged with an inducible degron tag were treated with drugs to induce ZNF143 degradation for 30 minutes, then collected for ATAC-seq.

first: Michael

last: Guertin


title: Bulk RNA-sequencing results of p16INK4A-high and -low glioblastoma, IDH-wild type, grade 4.

status: Public on May 10 2024

city: Suwon

first: Soon Sang

last: Park

address: Worldcup-Ro 206

country: South Korea

institute: Ajou University School of Medicine

summary: The expression of p16INK4A is widely used to identify fully senescent cells in noncancerous tissues, as well as senescent cancer cells in various types of cancer. In our study, we observed diffuse expression of p16INK4A in around 40 % of the patients with wild-type CDKN2A gene without mutation showing senescence phenotype, correlating with significant differences in overall survival between patients with high and low levels of p16INK4A expression. This prognostic distinction led us to hypothesize that cellular senescence in glioblastoma cells might confer a favorable effect on patient outcomes. To investigate the mechanism underlying this phenotype, we conducted RNA sequencing on 11 samples of glioblastoma, IDH-wild type along with corresponding in vivo and in vitro studies.

overall_design: To investigate the characteristics of p16INK4A-high and -low cancers, bulk RNA-sequencing of surgically removed glioblastoma tissues was performed in Ajou University Hospital, Korea.

first: Soon

middle: S

first: Jang-Hee

last: Kim

first: Tae

middle: H

last: Roh

first: Se-Hyuk


title: Effect of YAP-S127A overexpression on gene expression profile

status: Public on May 04 2024

city: shanghai

first: Xialu

last: Hong

state: State...

address: 415 Fengyang Road

institute: Changzheng hospital

summary: To investigate the mechanism by which YAP regulates methionine metabolism, seven-week-old C57 mice were injected with AAV8-TBG-control or AAV8-TBG-YAP-S127A through tail vein. 8 weeks later, the primary hepatocytes from two groups were isolated for RNA sequence analysis.

overall_design: RNA sequence was performed in control primary hepatocytes and primary hepatocytes overexpressing human YAP-S127A.

first: Chenkai

last: Huang

first: Hui

last: Qian


title: RNA-Seq of B-Cell Lines Clones, parental and resistant to Idelalisib or IMGN529

status: Public on May 06 2024

city: Bellinzona

first: Luciano

last: Cascione

state: Ticino

address: Via Chiesa 5

department: Lymphoma & Genomics Research Program

country: Switzerland

institute: IOR

summary:
Idelalisib was the first-in-class PI3Kδ inhibitor and additional compounds are undergoing clinical investigation. To identify modalities to overcome resistance to these agents, we have developed  idelalisib-resistant model derived from marginal zone lymphoma cell line (VL51). Cells were kept under idelalisib until acquisition of resistance (VL51-RER)  or with no drug (parental). In this experiment we identified the modulated genes between the resistant cell line and the parental counterpart (sensitive to the drug)
We invastigated the transcriptomic profiles of parental K1718 B-cell lymphoma cell lines and the same cell line made resistant to Idelalisib. in addition two B-Cells (SUDHL2 and SUDHL4) are made resistant to IMGN529 (Naratuximab emtansine) an antibody-drug conjugate (ADC) incorporating an anti-CD37 monoclonal antibody conjugated to the maytansinoid DM1 as payload.

overall_design: Cell lines were deep sequenced on a Illumina NextSeq using High Output, PhiX was spiked into the library at 5%.

