title: Regulation of meiotic progression by Sertoli-cell androgen signaling

city: Ann Arbor

first: Qianyi

last: Ma

state: MI

address: 1141 E Catherine St

department: Human Genetics

country: USA

institute: University of Michigan

summary: We combine synchronization of spermatogenesis, cytological analyses and single-cell RNA-seq (scRNA-seq) in the Sertoli cell androgen receptor knockout (SCARKO) mutant and control mice, and demonstrate that SCARKO mutant spermatocytes exhibited normal expression and localization of key protein markers of meiotic prophase events, indicating that initiation of meiotic prophase is not androgen dependent, whereas scRNA-seq analysis revealed a molecular transcriptomic block in an early meiotic prophase state (leptotene/zygotene) in mutant germ cells, and identified several misregulated genes in SCARKO Sertoli cells, many of which have been previously implicated in male infertility.

overall_design: Single-cell RNA-seq (scRNA-seq) analysis on whole testes in the Sertoli cell androgen receptor knockout (SCARKO) mutant and control mice at eight, thirteen and sixteen days past spermatogenesis synchronization and twenty-one days post-partum unsynchronized whole testes.

first: Saher

last: Hammoud

first: Mary

last: Handel


title: Dissecting the effect of a 3D micro-scaffold on the transcriptome of Neural Stem Cells with computational genomics

status: Public on Oct 22 2020

city: Pavia

first: Stella

last: Gagliardi

state: Pavia

address: Via Mondino, 2

department: GPG center

country: Italy

institute: IRCCS Mondino Foundation

type: Expression profiling by high throughput sequencing

summary: 3D cell cultures are becoming more and more important in the field of regenerative medicine due to their ability to mimic the cellular physiological microenvironment. Among the different types of 3D scaffolds there is the Nichoid: an engineered 3D micro-scaffold with a structure inspired by the natural staminal niche. The Nichoid can activate cellular response simply by subjecting the cells to mechanical stimuli. This kind of influence results in different cellular morphology and organization, but the molecular bases of these changes remain largely unknown. Here, we show that the main cellular response to the Nichoid is an alteration of the mechanotransduction, which in turn affects the global cellular activity. Through RNA-seq approach on murine Neural Precursors Stem Cells (NPCs) expanded inside the Nichoid, we investigated the deregulated genes and pathways showing that the Nichoid causes alteration in genes strongly connected to mechanical properties. We found 1934 deregulated RNAs, 81% of which were coding genes. Moreover, we fully dissected this mechanism highlighting how the changes start at a membrane level, with subsequent alterations in the cytoskeleton, signalling pathways, and metabolism, all leading to a final alteration in gene expression. The results shown here demonstrate that the 3D scaffold Nichoid influences the biological and genetic response of stem cells thorough specific alterations of cellular signaling.

overall_design: 3 Neural Precursor Stem Cells (NPCs) grown in standard floating condition (CTRL) and 3 NPCs grown inside the Nichoid (NIC)

first: Federica

last: Rey

first: Cecilia

last: Pandini

first: Bianca

last: Barzaghini

first: Toniella

last: Giallongo

first: Letizia

last: Messa

first: Orietta

last: Pansarasa

first: Matteo

last: Brilli

first: Gian

middle: V

last: Zuccotti

first: Cristina

last: Cereda

first: Manuela

middle: T

last: Raimondi

first: Stephana

last: Carelli


title: Macrophage maturation from blood monocytes is altered in people with HIV, and is linked to serum lipid profiles and activation indices: a model for studying atherogenic mechanisms

city: Cleveland

first: Mark

last: Cameron

state: OH

address: 10900 Euclid Ave

department: Population and Quantitative Health Sciences

institute: Case Western Reserve University

summary: People with HIV (PWH) are at increased risk for atherosclerotic cardiovascular disease (ASCVD). Proportions of vascular homing monocytes are enriched in PWH; however, little is known regarding monocyte-derived macrophages (MDMs) that may drive atherosclerosis in this population.  We isolated PBMCs from people with and without HIV, and cultured these cells for 5 days in medium containing autologous serum to generate MDMs. Differential gene expression (DGE) analysis of MDMs from PWH identified broad alterations in innate immune signaling (IL-1β, TLR expression, PPAR βδ) and lipid processing (LXR/RXR, ACPP, SREBP1).  Transcriptional changes aligned with the functional capabilities of these cells.  Expression of activation markers and innate immune receptors (CD163, TLR4, and CD300e) was altered on MDMs from PWH, and these cells produced more TNFa, reactive oxygen species (ROS), and matrix metalloproteinases (MMPs) than did cells from people without HIV.  MDMs from PWH also had greater lipid accumulation and uptake of oxidized LDL. PWH had increased serum levels of free fatty acids (FFAs) and ceramides, with enrichment of saturated FAs and a reduction in polyunsaturated FAs.  Levels of lipid classes and species that are associated with CVD correlated with unique DGE signatures and altered metabolic pathway activation in MDMs from PWH.  Here, we show that MDMs from PWH display a pro-atherogenic phenotype; they readily form foam cells, have altered transcriptional profiles, and produce mediators that likely contribute to accelerated ASCVD.

overall_design: Comparison of transcriptomic differences among monocyte-derived macrophages isolated from people with and without HIV

first: E

last: Bowman

first: C

first: B

last: Richardson

first: M

last: Kulkarni

first: J

last: Gabriel

last: Cichon

first: K

last: Riedl

first: Y

last: Mustafa

last: Cartwright

last: Snyder

first: S

last: Raman

first: D

last: Zidar

last: Koletar

last: Playford

first: N

last: Mehta

last: Sieg

last: Freeman

last: Lederman

last: Funderburg


title: Comprehensive analysis of metabolic and mitochondrial characteristics of extraembryonic endoderm cells

city: London

first: Mohamed

middle: I

last: Gatie

state: Ontario

address: 1151 Richmond street, Chemistry Building Room 003, Dock 11

department: Biology

country: Canada

institute: University of Western Ontario

summary: The purpose of the study was to identify transcriptomic changes between feeder-free ES and XEN cells. ES cells were cultured feeder-free on 0.1% gelatin in 2iLIF in N2B27 media. XEN cells were also cultured feeder-free on 0.1% gelaton  in RPMI1640 supplemented with 15% FBS, 2mM GlutaMAX and BME.

overall_design: Bulk RNAseq of feeder-free ES and XEN cells under normal conditions

first: Gregory

middle: M

last: Kelly


title: Dissecting the effect of 3D micro-scaffolds on the transcriptome of Neural Precursors Stem Cells with computational approaches: a focus on mechanotrasduction

first: Stella

last: Gagliardi

summary: 3D cell cultures are becoming more and more important in the field of regenerative medicine due to their ability to mimic the cellular physiological microenvironment. Among the different types of 3D scaffolds, we focus on the Nichoid, a miniaturized scaffold with a structure inspired by the natural staminal niche. The Nichoid can activate cellular responses simply by subjecting the cells to mechanical stimuli. This kind of influence results in different cellular morphology and organization, but the molecular bases of these changes remain largely unknown. Through RNA-seq approach on murine Neural Precursors Stem Cells expanded inside the Nichoid, we investigated the deregulated genes and pathways showing that the Nichoid causes alteration in genes strongly connected to mechanobiological functions. Moreover, we fully dissected this mechanism highlighting how the changes start at a membrane level, with subsequent alterations in the cytoskeleton, signaling pathways, and metabolism, all leading to a final alteration in gene expression. The results shown here demonstrate that the Nichoid influences the biological and genetic response of stem cells thorough specific alterations of cellular signaling. The characterization of these pathways elucidates the role of mechanical manipulation on stem cells, with possible implications in regenerative medicine applications.