first: Francesco

last: Bertoni

first: Alberto

last: Arribas


title: Transcriptomic Landscape of Hras12V Oncogene-Induced Hepatocarcinogenesis with Gender Disparity

city: DaLian

first: aiguo

last: Wang

address: DaLian

institute: Dalian Medical University

summary: The genesis of hepatocellular carcinoma (HCC) is closely related to hyper-activated Ras signals and male factors. This study aimed to characterize the underlying mechanisms by examining the transcriptome of Hras12V transgenic mice (Ras-Tg, HCC model). A transcriptomic database was established via RNA-Seq of HCC (T) and the adjacent precancerous liver tissue (P) of Ras-Tg and the normal liver tissue of wild-type mice (W) of both sexes. Bioinformatics analysis as well as metabolite and gene expression assays were performed. Totally, 2708 differentially expressed genes (DEGs) were screened out (FC ≥ 2 and q < 0.05). Longitudinal comparative analysis within W, P, and T and correlation expression pattern analysis revealed common/unique cluster-enriched items towards HCC between the sexes. Specifically, the numbers of DEGs were much higher in females than in males, and tumor suppressor genes, such as p21Waf1/Cip1 and C6, were significantly higher in the female P. This finding denotes the higher sensitivity of female hepatocytes to the Ras oncogene and, therefore, the difficulty in developing HCC. Moreover, convergence in HCC between the sexes suggests the underlying mechanisms for ineffectiveness of sex hormone therapies. Additionally, expression pattern analysis revealed that the DEGs and their relevant pathways were either positively or negatively associated with the HCC/Ras oncogene. Among them, the vital role of glutathione metabolism in HCC was established. The systemic transcriptomic database of the Ras oncogene-induced HCC with sex disparities provides valuable clues for elucidating the underlying mechanisms, selecting the diagnostic biomarker, and planning the clinical therapy in HCC.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for tumor tissue and its adjacent precancerous liver tissues of Hras12V transgenic mice, and normal liver tissues of wild-type C57BL/6 mice from males and females.

first: Aiguo


title: ZNF143 binds DNA and stimulates transcription initiation to activate and repress direct target genes (ChIP-seq)

overall_design:
HEK293T cells with ZNF143 tagged with an inducible degron tag were treated with drugs to induce ZNF143 degradation for 30 minutes, then collected for ChIP-seq for ZNF143 (via HA tag) and SP1.
HEK293T cells with ZNF143 tagged with an inducible degron tag were treated with drugs to induce ZNF143 degradation for 30 minutes, then collected for ChIP-seq targeting Nrf1 or YY1.


title: RNA-seq analyses of periodontal tissue and liver of periodontitis-affected mice

status: Public on May 03 2024

city: Okayama

first: Shigeki

last: Suzuki

state: Okayama

address: 2-5-1 Shikata-cho, Kita-ku

department: Department of Operative Dentistry

institute: Okayama University

summary:
Background/purpose
Periodontal diseases exacerbate hepatic inflammation and diseases like non-alcoholic fatty liver disease via circulating pathogenic factors from periodontal tissue. Long-term pre-symptomatic state eventually leads to the development of such hepatic diseases. However, it is uncertain if periodontitis contributes in the onset of hepatic pre-symptomatic state. Herein, we conducted a hepatic whole transcription analysis of periodontitis-affected mice and healthy mice to understand the early functional changes in the hepatic system in periodontitis-affected mice.
Materials and methods
Silk ligatures were tied around mice second maxillary molars for 14 days to develop periodontitis. RNA-seq samples were collected from periodontal tissues and liver tissues of mice with periodontitis and healthy mice. Lipidomic analysis of hepatic omega-3 fatty acids in periodontitis-affected and healthy mice was conducted. The anti-inflammatory effects of omega-3 fatty acids and their metabolites were elucidated using hepatocytes HepG2 cells.
Results
In the liver of mice with periodontitis, genes coding for cytochrome P450 such as Cyp4a12a and Cyp4a12b were identified as significantly down-regulated genes. Lipidomic analyses identified that epoxidation and subsequent hydrolysis of hepatic omega-3 fatty acids were inhibited in periodontitis-affected mice. Eicosapentaenoic acid metabolites, epoxy eicosatetraenoic acid and dihydroxyeicosatetraenoic acid, inhibited inflammatory responses of HepG2 cells.
Conclusion
These results suggest that, in the liver of periodontitis-affected mice, due to the reduced activity of omega-3 fatty acid epoxidation, pre-symptomatic state  with pro-inflammatory  status develop. Therefore, early intervention of periodontitis might contribute to the prevention of the onset of hepatic diseases.