overall_design: 3 NPCs grown inside the Nichoid (NIC) and replated in standard floating condition for 7 more days

first: Federica

last: Rey

first: Cecilia

last: Pandini

first: Bianca

last: Barzaghini

first: Letizia

last: Messa

first: Toniella

last: Giallongo

first: Orietta

last: Pansarasa

first: Matteo

last: Brilli

first: Gian

last: Zuccotti

last: Cereda

last: Raimondi

last: Carelli


title: Expression data from whole blood from healthy human individuals

city: Stanford

first: Aida

last: Habtezion

state: CA

address: 291 Campus Drive

department: Medicine

institute: Stanford University

summary:
Blood is one of the most common biospecimens used in clinical and translational research. However, minimizing the variability in collection and processing of human blood samples remains a challenge. Delaying plasma or serum isolation after phlebotomy (processing delay) can cause perturbations of numerous analytes used in research studies and clinical practice.
We used microarrays to investigate the effects of time delay and temperature changes on whole blood transcriptomics.

overall_design: Blood from healthy donors was subjected to processing delays that are commonly encountered in blood sample collection: 2hr RT, 5hr RT, 5hr 4C, and 15hr (ON) 4C.

first: Lawrence

last: Bai


title: Gene therapy with secreted acid alpha-glucosidase rescues Pompe disease in a novel mouse model with early-onset spinal cord and respiratory defects

city: Madrid

first: Manuel

middle: J

last: Gomez

state: Madrid

address: Melchor Fernández Almagro, 3

country: Spain

institute: CNIC

summary: Pompe disease (PD) is a neuromuscular disorder caused by deficiency of acid-alpha-glucosidase (GAA), leading to motor and respiratory dysfunctions. Available Gaa knock-out (KO) mouse models do not accurately mimic PD, particularly the highly impaired respiratory phenotype. Methods. Here we developed a new mouse model of PD crossing Gaa KO B6;129 with DBA2/J mice. Findings. Male Gaa KODBA2/J presents most of the key features of the human disease, including early lethality, severe respiratory impairment, cardiac hypertrophy and muscle weakness. Transcriptome analyses of Gaa KODBA2/J, compared to the parental Gaa KOB6;129 mice, revealed a profoundly impaired gene signature in the spinal cord and a similarly deregulated gene expression in skeletal muscle. Muscle and spinal cord transcriptome changes in Gaa KODBA2/J, were significantly improved upon gene therapy with AAV vectors expressing a secreted GAA enzyme.

overall_design: Transcriptomic analysis comparing the gene expression profiles of quadriceps muscle and whole spinal cord from:  Gaa WT and Gaa KO mice in B6;129 background; Gaa WT and Gaa KO mice in  B6;129;DBA2/J background and Gaa KO mice in B6;129;DBA2/J strain treated with AAV-mediated gene therapy with secreted acid-alpha-glucosidase.

first: P

last: Colella

first: M

first: U

last: Cagin

first: F

last: Mingozzi


title: The brlA gene is involved in conidiogenesis and secondary metabolism in Penicillium expansum

city: TOULOUSE

first: Olivier

last: PUEL

address: 180 chemin de Tournefeuille

department: TOXALIM UMR 1331

country: France

institute: INRAE

summary: In Aspergillus sp.,  BrlA is a master transcription factor essential for the induction of conidiogenesis. Sometimes it is also involved in the secondary metabolism. The aim of this study was to characterize the role of BrlA in Penicillium expansum. For that a null mutant PeΔBrlA was generated.

overall_design: PeΔBrlA and WT strains were grown in the same conditions ( 25°C for 5 days in the dark) on MEA medium. The RNAs were extracted and their transcriptomes were analyzed.

first: Chryztian

last: Zetina-Serrano

first: Ophélie

last: Rocher

first: Sophie

last: Lorber

last: Puel

first: Claire

last: Naylies

first: Yannick

last: Lippi


title: RNA sequencing from T47D and MCF-7 breast cancer cells

city: San Antonio

first: Tim

middle: Hui-Ming

last: Huang

state: TX

address: 7703 Floyd Curl Dr

institute: University of Texas Health at San Antonio

summary: We report the RNA sequencingn onT47D and MCF-7 cells infected by either vehicle or shMEN1 virus

overall_design: Investigate downstream genes of MEN1 in breast cancer cells.

middle: H

first: Chun-Liang

last: Chen

first: Chih-Wei

last: Chou

first: Chun-Lin

last: Lin


title: iMEF SDHC-loss stable line histone acetyltransferase inhibitor ATAC-seq

city: Rochester

first: John

middle: Andrew

last: Smestad

state: Minnesota

address: 200 1st St. SW

department: Biochemistry and Molecular Biology

institute: Mayo Clinic

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We have developed a tet-inducible SV-40 large T antigen-expressing lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of mitochondrial electron transport chain (ETC) dysfunction involving complex II (succinate dehydrogenase; SDH) in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl). Following doxycycline induction, dilutional subcloning was employed to obtain stable Sdhc -/- (SDHC KO) cell lines. Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR. Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2. Following cell line derivation and verification of Sdhc genetic status, cells were treated with histone acetyltransferase inhibitors C646 (10 µM) and MB-3 (100 µM) or vehicle for 3 days. Accessible chromatin regions were subsequently interrogated by ATAC-seq.

overall_design: Included in this dataset are dual replicate ATAC-seq data for a SDHC KO (R26M2rtTA/+;TetOcre;Sdhc-/-) iMEF cell line treated with histone acetyltransferase inhibitors C646 (10 µM) and MB-3 (100 µM) or vehicle for 3 days.

first: Louis

last: Maher

middle: A

first: Xiong

last: Yuning


title: Detecting Tumor Specific Antigen-Reactive T cells from Tumor Infiltrating Lymphocytes via Interaction Dependent Fucosyl-biotinylation [TCR-seq]

city: San Diego

first: Peng

last: Wu

address: 10550 North Torrey Pines Road

department: Molecular Medicine

institute: Scripps Research

summary: Reactivation and clonal expansion of tumor specific antigen (TSA)-reactive T cells are key to the success of checkpoint blockade and adoptive transfer of tumor-infiltrating lymphocyte (TIL) based therapies. There are no reliable markers to specifically identify the repertoire of TSA-reactive T cells due to their heterogeneous composition. Here we introduce FucoID as a general platform to detect endogenous antigen-specific T cells and study their biology. Through this interaction dependent labeling approach, TSA-reactive T cells can be detected and separated from bystander T cells in primary tumor digests based on their cell-surface enzymatic fucosyl-biotinylation. Compared to bystander TILs, TSA-reactive TILs possess a unique gene features according to RNA-seq.

overall_design: TCRbeta sequencing of two populations (B- and B+) of TILs isolated from E0771 and MC38 tumor (15 days) on C57BL/6 mice, in triplicate.

first: Zilei

last: Liu


title: Detecting Tumor Specific Antigen-Reactive T cells from Tumor Infiltrating Lymphocytes via Interaction Dependent Fucosyl-biotinylation [RNA-seq]

overall_design: RNA-seq of three populations of TILs isolated from MC38 tumor (15 days) on C57BL/6 mice, in triplicate.