overall_design: Gene expression analysis in periodontal tissue and liver tissue of periodontitis-affected and healthy mice


title: Aberrant activation of wound healing programs within the metastatic niche facilitates lung colonization by osteosarcoma cells

status: Public on May 09 2024

city: Columbus

first: Matt

last: Cannon

state: OH

address: 700 Children's Dr.

department: Center for Childhood Cancer

institute: Nationwide Children's Hospital

summary: Purpose: Lung metastasis is responsible for nearly all deaths caused by osteosarcoma, the most common pediatric bone tumor. How malignant bone cells coerce the lung microenvironment to support metastatic growth is unclear. This study delineates how osteosarcoma cells educate the lung microenvironment during metastatic progression.   Experimental design: Using single-cell transcriptomics (scRNA-seq), we characterized genome- and tissue-wide molecular changes induced within lung tissues by disseminated osteosarcoma cells in both immunocompetent murine models of metastasis and patient samples. We confirmed transcriptomic findings at the protein level and determined spatial relationships with multi-parameter immunofluorescence. We evaluated the ability of nintedanib to  impair metastatic colonization and prevent osteosarcoma-induced education of the lung microenvironment in both immunocompetent murine osteosarcoma and immunodeficient human xenograft models. Results: Osteosarcoma cells induced acute alveolar epithelial injury upon lung dissemination. scRNA-seq demonstrated that the surrounding lung stroma adopts a chronic, non-resolving wound-healing phenotype similar to that seen in other models of lung injury. Accordingly, metastasis-associated lung demonstrated marked fibrosis, likely due to the accumulation of pathogenic, pro-fibrotic, partially-differentiated epithelial intermediates.  Inhibitionintermediates. Inhibition of fibrotic pathways with nintedanib prevented metastatic progression in multiple murine and human xenograft models. Conclusions: Our work demonstrates that osteosarcoma cells co-opt fibrosis to promote metastatic outgrowth. When harmonized with data from adult epithelial cancers, our results support a generalized model wherein aberrant mesenchymal-epithelial interactions are critical for promoting lung metastasis. Adult epithelial carcinomas induce fibrotic pathways in normal lung fibroblasts, whereas osteosarcoma, a pediatric mesenchymal tumor, exhibits fibrotic reprogramming in response to the aberrant wound-healing behaviors of an otherwise normal lung epithelium, which are induced by tumor cell interactions.

overall_design: We utilized multiple immunocompetent murine osteosarcoma lung metastasis models coupled with single-cell RNA transcriptomics to characterize the cellular and molecular changes induced to the lung microenvironment by disseminated osteosarcoma cells. Transcriptomic findings were confirmed at the protein level with immunohistochemistry in mouse models as well as human patient samples. We tested the effect of nintedanib, an orally available tyrosine kinase inhibitor, for its ability to impair metastasis and prevent osteosarcoma education of the lung microenvironment in both immunocompetent murine osteosarcoma models and immunodeficient human xenograft models.

first: James

last: Reinecke

first: Amy

last: Gross

first: Maren

last: Cam

first: Leyre

last: Jimenez Garcia

first: Matthew

first: Ruben

last: Dries

first: Berkley

middle: E

last: Gryder

first: Ryan

last: Roberts


title: RNA-seq data of 293T cells trasfected with different viral PA-X proteins of influenza viruses

status: Public on May 02 2024

city: Yangzhou

first: Jiao

last: Hu

address: 48 Wenhui Road

institute: Yangzhou University

summary:
PA-X is a small accessory protein that modulates the virulence of various influenza A virus both in mammals and birds.
However, the specific role of PA-X in the pathogenesis of highly pathogenic avian influenza virus (HPAIV) H7N9 subtype in mammals and avian species is largely unknown.
By functional analysis, we want to investigate critical amino acids that contribute to the host shutoff ability of the PA-X protein of the H7N9 virus in 293T cells

overall_design:
Two time points were set. Each time point, a total of three biological samples were analyzed, each of them had triplication.The  samples are named PAX-24h, PAX-48h, PAX-N3-24h,  PAX-N3-48h,  PAX-N6-24h,  PAX-N6-48h, PCAGGS, repectively.

PA-X = the parental PA-X protein of the H7N9 influenza virus
PAX-N3 = the mutant isoform of PA-X carrying I94V mutation in the parental PA-X
PAX-N6 = the mutant isoform of PA-X carrying A100V in the parental PA-X

first: Xia

last: Chen


title: RNA-seq of mouse peritoneal macrophages infected with SeV

status: Public on May 01 2024

city: Shanghai

first: Qihang

last: Zhao

address: 800 xiangyin road

institute: National laboratory of medical immunology

summary: RNA seq of wild-type C57B6/L mouse peritoneal macrophages with infection of SeV (MOI=1) for 12 hours.

overall_design: To analyze variation of gene expression by comparing group of NC with group of SeV.

first: Pin

first: Zhengdong

last: Yang


title: Endothelial IGFBP6 Suppresses Vascular Inflammation and Atherosclerosis

city: Hefei

first: Meiming

last: Su

address: Huangshan Road

institute: USTC

summary: It has previously been reported that expression levels of insulin-like growth factor binding protein 6 (IGFBP6) are reduced in unstable plaques compared to stable plaques in patients with coronary heart disease (CAD). However, the exact role and mechanism of IGFBP6 in endothelial homeostasis and inhibition of focal atherosclerotic plaque development in vivo remains unclear. Transcripome sequencing and bioinformatics analysis of IGFBP6-overexpressed human umbilical vein endothelial cells (HUVECs) suggest that pathway analysis and differentially expressed mRNA may help clarify the role and potential mechanism of IGFBP6 in the treatment of cardiovascular diseases.

overall_design: Human umbilical vein endothelial cells (HUVECs) were seeded in 0.2% gelatin-coated petri dishes the day before the experiment.   The next day, cells were treated with AdNC or Ad-IGFBP6 for 24 hours and with or without TNF-α (10ng/ml) for 6 hours.   After treatment, total RNA was isolated using the RNeasy kit.   RNA libraries were prepared for sequencing using the rRNA removal library construction protocol.

first: Wenqi

first: Suowen

last: Xu

first: Jianping

last: Weng


title: Histone H3 E50K mutation confers oncogenic activity and supports an EMT phenotype

first: Jennifer

middle: Marie

last: Spangle

state: Georgia

address: 1365C Clifton Rd NE

summary: Sequencing of human patient tumors has identified recurrent missense mutations in genes encoding core histones. We report that mutations that convert histone H3 amino acid 50 from a glutamate to a lysine (H3E50K) supports an oncogenic phenotype in human cells. Expression of H3E50K is sufficient to transform human cells as evidenced by a dramatic increase in cell migration and invasion, and a statistically significant increase in proliferation and clonogenicity. H3E50K also increases the invasive phenotype in the context of co-occurring BRAF mutations, which are present in patient tumors characterized by H3E50K. E50 lies on the globular domain surface in a region that contacts H4 within the nucleosome. We find that H3E50K perturbs global proximal H3 posttranslational modifications and dysregulates gene expression, activating the epithelial to mesenchymal transition. Functional studies using S. cerevisiae reveal that, while yeast cells that express H3E50K as the sole copy of histone H3 show sensitivity to cellular stressors including caffeine, H3E50K cells display growth properties that are distinct from the characterized H3K36M oncohistone yeast model. Taken together, these data suggest that additional histone H3 mutations have the potential to be oncogenic drivers and function through distinct mechanisms that dysregulate gene expression

overall_design: To investigate whether the histone H3.3 E50K mutant differentially regulates the global transcriptome compared to H3.3 WT. We are using immortalized but untransformed HMEC DD cells with dominant negative p53 cells that stably overexpressed a C-terminal TY1 epitope-tagged H3.3 WT and E50K.