title: Genome-wide transcriptome analysis of the STAT6-regulated genes in advanced-stage cutaneous T-cell lymphoma [scRNA-seq]

city: Pittsburgh

first: Patrizia

last: Fuschiotti

state: PA

address: 200 Lothrop St

institute: University of Pittsburgh

summary: The signal transducer and activator of transcription 6 (STAT-6) is a critical up-stream mediator of IL-13 and IL-4 signaling and is constitutively activated in malignant lymphocytes from Sezary syndrome (SS) and mycosis fungoides (MF), the most common subtypes of cutaneous T-cell lymphomas. By combining genome-wide expression profiling with pharmacological STAT-6 inhibition, we have identified the genes regulated by STAT-6 in MF/SS tumors. We found that STAT-6 regulates several common pathways in MF/SS malignant lymphocytes that are associated with control of cell cycle progression and genomic stability as well as production of Th2 cytokines. Using ex vivo skin explants from cutaneous MF tumors as well as Sezary cells derived from the blood of SS patients, we demonstrated that inhibition of STAT-6 activation down-regulates cytokine production and induces cell cycle arrest in MF/SS malignant lymphocytes, inhibiting their proliferation but not their survival. Furthermore, we show that STAT-6 promotes the pro-tumoral M2-like phenotype of tumor-associated macrophages in the tumor microenvironment of advanced-stage MF by up-regulating the expression of genes associated with immunosuppression, chemotaxis, and tumor matrix remodeling. Thus, we show STAT-6 to be a major factor in the pathogenesis and progression of MF/SS, promoting proliferation and invasion of the malignant lymphocytes while inducing a progressive depression of the anti-tumor immune response. Together, our results provide new insights into disease pathogenesis and offer new prospective targets for therapeutic intervention.

overall_design:
3 CTCL samples were analyzed and 4 normal controls were analyzed.

***Raw Data will be submitted to dbGAP***

first: Alyxzandria

last: Gaydosik

first: Dawn

middle: S

last: Queen

first: Oleg

middle: E

last: Akilov

first: Larisa

last: Geskin


title: Genome-wide transcriptome analysis of the STAT6-regulated genes in advanced-stage cutaneous T-cell lymphoma

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

overall_design: Refer to individual Series


title: Senescence induced by BMI1 inhibition is a therapeutic vulnerability in H3K27M-mutant DIPG

city: Aurora

first: Etienne

last: Danis

state: CO

institute: University of Colorado, Denver - Anschutz Medical Campus

summary:
We studied the effect of BMI1 inhibition in diffuse intrinsic pontine glioma cells (DIPG).
We reported the application of chromatin immunoprecipitation sequencing for high-thoughput profiling of BMI1 and H2AK119ub in diffuse intrinsic pontine glioma cells (DIPG) in response to BMI1 inhibition.
We also performed bulk RNA sequencing of DIPG cells in response to pharmacological inhibition or knockdown of BMI1.

overall_design:
ChIP-seq samples were obtained using anti-BMI1 and anti-H2AK119ub antibodies in SU-DIPG04 and SF8628 DIPG cells with or without the BMI1 inhibitior, PTC-028.
RNA-seq samples were obtained from SU-DIPG04 and SF86828 DIPG cells treated with or without PTC-028 and from SU-DIPG04 and SF7761 DIPG cells expressing control shRNA or BMI1 shRNAs.

first: Ilango

last: Balakrishnan

first: Rajeev

last: Vibhakar

first: Sujatha

last: Venkataraman


title: Genome-wide transcriptome analysis of the STAT6-regulated genes in advanced-stage cutaneous T-cell lymphoma [Bulk-seq]

type: Expression profiling by genome tiling array

overall_design: 3 MF samples, 4 SZ samples and 3 normal control samples were cultured for 48 hours with or without the STAT6 inhibitor and then analyzed with RNAseq.


title: Efficient and cost-effective bacterial mRNA sequencing from low input samples through ribosomal RNA depletion

city: Santa Barbara

first: Kellie

last: Heom

state: California

address: UC Santa Barbara

department: Chemical Engineering

institute: University of California Santa Barbara

summary: RNA sequencing is a powerful approach to quantify the genome-wide distribution of mRNA molecules in a population to gain deeper understanding of cellular functions and phenotypes. However, unlike eukaryotic cells, mRNA sequencing of bacterial samples is more challenging due to the absence of a poly-A tail that typically enables efficient capture and enrichment of mRNA from the abundant rRNA molecules in a cell. Moreover, bacterial cells frequently contain 100-fold lower quantities of RNA compared to mammalian cells, which further complicates mRNA sequencing from non-cultivable and non-model bacterial species which are often present in low abundance. To overcome these limitations, we report EMBR-seq (Enrichment of mRNA by Blocked rRNA), a method that efficiently depletes 5S, 16S and 23S rRNA using blocking primers to prevent their amplification. EMBR-seq results in greater than 80% of the sequenced RNA molecules deriving from mRNA. We demonstrate that this increased efficiency provides a deeper view of the transcriptome without introducing technical amplification-induced biases. Moreover, compared to recent methods that employ a large array of oligonucleotides to deplete rRNA, EMBR-seq employs a single oligonucleotide per rRNA, thereby making this new technology significantly more cost-effective, especially when applied to varied bacterial species. Finally, compared to existing commercial kits, we show that EMBR-seq can be used to successfully quantify the transcriptome from more than 500-fold lower starting total RNA. Thus, EMBR-seq provides an efficient and cost-effective approach to quantify global gene expression profiles from low input bacterial samples.

sample_organism: Escherichia coli str. K-12 substr. MG1655

overall_design: In this study, we developed a new bacterial mRNA-sequencing method (EMBR-seq) by depleting rRNA from total RNA. After validating the method, we used EMBR-seq to deplete rRNA and quantify gene expression from low input material, applying our method to 100, 20, 2, 0.2 and 0.02 ng of starting total RNA.

first: Chatarin

last: Wangsanuwat

first: Estella

first: Michelle

last: O'Malley

first: Siddharth

last: Dey


title: MicroRNA profiling of H9N2 subtype avian influenza virus-infected chicken dendritic cells using high-throughput sequencing

city: nanjing

first: liu

last: qingtao

state: jiangsu

address: zhongling road NO. 50

department: Institute of veterinary medicine

country: China

institute: jiangsu academy of agricultural sciences

type: Non-coding RNA profiling by high throughput sequencing

summary: The microRNA expression changes were analysed in chicken dendritic cells during H9N2 avian influenza virus infection

overall_design: MicroRNA expression profiles of chicken DCs infected without or with H9N2 influenza virus were generated by deep sequencing, in triplicate, using Illumina HiSeq X

first: Qingtao

first: Jing

last: Yang


title: Reponses of Galleria mellonella hemocytes to  Beauveria bassiana

city: Hangzhou

first: Shenghua

last: Ying

state: Zhejiang

address: Yuhangtang Road 866

institute: Zhejiang University

summary: Insect cellular immunue plays an important role in recognition and eliminating the intruding entomopathogenic fungi.  The genome-wide exprssion analysis involved in the cellular responses was analyzed by using high throughput sequencing (RNA-Seq). Our transcriptional profiles revealed that numerous differentially expressed genes (DEGs), of which involved in metabolism, immune system, cellular processing and so on, were significantly involved in the responses of G. mellonnella hemocytes.

overall_design: Total RNA was obtained from G. mellonella hemocytes challenged by B. bassiana at different time-points. Comparison were performed between the indicated time-point and control group (without infection by fungus) .

first: Sheng-Hua

first: Ming-Guang

last: Feng


title: GLI1 ChIP-Seq in non-irradiated versus irradiated SUM1315 cells

city: Birmingham

first: Dongquan

last: Chen

state: AL

address: 1717 11th Ave South

institute: Univ of Alabama at Birmingham

summary: Changes in the GLI1 cistrome following ionizing radiation were investigated using ChIP-Seq in SUM1315 triple-negative breast cancer cells.

overall_design: 8 samples analyzed in total, 2 conditions with paired 2% input controls done in duplicate, SUM1315 cells 4 hours after 4 Gy ionizing radiation or nonirradiated

first: Tshering

middle: D

last: Lama-Sherpa

first: Victor

last: Lin

first: Brandon

last: Metge

first: Shannon

last: Weeks

last: Samant

first: Lalita

last: Shevde


title: A transcriptomic overview of lung and liver changes one day after pulmonary exposure to graphene and graphene oxide

status: Public on Oct 21 2020

city: Ottawa

first: Sabina

last: Halappanavar

address: 251 Sir Frederick Banting Driveway, Building 22

institute: Health Canada

summary: This study investigates transcriptomic differences in the lung and liver after pulmonary exposure to two Graphene based materials with similar physical properties, but different surface chemistry. Female C57BL/6 mouse were exposed by a single intratracheal instillation of 0, 18, 54 or 162 μg/mouse of graphene oxide (GO) or reduced graphene oxide (rGO). Pulmonary and hepatic transcriptional changes were compared to identify commonly and uniquely perturbed functions and pathways by GO and rGO. These changes were then related to previously analyzed endpoints. GO exposure induced more differentially expressed genes, affected more functions, and perturbed more pathways compared to rGO, both in the lung and liver.

overall_design: Lung tissue were collected for total RNA extraction from seven weeks old C57BL/6J female mice exposed to 0, 18, 54 and 162 µg/mouse of graphene oxide (GO) or reduced graphene oxide (rGO) or or carbon black (CB) or vehicle control for 1 day, 28 days and 90 days.

first: Stefan

last: Bengtson

first: Andrew

last: William

first: Ulla

last: Vogel


title: iMEF SDHC-loss stable line ATAC-seq

first: John

last: Smestad

summary: We have developed a tet-inducible SV-40 large T antigen-expressing lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of mitochondrial electron transport chain (ETC) dysfunction involving complex II (succinate dehydrogenase; SDH) in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl).  For comparison, we include an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt) that retains one intact copy of Sdhc upon doxycycline exposure.  Following doxycycline induction of both cell lines, dilutional subcloning was employed to obtain stable Sdhc -/- (SDHC KO) and Sdhc +/- (Control) cell lines.  Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR.  Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2.  Following cell line derivation and verification of Sdhc genetic status, accessible chromatin regions in each cell line were interrogated by ATAC-seq.

overall_design: Included in this dataset are single replicate ATAC-seq data for two SDHC KO (R26M2rtTA/+;TetOcre;Sdhc-/-) and  two control (R26M2rtTA/+;TetOcre;Sdhc+/-) cell lines derived by dilutional subcloning following induction of Sdhc gene rearrangement with doxycycline.

first: Louis

last: Maher

first: Xiong

last: Yuning


title: iMEF SDHC-loss stable line CUT&RUN

summary: We have developed a tet-inducible SV-40 large T antigen-expressing lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of mitochondrial electron transport chain (ETC) dysfunction involving complex II (succinate dehydrogenase; SDH) in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl).  For comparison, we include an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt) that retains one intact copy of Sdhc upon doxycycline exposure.  Following doxycycline induction of both cell lines, dilutional subcloning was employed to obtain stable Sdhc -/- (SDHC KO) and Sdhc +/- (Control) cell lines.  Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR.  Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2.  Following cell line derivation and verification of Sdhc genetic status, chromatin epigenomic marks including H3K4me3, H3K27me2, H3K27ac, CTCF, acetyllysine, propionyllysine, and butyryllysine were characterized by CUT&RUN.

overall_design: Included in this dataset are biological replicate CUT&RUN data for two SDHC KO (R26M2rtTA/+;TetOcre;Sdhc-/-) and  two control (R26M2rtTA/+;TetOcre;Sdhc+/-) cell lines derived by dilutional subcloning following induction of Sdhc gene rearrangement with doxycycline.  For each line, epigenomic marks including H3K4me3, H3K27me2, H3K27ac, CTCF, acetyllysine, propionyllysine, and butyryllysine were characterized by CUT&RUN.


title: Subunit vaccine elicited lung TRM cells use fibroblast IL-17R signaling to provide serotype independent immunity against hypervirulent K. pneumoniae

city: New Orleans

first: Jay

last: Kolls

state: LA

address: 1430 Tulane Ave

institute: Tulane University

summary: Subunit vaccine conferred lung protection against heterologous strain through the augmentaion of lung tissue resident memmory CD4+ T-cells.

overall_design: ①We immunized WT C57BL/6 mice with OmpX+LTA1 or heat killed Klebsiella pneumoniae (ATCC 43816) twice 3 weeks apart, and 1 more week later we euthanized and removed the lung for single cell suspensions in addition to naive spleen cells. ②Our data had indicated that 24 hours post pulmonary infection of K1 strain at 10e4 CFU reveals quite higher bacterial burden in Il17raDermoCre postivie mice than in Il17raDermoCre negative mice, and therefore we harvested lung tissues to obatin RNA for bulk RNAseq at 4 hours post infection because any differences in gene expression would not be due to differences in lung bacterial burdens.

middle: K


title: An integrated multi-omics approach identifies epigenetic alterations associated with Alzheimer’s disease (5hmC) [5hmC-Seal]

city: Philadelphia

first: Yemin

last: Lan

address: 3400 Civic Center Blvd

institute: University of Pennsylvania

type: Methylation profiling by high throughput sequencing

summary: We report the 5hmC in human hippocampus subject to normal aging and Alzheimer's disease.

overall_design: 5hmC-Seal in postmortal lateral temporal lobe of Alzheimer's disease affected brains (AD), control Old (Old) and control Young (Young).

first: Raffaella

last: Nativio

first: Berger

last: Shelley


title: Endothelial GATA4 controls liver fibrosis and regeneration by preventing a pathogenic switch in angiocrine signaling

city: Mannheim

first: Gergana

last: Dobreva

state: Baden Württemberg

address: Ludolf-Krehl-Str. 7-11

department: Cardiovascular Genomics and Epigenomics

country: Germany

institute: Medical Faculty Mannheim/University of Heidelberg


title: Endothelial GATA4 controls liver fibrosis and regeneration by preventing a pathogenic switch in angiocrine signaling [ATAC-seq]

summary: Angiocrine signaling by liver sinusoidal endothelial cells (LSEC) regulates liver functions such as liver growth, metabolic maturation, and regeneration. Recently, we identified GATA4 as the master regulator of LSEC specification during development. Here, we studied endothelial GATA4 in the adult liver and in hepatic disease pathogenesis.   We generated adult Clec4g-icretg/0xGata4fl/fl (Gata4LSEC KO) mice with deficiency of Gata4 in LSEC. Livers were analyzed by histology, electron microscopy, immunohistochemistry/immunofluorescence, in-situ hybridization, and by expression profiling and ATAC-sequencing of isolated LSEC. For liver regeneration, partial hepatectomy was performed. As models of liver fibrosis, CDAA diet and chronic CCl4 exposure were applied. Human single cell RNAseq data sets were analyzed for endothelial alterations in healthy and cirrhotic livers.  Genetic Gata4 deficiency in LSEC in adult mice caused perisinusoidal liver fibrosis, hepatopathy and impaired liver regeneration. Sinusoidal capillarization and LSEC-to-continuous endothelial transdifferentiation were accompanied by a profibrotic angiocrine switch including de novo endothelial expression of hepatic stellate cell-activating cytokine PDGFB. Increased chromatin accessibility and amplification by activated Myc mediated angiocrine PDGFB expression. In CDAA diet-induced perisinusoidal liver fibrosis, LSEC showed repression of GATA4, activation of MYC and the profibrotic angiocrine switch already detected in Gata4LSEC KO mice. Comparison of CDAA-fed Gata4LSEC KO and control mice demonstrated that endothelial Gata4 indeed protects from dietary-induced perisinusoidal liver fibrosis. In human cirrhotic livers, Gata4-positive LSEC and endothelial Gata4 target genes were reduced, while non-LSEC endothelial cells and Myc target genes including PDGFB were enriched.   Endothelial GATA4 protects from perisinusoidal liver fibrosis by repressing MYC activation and profibrotic angiocrine signaling on the chromatin level. Therapies targeting the GATA4/MYC/PDGFB/PDGFRβ axis offer a promising strategy for the prevention and treatment of liver fibrosis.