middle: M

first: Anita

last: Corbett

first: Kirti

last: Sad

first: Celina

last: Jones

first: Miranda

last: Adams

first: Severin

last: Lustenberger

first: Richard

last: Lee

first: Satvik

last: Elayavalli

first: Jonathan

last: Farhi

first: Laramie

last: Lemon

first: Milo

middle: B

last: Fasken


title: Genomic and phenotypic stability of fusion-driven pediatric sarcoma cell lines [array]

city: Heidelberg

first: Thomas

middle: Georg Philipp

last: Grunewald

address: Im Neuenheimer Feld 280

department: Division of Translational Pediatric Sarcoma Research

institute: German Cancer Research Center (DKFZ)

summary: Human cancer cell lines are the mainstay of cancer research. Recent reports showed that highly mutated adult carcinoma cell lines (mainly HeLa and MCF-7) present striking diversity across laboratories and that long-term continuous culturing results in genomic/transcriptomic heterogeneity with strong phenotypical implications. However, to what extent these limitations in terms of reproducibility may be generalized to all cancer cell lines remained unexplored. Here, we hypothesized that oligomutated pediatric sarcoma cell lines mainly driven by a fusion transcription factor, such as Ewing sarcoma (EwS), were genetically and phenotypically more stable than the previously investigated (adult) carcinoma cell lines. A comprehensive molecular and phenotypic characterization of multiple EwS cell line strains, together with a simultaneous analysis during 12 months of continuous cell culture showed that fusion-driven pediatric sarcoma cell line strains are genomically more stable than adult carcinoma strains, display remarkably stable and homogenous transcriptomes, and exhibit uniform and stable drug response. Additionally, the analysis of multiple EwS cell lines subjected to long-term continuous culture revealed that variable degrees of genomic/transcriptomic/phenotypic changes among fusion-driven cell lines, further exemplifying that the potential for reproducibility of in vitro scientific results may be rather understood as a spectrum, even within the same tumor entity.

overall_design: Human cell lines were cultured continously over a period of 12 months and subjected to DNA methylation profiling using Illumina EPIC arrays at three different time points (m0=ITP, m6=MTP, m12=FTP)

first: Merve

last: Kasan

first: Florencia

last: Cidre-Aranaz

middle: G

last: Grünewald


title: Gene expression data of cancer cells overexpressing p53-R282W with or without  decitabine (DAC) treatment

status: Public on Apr 30 2024

first: Yigang

last: Tang

address: No 197 RUIJIN ER RD

institute: Shanghai Jiaotong University

summary: The goals of this study are to compare transcriptome of tumor cells with p53-R282W treated by DAC

overall_design: Treat isogenic p53 null tumor cells (HCC1937 and THP-1) overexpressing p53-R282W with  DAC

first: Haoyu

last: Wang

first: Min

last: Lu


title: ZNF143 binds DNA and stimulates transcription initiation to activate and repress direct target genes (PRO-seq)

first: Jinhong

last: Dong

overall_design: HEK293T cells with ZNF143 tagged with an inducible degron tag were treated with drugs to induce ZNF143 degradation for 30 minutes, then collected for PRO-seq.