overall_design: Six samples

first: Julio

last: Cordero

first: Christian

last: Schmid


title: The short- and long-range RNA Interactome of SARS-CoV-2

city: cambridge

first: Jonathan

middle: Lewis

last: Price

address: The Sanger Building Tennis Court Road

department: Biochemistry

country: United Kingdom

institute: University of Cambridge

summary: The Coronaviridae are a family of positive- strand RNA viruses that includes SARS-CoV-2, the etiologic agent of the COVID-19 pandemic. Bearing the largest single stranded RNA genome in nature, coronaviruses are critically dependent on long-distance RNA-RNA interactions to regulate the viral transcription and replication pathways. Here we experimentally mapped the in vivo long-range RNA interactome of the full-length SARS-CoV-2 genome and the subgenomic mRNAs. We uncover a network of RNA-RNA interactions spanning tens of thousands of nucleotides that facilitate the unique transcription mode of coronaviruses, and reveal that the viral genome adopts alternative topologies inside cells and undergoes genome cyclization. Moreover, we discover long RNA-bridges between adjacent open reading frames that encircle the programmed ribosomal frame-shifting element, and demonstrate their conservation in vivo for the related MERS-CoV. Finally, the SARS-CoV-2 genome and subgenomic mRNAs engage in different sets of interactions with cellular RNAs. Our findings illuminate RNA-based mechanisms governing replication, discontinuous transcription, and translation of coronaviruses, and will aid future efforts to develop antiviral strategies.

sample_organism:
Homo sapiens
Chlorocebus pygerythrus

platform_organism:
Homo sapiens
Chlorocebus pygerythrus

overall_design: We recently developed Crosslinking Of Matched RNAs And Deep Sequencing (COMRADES) for in-depth RNA conformation capture in living cells (Ziv et al., 2018). COMRADES can detect base-paired regions in RNA inside the cell,  using a clickable psoralen derivative to specifically crosslink double-stranded RNA, and high throughput sequencing to retrieve the base-pairing information (Figure S1A). Following in vivo crosslinking, the viral RNA is selectively captured, fragmented and subjected to a click-chemistry reaction to add a biotin tag to crosslinked regions. Crosslinked RNA duplexes are then selectively captured using streptavidin affinity purification. Half of the resulting RNA is proximity ligated, following reversal of the crosslink to create chimeric RNA templates for high throughput sequencing. The other half is used as a control, in which reversal of the crosslink precedes the proximity ligation, and accurately represents the background level of non-specific ligation. The coupling of two enrichment steps, first of viral RNA, and second of crosslinked RNA duplexes provides high structural depth for identification of both long- and short-lived conformations. COMRADES can therefore measure (i) the structural diversity of alternative RNA conformations that coexist inside cells; (ii) short-distance, as well as long-distance (over tens of thousands of nucleotides) base-pairing within the same RNA molecule; and (iii) base-pairing between different RNA molecules, such as those of host and viral origin (Kudla et al., 2020; Ziv et al., 2018).     Here we applied COMRADES to study the structural diversity of SARS-CoV-2 gRNA and sgmRNAs inside cells. Whereby we have conducted in duplicate - MERS and SARS-CoV-2 gRNA as well as MERS and SARS-CoV-2 sgmRNAs.

first: Omer

last: Ziv

first: Lyudmila

last: Shalamova

first: Tsveta

last: Kamenova

first: Ian

last: Goodfellow

first: Friedemann

last: Weber

first: Eric

last: Miska


title: DNA methylation analysis in malignant melanoma clinical samples [EPIC array]

city: Chiba

first: Masaki

last: Fukuyo

address: 1-8-1 Inohana, Chuo-ku

department: Department of Molecular Oncology

country: Japan

institute: Chiba University

type: Methylation profiling by genome tiling array

summary: Comprehensive DNA methylation analysis in malignant melanoma clinical samples and primary melanocyte culture. Infinium MethylationEPIC BeadChip was used to obtain DNA methylation profiles. Samples included 8 malignant melanoma cases and primary melanocyte culture.

overall_design: Bisulphite converted DNA from the 8 samples and primary melanocyte culture were hybridised to the Infinium HumanMethylation450 BeadChip.

first: Yosuke

last: Yamamoto

first: Keisuke

last: Matsusaka

first: Bahityar

last: Rahmutulla

first: Hiroyuki

last: Matsue

first: Atsushi

last: Kaneda


title: DNA methylation analysis in malignant melanoma clinical samples


title: DNA methylation analysis in malignant melanoma clinical samples [450K array]

summary: Comprehensive DNA methylation analysis in malignant melanoma clinical samples. Infinium HumanMethylation450 BeadChip was used to obtain DNA methylation profiles across 485,577 CpG sites. Samples included 54 malignant melanoma cases and primary melanocyte culture.

overall_design: Bisulphite converted DNA from the 45 samples were hybridised to the Infinium HumanMethylation450 BeadChip.


title: Whole-genome DNA methylation profiling of ESR1 Q375H and R394H mutations from the human breast cancer MB-231 stable cells.

city: RTP

first: NIEHS

last: Microarray Core

state: NC

address: 111 T.W. Alexander Drive

department: DIR

institute: NIEHS

summary:
Estrogen insensitivity syndrome (EIS) arises from rare mutations in estrogen receptor α (ERα, encoded by ESR1 gene) resulting in the inability of estrogen to exert its biological effects. Due to the rarity, mutations in ESR1 gene and the underlying molecular mechanisms of EIS have not been thoroughly studied.
We used human EPIC DNA methylation array to comparison of the transcriptome between the WT ESR1 and Q375H and R394H clinical mutants in MB231 stable cell lines.

overall_design: MB231 cells stably exressing the WT, Q and R mutation were tretaed with veh, E2 or DES for 18 hours.

first: Yin

last: Li

first: Laura

last: Wharey

first: Kevin

last: Gerrish

first: Jian-Liang


title: Trib1 promotes acute myeloid leukemia progression by modulating transcriptional programs of Hoxa9

city: Tokyo

first: Takuro

last: Nakamura

address: 3-8-31 Ariake, Koto-ku

department: The Cancer Institute

institute: Japanese Foundation for Cancer Research

overall_design: Refer to individual Series


title: Trib1 promotes acute myeloid leukemia progression by modulating transcriptional programs of Hoxa9 [array]

summary:
The pseudokinase Trib1 functions as a myeloid oncogene that recruits E3 ubiquitin ligase COP1 to C/EBPa, and interacts with MEK1 to enhance ERK phosphorylation. Close genetic interaction between Trib1 and Hoxa9 has been observed in myeloid leukemogenesis as Trib1 overexpression significantly accelerates Hoxa9-induced leukemia onset. Yet the mechanism how Trib1 functionally modulates Hoxa9 transcription activity is unclear. Here we provide evidence that Trib1 modulates Hoxa9-associated super-enhancers. ChIP-seq analyses identified increased Histone H3K27Ac signals at the super-enhancers such as Erg, Aff3, Spns2 and Coro2 loci, as well as increased mRNA expression of these genes. The modification of super-enhancer activities is mostly achieved by degradation of C/EBPa by Trib1 and the MEK/ERK pathway only slightly contributes to the activities. Silencing of Erg abrogates the growth advantage acquired by Trib1 overexpression, indicating that Erg is a critical downstream target of the Trib1/Hoxa9 axis. Moreover, treatment of AML cells with the BRD4 inhibitor JQ1 showed growth inhibition in the Trib1/Erg-dependent manner. Upregulation of ERG by TRIB1 was also observed in human AML cell lines, suggesting that Trib1 is a potential target of Hoxa9-associated AML.
We used microarrays to detail the global program of gene expression in mouse AML

overall_design: Murine AML cell lines expressing Hoxa9 with or without Trib1 were examined.

first: Seiko

last: Yoshino


title: Transiently enhanced multi-drug resistance by epigenetic mechanism from reprogrammed cancer cell model

city: Seoul

first: Yourae

last: Hong

address: 81, Irwon-ro

department: Samsung Genome Institute

country: South Korea

institute: Samsung Medical Center

summary: Background: Drug resistance by epigenetic modulation in cancer cells has been suggested, and the epigenetic-driven heterogeneity has been suggested as an important mechanism. To elucidate the epigenetic mechanism further, epigenetically reprogrammed cancer (R-cancer) cells were established and their resistance during differentiation was analyzed.  Methods: R-cancer cells for H460 (R-H460) were established by transient introduction of reprogramming factors. Then, differentiated R-H460 (dR-H460) cells were prepared by withdrawal of stem cell media (SCM) for various durations, and the changes in drug resistance were tested.  Results: dR-H460 cells with SCM-withdrawal for about 2 weeks (mdR-H460 cells) formed significantly more drug-resistant colonies to both cisplatin and paclitaxel. The resistant phenotype of the cisplatin resistant colonies from mdR-H460 cells, however, was lost after long-term (about 70-90 days) cisplatin-withdrawal, suggesting a transient epigenetic mechanism. In another R-cancer cell model with N87, the increased drug-resistance was also observed for both cisplatin and paclitaxel after SCM-withdrawal. In single cell analyses, heterogeneity did not increase significantly in mdR-H460, although relatively higher fraction of mdR-H460 cells was observed in several clusters of parent H460 cells.  Conclusion: Based on R-cancer model, increased cancer cell population under epigenetic transition, rather than heterogeneity from epigenetic changes, may confer the epigenetic-driven drug resistance, which can provide a new strategy to fight cancers by control of cancer cells under epigenetic transition.

overall_design: Single cell RNA sequencing was performed on H460, mdR-H460, and ldR-H460 cells. After quality filtration and integration of  parent H460 and two epigenetic H460 variants of mdR-H460 and ldR-H460, we obtained an integrated single cell transcriptome for 14,638 cells.

first: Shiv

last: Poojan

first: Seung-Hyun

last: Bae

first: Jae-Woong

last: Min

first: Eun

middle: Y

last: Lee

first: Yura

last: Song

first: Hee

last: Kim

first: Hye

middle: W

last: Sim

first: Eun-Kyung

last: Kang

first: Young-Ho

first: Hae-Ock

first: Woong-Yang

last: Park

first: Hyonchol

last: Jang

first: Kyeong-Man


title: The Thermus thermophilus DEAD-box protein Hera is a general RNA chaperone and plays a key role in tRNA metabolism

city: La Jolla

first: Gene

last: Yeo

address: 2880 Torrey Pines Scenic Dr. Room 3805/Yeo Lab

institute: UCSD

summary: RNA helicases catalyze the ATP-dependent destabilization of RNA duplexes. DEAD-box helicases share a helicase core that mediates ATP binding and hydrolysis, RNA binding and unwinding. Most members of this family contain domains flanking the core that can confer RNA substrate specificity and guide the helicase to a specific RNA. However, the in vivo RNA substrates of most helicase are currently not defined.   The DEAD-box helicase Hera from Thermus thermophilus contains a helicase core, followed by a dimerization domain (DD) and an RNA binding domain (RBD) that folds into an RNA recognition motif (RRM). The RRM mediates high affinity binding to an RNA hairpin, and an adjacent duplex is then unwound by the helicase core. Hera is a cold-shock protein, and has been suggested to act as an RNA chaperone under cold-shock conditions. Using immunoprecipitation of Hera-RNA complexes and RNA-Seq, we show that Hera binds to a large fraction of T. thermophilus RNAs under normal-growth and cold-shock conditions without a strong sequence preference, in agreement with a structure-specific recognition of RNAs and a general function in RNA metabolism. Under cold-shock conditions, Hera is recruited to RNAs with high propensities to form stable secondary structures. We show that selected RNAs identified, including a set of tRNAs, bind to Hera in vitro, and activate the Hera helicase core. Gene ontology analysis reveals an enrichment of genes related to translation, including mRNAs of ribosomal proteins, tRNAs, tRNA ligases, and tRNA-modifying enzymes, consistent with a key role of Hera in ribosome assembly and tRNA metabolism.

overall_design: eCLIP-seq in Thermus thermophilus HB27 (DSM 7039, DSMZ; ATCC BAA-163)

first: Pascal

last: Donsbach

first: Brian

last: Yee

first: Stefan

last: Aigner

first: Dagmar

last: Klostermeier


title: Experimental data of ex vivo treated human cord blood (CB) derived hematopoietic stem and progenitor cells (HSPCs) with Ciclopirox ethanolamine.

status: Public on Oct 20 2020

city: Lund

first: Mehrnaz

last: Safaee Talkhoncheh

address: Sölvegatan 17

country: Sweden

institute: Lund University

summary: Ciclopirox ethanolamine (CPX), is a novel regulator of HSCs and preserve the primitive stem cell state of these cells. By treateing human CB-HSPCs with CPX we observed a substantial increas on number of stem and porgenitor cells detected by phenotypic monitoring of stem cell markers. we performed this microarray analysis to identify the differentialy expressed genes upon CPX treatment.

overall_design: Cord blood-derived enriched fraction (CD34+CD90+) cells were treated with either DMSO control or CPX for 24 hours or 6 days ex vivo. Cells were harvested at these time points and analyzed for differentially expressed genes.

first: Jonas

last: Larsson


title: Tumor Repopulating Cell Transcriptome reveals Specific Cell Surface Biomarkers and Signaling Pathways

city: Bar Harbor

first: Michael

middle: C.

last: Saul

state: ME

address: 600 Main St

institute: The Jackson Laboratory

summary: Cancer metastasis is the most deadly stage in cancer progression. Despite significant efforts over the last few decades, it remains elusive why only a very small fraction of tumorigenic cancer cells is able to generate metastatic colonization. Recently, we have shown that mechanically selected tumor-repopulating cells (TRCs), a highly tumorigenic subpopulation of mouse melanoma cells, can grow in 3D soft fibrin gels. However, the novel cell surface biomarkers and gene regulatory features of TRCs remain largely unknown. Here, we utilized RNA-sequencing method that uses next-generation sequencing (NGS) techniques for whole transcriptome profiling to elucidate novel cell surface biomarkers and active gene regulatory features. RNA-sequencing data was utilized in DAVID for identification of differentially expressed clusters of genes, including cell adhesion cluster. This allowed us to identify novel cell surface biomarkers such as Col2a1, NCAM1, F11R, and Negr1. We have validated the expression of these genes by real-time qPCR. Among the biomarkers, expression level of Col2a1 was found to be low in TRCs but 20 fold higher compared to parental B16F1 cells, thus making the marker very specific for TRCs. Our immunofluorescence microcopy study also shows high expression of Col2a1 in TRCs compared with parental B16F1 cells. KEGG pathway analysis showed JAK/STAT pathway, hypoxia, and Akt3 pathway to be highly active in TRCs. In addition, aerobic glycolysis pathway was found to be very active, indicating a typical Warburg effect of highly tumorigenic cells. Together, our study revealed novel and specific biomarkers of melanoma TRCs and their cryptic cell signaling pathways. Future studies will uncover if perturbation of these pathways can successfully neutralize TRCs.