first: Michael

middle: J

last: Guertin


title: Netrin1 patterns the dorsal spinal cord through modulation of Bmp signaling

status: Public on Apr 23 2024

city: Los Angeles

first: Riki

last: Kawaguchi

state: CA

address: 760 Westwood Plaza, Room 37-420

department: Department of Psychiatry and Neurology Department of Psychiatry

institute: University of California, Los Angeles

summary: We directed mESCs towards the dorsal sensory interneurons (dIs) using RA-/+ BMP4 and tested the effects of an axon guidance molecule netrin1 on dI differentiation in vitro. Using bulk-RNA seq analysis, we investiagted when the addition of netrin 1 will affect the transcriptome of differentiating dIs. For this analysis, we used three timewindows (conditions 1-3) for netrin addition (1) condition 1  netrin 1 was added together with RA+BMP4 at day 4, and RNA seq was conducted at day 5, (2) netrin1 was added after the RA+BMP4 mediated patterning at day 5 and RNA seq was conducted at day 6, and (3) netrin1 was added for the extended duration between day 5-day9 to investigate the netrin1 effects throughout the dI differentiation and RNA seq was conducted at day 9.  Through this analysis we identified that netrin1 downregulate BMP signaling pathway, possibly through modlulating the mRNA processing of key mediator of BMP signaling such as Id genes and BMP signaling inhibitor Smad7 and affect dI differentiation. This is the first report to show that netrin1 regulate dI fate specification in the dorsal spinal cord.

overall_design: 3- replicate samples were collected at day 5 (condition 1), and day 6 (condition 2), while 2 replicates passed the quality control for day 9 RNA seq (condition 3), Total 16 samples were collected from 3-independent differentiations.

first: Sandy

last: Alvarez

first: Sandeep

last: Gupta

first: Kaitlyn

last: Honeychurch

first: Yesica

last: Mercado-Ayon

first: Samantha

last: Butle


title: Effect of HNF4α overexpression on gene expression profile

status: Public on Apr 26 2024

first: Bonan

last: Xu

address: 415，Fengyang Road

department: gastroenterology

institute: Shanghai Changzheng Hospital

summary: In order to explore the potential mechanism of HNF4α in ICC, RNA-seq was performed to obtain differentially expressed genes (DEGs) between HuCCT-1 cells infected with AdGFP or AdHNF4α


title: Large-scale transcript variants dictate neoepitopes for cancer immunotherapy

status: Public on Apr 21 2024

city: nan jing

first: shiliang

last: ji

address: No. 163，Xianlin Avenue

department: State Key Laboratory of Pharmaceutical Biotechnology, School of Life Sciences

institute: Nanjing University

summary: Neoantigen tumor vaccines have shown promising anti-tumor efficacy in early clinical trials. However, the underlying mechanism regarding response or resistance to this treatment remains unclear. Therefore, further studies using advanced technologies such as single-cell RNA sequencing (scRNA-seq) were performed to reveal significant changes in tumor microenvironment after vaccination.

overall_design: Tumor-infiltrating immune cells of MC38 tumors were isolated by Fluorescence activated cell sorting (FACS) according to the presence or absence of CD45+ cells, and analyzed using scRNA-seq/TCR-seq.

first: Shiliang

last: Ji

first: Zengding

last: Wu

first: Zhen

last: Huang

first: Junfeng

last: Zhang


title: Liver cancer development driven by the AP-1/c-Jun~Fra2 dimer through c-Myc