overall_design: Four total samples were collected: two control samples (A1 and A3) and two tumor repopulating cell samples (B1 and B3).

first: Bhavana

last: Talluri

first: Kshitij

last: Amar

first: Tasnim

last: Shireen

first: Vjollca

last: Konjufca

first: Jian

last: Ma

first: Taekjip

last: Ha

first: Farhan

last: Chowdhury


title: iMEF SDHC-loss stable line eHi-C

city: Rochester

first: John

last: Smestad

address: 200 1st St. SW

institute: Mayo Clinic

summary: We have developed a tet-inducible SV-40 large T antigen-expressing lentivirus-immortalized mouse embryonic fibroblast (iMEF) cell culture model of mitochondrial electron transport chain (ETC) dysfunction involving complex II (succinate dehydrogenase; SDH) in which silencing gene rearrangement of the Sdhc floxed allele is driven by doxycycline-dependent expression of cre-recombinase from a tet-inducible promoter (R26M2rtTA/+;TetOcre;Sdhcfl/fl).  For comparison, we include an isogenic Sdhc wt control line (R26M2rtTA/+;TetOcre;Sdhcfl/wt) that retains one intact copy of Sdhc upon doxycycline exposure.  Following doxycycline induction of both cell lines, dilutional subcloning was employed to obtain stable Sdhc -/- (SDHC KO) and Sdhc +/- (Control) cell lines.  Sdhc gene rearrangement status for each clonally-derived cell line was confirmed by PCR.  Cell lines were grown in standard DMEM containing penicillin/streptomycin antibiotics (0.5 mg/mL), non-essential amino acids (100 micromolar each of glycine, alanine, asparagine, aspartic acid, glutamic acid, proline, and serine), sodium pyruvate (1 mM), and HEPES buffer (10 mM) at 21% O2 and 5% CO2.  Following cell line derivation and verification of Sdhc genetic status, long-range genomic contacts were profiled using eHi-C.

overall_design: Included in this dataset are biological replicate eHi-C data for two SDHC KO (R26M2rtTA/+;TetOcre;Sdhc-/-) and  two control (R26M2rtTA/+;TetOcre;Sdhc+/-) cell lines derived by dilutional subcloning following induction of Sdhc gene rearrangement with doxycycline.  For each cell line, genome-wide topological contact maps were generated using eHi-C.

first: Louis

last: Maher


title: Inhibitory effects of Fusobacterium nucleatum on osteogenic differentiation of osteoblasts and its molecular mechanism

city: 济南市

first: Hui

last: Gao

state: 山东省

address: 济南药谷（经十东路邢村立交桥东）A座2504室

institute: 山东大学

summary: Background: F.nucleatum is one of the important pathogens of periodontitis, and the main symptom of periodontitis is alveolar bone loss. The study of F.nucleatum in alveolar bone loss caused by periodontitis is still very insufficient. In this study, the biological behavior of osteoblasts stimulated by F.nucleatum was studied to elucidate the mechanism of bone loss in periodontitis.  Methods:A primary rat calvarial osteoblast culture model was established and invaded repeatedly with F.nucleatum (ATCC 25586). Transcriptome analysis at multiple time points(days 1, 3, 7, 14, 21 and 28) was applied to identify gene expression changes in osteoblasts stimulated by F.nucleatum. Cell-counting and EDU-labeling assay were used to analyze the effects of F.nucleatum on the cell proliferation. Flow cytometry and enzyme linked immunosorbent assay (ELISA) was used to analysis the cell apoptosis, cell cycle and secreted inflammatory cytokines changes. ALP activity testing, Alizarin red mineralized nodules staining and calcium content assays, qPCR and western blot were carried on to evaluate the osteogenic differentiation and mineralization of osteoblasts.   Results: Cytology experiments and RNA-seq analysis showed that F.nucleatum could inhibit cell proliferation, promote cell apoptosis, block cell cycle of osteoblasts and promote inflammatory cytokines production in a dose- and time-dependent manner. Besides, F. nucleatum could significantly inhibited osteoblasts differentiation and mineralization via a series inhibition of the differentiation regulatory genes and proteins. Whole-transcriptome analysis identified a total of 235 transcripts that were differentially expressed in all six time-points, most of which were inflammatory-related genes. The genes, Ccl2, Ccl20, Csf1, Cx3cl1, Cxcl1, Cxcl3, Il6, Birc3, Map3k8, Nos2, Nfkb2, Tnfrsf1b and Vcam1, played core roles in a PPI network and interacted closely with other ones in the infection. In addition, 133 osteogenesis-related DEGs were time-serially dynamically changed in a short time-series expression miner (STEM) analysis, which were enriched in multiple cancer-related pathways. The core dynamic DEGs (Mnda, Cyp1b1, Comp, Phex, Mmp3, Tnfrsf1b, Fbln5 and Nfkb2) had been reported to be closely related to the development and metastasis in tumor and cancer progress. This study is the first to evaluate the long-term interaction of F. nucleatum on osteoblasts, which might increase the risk of cell carcinogenesis of normal osteoblasts, and provides new insight into the pathogenesis of bacterial-induced bone destruction.

overall_design: In order to observe the changes of osteoblasts under F.nucleatum stimulation at the genetic level,  mRNA was were analyzed from 36 cell samples cultured in osteogenic inductive medium (OIM, medium supplemented with 10% FBS, 50 mg/L ascorbic acid and 10 mmol/L β-glycerophosphate (Sigma-Aldrich)) at days 1 , 3, 7, 14, 21, 28 from 3 individuals (each individual involved 12 samples with or without F. nucleatum infected (MOI=0, 50)) .

first: Qiang

last: Feng

first: Jun

last: Zhang


title: Regulation of microRNA-221, -222, -21 and -27 in articular cartilage subjected to abnormal compressive forces

city: NEWCASTLE UPON TYNE

first: David

last: Young

address: Centre For Life

institute: Newcastle University

summary:
Objective: microRNAs (miRs) are small non-coding molecules that regulate post-transcriptional target gene expression and are involved in mechano-regulation of cellular activities in all physiological systems. It is unknown whether such epigenetic mechanisms are regulated in response to increasing magnitudes of load. This study investigated mechano-regulation of miRs in articular cartilage subjected to ‘physiological’ and ‘non-physiological’ compressive loads in vitro as a model system and validated findings in an in vivo model of abnormal joint loading.

Design: Bovine full-depth articular cartilage explants were loaded to 2.5MPa (physiological) or 7MPa (non-physiological) (1Hz, 15 minutes) and mechanically-regulated miRs identified using Next Generation Sequencing and verified using quantitative PCR. Downstream targets were verified using miR-specific mimics or inhibitors in conjunction with 3’-UTR luciferase activity assays.

Results: A subset of miRs were mechanically-regulated in ex vivo cartilage explants and in vivo joint cartilage. miR-221, miR-222, miR-21-5p, and miR-27a-5p were increased and miR-483 levels decreased with increasing load magnitude. Tissue Inhibitor of Metalloproteinase 3 (TIMP3) and Cytoplasmic Polyadenylation Element Binding Protein 3 (CPEB3) were identified as putative downstream targets.