status: Public on Apr 22 2024

city: Vienna

first: Latifa

last: Bakiri

address: Lazarettgasse 14, BT25/2, 6th floor Lab 2

department: Laboratory Medicine

country: Austria

institute: Medical University of Vienna

summary: Hepatocellular carcinoma (HCC) is a leading cause of cancer-related death. HCC incidence is on the rise, while treatment options remain limited. Thus, a better understanding of the molecular pathways involved in HCC development has become a priority to guide future therapies. While previous studies implicated the AP-1 (Fos/Jun) transcription factor family members c-Fos and c-Jun in HCC formation, the contribution of Fos-related antigens 1 and 2 (Fra-1/2) is unknown. Here we show that hepatocyte-restricted expression of a single chain c-Jun~Fra-2 protein, which functionally mimics the c-Jun/Fra-2 AP-1 dimer, results in spontaneous HCC formation in c-Jun~Fra-2hep mice. Several hallmarks of human HCC, such as cell cycle dysregulation and the expression of HCC markers are observed in liver tumors arising in c-Jun~Fra-2hep mice. Tumorigenesis occurs in the context of mild inflammation, low-grade fibrosis and Pparg-driven dyslipidemia. Subsequent analyses revealed increased expression of c-Myc, evidently directly regulated by AP-1 through a conserved distal 3’ enhancer. Importantly, c-Jun~Fra-2-induced tumors revert upon switching off transgene expression, suggesting oncogene addiction to the c-Jun~Fra-2 transgene. Tumors escaping reversion maintained c-Myc and c-Myc target gene expression, likely due to increased c-Fos. Interfering with c-Myc in established tumors using the BET bromodomain inhibitor JQ-1 diminished liver tumor growth in c-Jun~Fra-2 mutant mice. Thus, our data establish c-Jun~Fra-2hep mice as a novel model to study liver tumorigenesis and identify the c-Jun/Fra-2-Myc interaction as a potential target to improve HCC patient stratification and/or therapy.

overall_design:
RNA from murine (adult males, C57BL/6) liver samples was sequenced. Two sets were included in the experiment in twosequencing runs: the first set included 9 samples from mice sacrificed 2 months after transgene induction in 6 controls and 3 mutants. The second set included 10 samples from mice sacrificed 9 months after transgene induction in 3 controls and 3 mutants. in the 3 mutants, 3 samples corresponded to areas that were macroscopically tumor free (periT) and 4 samples corresponded to macroscopically visible tumors: two tumors from mutant -1 and one tumor from each of mutant -2 and mutant-3. Comparative expression profiling was conducted across genotype (mutants compared to controls) and time. Transgene expression in the mutants was always started at weaning (3 weeks of age) by doxycycline removal. Controls were littermates that received the same treatment.

Grant ID: 741888 (ERC AdG 2016)
Grant title: CSI-Fun
Funding source: European Research Council
Grantee name: Erwin F.Wagner

Grant ID: 859860 (ITN 2019)
Grant title: CANCERPREV
Funding source: H2020 - MSCA
Grantee name: Erwin F. Wagner


title: Pooled screening for CAR function in glioblastoma

status: Public on Apr 19 2024

city: Cambridge

first: Caleb

last: Perez

email: crperez@mit.edu, mbirnb@mit.edu

address: 77 Massachusetts Ave

department: Biological Engineering

institute: MIT

summary: Human primary CD3+ T cells expressing IL13Ra2-targeted chimeric antigen receptors (CARs) that had been identified through high throughput pooled screening were rechallenged with an IL13Ra2+ human glioblastoma cell line , then stained with CITESEQ antibodies indicative of T cell memory and exhaustion state and prepared for single cell sequencing in order to determine their phenotypic responses to chronic tumor challenge relative to that of a clinical standard CAR.

overall_design: CD3+ T cells from two donors expressing two novel CARs and one control CAR were subject to antigen stimulation with mitomycin treated IL13Ra2+ U87 cells four times in vitro. These were then stained with TotalSeq-C hashing, sorted to remove residual tumor cells, and pooled before encapsulation in two channels of the Chromium Single Cell 3′ v3.1 platform (10X Genomics). Gene expression (GEX) libraries were constructed based on manufacturer’ instructions, while hashing antibody libraries were constructed as reported previously. The resulting libraries were pooled at 12.5% antibody to 87.5% GEX before sequencing on an Illumina NextSeq500 to a depth of 34,380 reads per cell. Reads were aligned to the Genome Reference Consortium Human Build 38 (GRCh38), and a cell–gene matrix was generated using the CellRanger pipeline (10X Genomics; v4.0.0). Downstream analysis was performed using the Seurat package (v4.0.0).

first: Khloe

last: Gordon

first: Andrea

last: Garmilla

first: Maxine

last: Lam

first: Joey

last: Aw

first: Douglas

middle: A

last: Lauffenburger

last: Pavesi

last: Birnbaum