Conclusion: Our data confirms miR-221 and -222 mechano-regulation and demonstrates novel mechano-regulation of miR-21-5p and miR-27a-5p in ex vivo and in vivo cartilage loading models. TIMP3 and CPEB3 are putative miR targets in chondrocytes. Identification of specific miRs that are regulated by increasing load magnitude, and their potential to impact on tissue homeostasis, has direct relevance to other mechano-sensitive physiological systems and provides a mechanism by which load-induced tissue behaviour is regulated, both in health and pathology.

overall_design: Bovine cartilage was cultured and mechanically loaded for various times and loads. RNA was isolated and subjected to small RNA-seq

first: Emma

last: Blain

first: Andrew

last: Skelton

first: Paulina

last: Stadnik


title: Transcriptome of rice idr1-1 mutant

city: Nanjing

first: gui

middle: Hong

last: La

address: XuanWu

institute: Nanjing Agricultural University

summary: To discover new mutants conferring enhanced tolerance to drought stress, we screened a mutagenized rice (O. sativa) population (cv. IAPAR9) and identified a mutant, named idr1-1 (for increased drought resistance 1-1), with obviously increased drought tolerance under upland field conditions. The idr1-1 mutant possessed a significantly enhanced ability to tolerate high-drought stresses. Map-based cloning revealed that the gene LOC_Os05g26890 (corresponding to D1 or RGA1 gene), residing in the mapping region of IDR1 locus, carried a single-base deletion in idr1-1 mutant. IDR1 protein was localized in nucleus and to plasma membrane or cell periphery. Further investigations indicated that the significantly increased drought tolerance in idr1-1 mutant stemmed from a range of physiological and morphological changes, including greater leaf potentials, increased proline contents, heightened leaf thickness, and upregulation of antioxidant-synthesizing and drought-induced genes, etc., under drought-stressed conditions. Especially, ROS production might be remarkably impaired, while ROS-scavenging ability appeared to be markedly enhanced as a result of significantly elevated expression of ROS-scavenging enzyme genes in idr1-1 mutant under drought-stressed conditions. Besides, idr1-1 mutant showed reduced expression of OsBRD1. Altogether, these results suggest that mutation of IDR1 leads to alterations of multiple layers of regulations, which ultimately confers obviously enhanced drought tolerance to the idr1-1 mutant.localized in nucleus and to plasma membrane or cell periphery. Further investigations indicated that the significantly increased drought tolerance in idr1-1 mutant stemmed from a range of physiological and morphological changes, including greater leaf potentials, increased proline contents, heightened leaf thickness, and upregulation of antioxidant-synthesizing and drought-induced genes, etc., under drought-stressed conditions. Especially, ROS production might be remarkably impaired, while ROS-scavenging ability appeared to be markedly enhanced as a result of significantly elevated expression of ROS-scavenging enzyme genes in idr1-1 mutant under drought-stressed conditions. Besides, idr1-1 mutant showed reduced expression of OsBRD1. Altogether, these results suggest that mutation of IDR1 leads to alterations of multiple layers of regulations, which ultimately confers obviously enhanced drought tolerance to the idr1-1 mutant.

overall_design:
The RNA-seq data of rice idr1-1 leaves under drought stress and non-drought stress
4 sets of data, each of them containing 3 biological replicates

first: Honggui

first: Xiaofeng

last: Zu

first: Huaqi

last: Wang

first: Yumei


title: CUT&RUN reveals unique positioning of pre-initiated RNA polymerase II in the steady state of transcription

city: Hong Kong

first: Michi

last: Miura

address: 21 Sassoon Road

country: Hong Kong

institute: The University of Hong Kong

summary: CUT&RUN is a powerful tool to study protein-DNA interactions in vivo. DNA fragments cleaved by the targeted micrococcal nuclease identify the footprints of DNA-binding proteins on the chromatin. We performed CUT&RUN on human lung carcinoma cell line A549 maintained in a multi-well cell culture plate to profile RNA polymerase II. Size-fractionation of aligned reads revealed that the collection of long (>270 bp) DNA fragments corresponded to the bimodal peak around the transcription start sites, as previously seen with chromatin immunoprecipitation. However, we found that shorter (<120 bp) fragments identify a well-defined peak localised at the transcription start sites. This distinct DNA footprint of shorter fragments, which constituted only about 5% of the total reads, indicates the unique positioning of the poised RNA polymerase II before transcription initiation, which has not been detected in the steady state of transcription by standard chromatin immunoprecipitation.

overall_design: CUT&RUN assays targeting RNA polymerase II.  Formaldehyde cell fixation was implemented in some of these assays.

first: Honglin

last: Chen


title: Endothelial GATA4 controls liver fibrosis and regeneration by preventing a pathogenic switch in angiocrine signaling [ChIP-seq]

first: Gergana

last: Dobreva

overall_design: 3 Samples

first: Julio

last: Cordero

first: Christian

last: Schmid


title: Trib1 promotes acute myeloid leukemia progression by modulating transcriptional programs of Hoxa9 [ChIP-seq]

first: Takuro

last: Nakamura

summary: The pseudokinase Trib1 functions as a myeloid oncogene that recruits E3 ubiquitin ligase COP1 to C/EBP, and interacts with MEK1 to enhance ERK phosphorylation. Close genetic interaction between Trib1 and Hoxa9 has been observed in myeloid leukemogenesis as Trib1 overexpression significantly accelerates Hoxa9-induced leukemia onset. Yet the mechanism how Trib1 functionally modulates Hoxa9 transcription activity is unclear. Here we provide evidence that Trib1 modulates Hoxa9-associated super-enhancers. ChIP-seq analyses identified increased Histone H3K27Ac signals at the super-enhancers such as Erg, Aff3, Spns2 and Coro2 loci, as well as increased mRNA expression of these genes. The modification of super-enhancer activities is mostly achieved by degradation of C/EBP by Trib1 and the MEK/ERK pathway only slightly contributes to the activities. Silencing of Erg abrogates the growth advantage acquired by Trib1 overexpression, indicating that Erg is a critical downstream target of the Trib1/Hoxa9 axis. Moreover, treatment of AML cells with the BRD4 inhibitor JQ1 showed growth inhibition in the Trib1/Erg-dependent manner. Upregulation of ERG by TRIB1 was also observed in human AML cell lines, suggesting that Trib1 is a potential target of Hoxa9-associated AML.

overall_design: Examination of binding sites of 2 transcription factors, Hoxa9 and C/EBPa, and Histone H3K27ac in a single cell line with 2 different conditions

first: Seiko

last: Yoshino


title: miRNA sequencing of extracellular vesicles derived from podocytes under high glucose stimulation.

city: Guangzhou

first: Ying

last: Huang

state: Guangdong

address: 106 Zhongshan 2 Road

department: Division of Nephrology

institute: Guangdong Provincial People's hospital

summary: Extracellular vesicles were isolated from the cell culture supernatants of podocytes under high glucose(HG), normal glucose(NG) and iso-osmolality stimulation (three replicates/group). miRNA sequencing was performed to identify differentially expressed miRNAs in podocyte-derived extracellular vesicles. After sequencing, A total of 1915 miRNAs were annotated from all samples . A comparison of the HG and NG groups showed that 11 miRNAs were differentially expressed (4 for up-regulated and 7 for down-regulated,|log2(fold change)| > 1, p-value < 0.05), while a comparison of the HG and iso-osmolality groups showed that 18 miRNAs were differentially expressed (1 for up-regulated and 17 for down-regulated, |log2(fold change)| > 1, p-value < 0.05). This study provides the results of miRNA alteration in podocytes extracellular vesicles under HG stimulation.

overall_design: Examination of miRNA expression profiles of podocyte-derived extracellular vesicles using high-throughput sequencing.

first: Xinling

last: Liang

first: Ruizhao

last: Li

first: Li

first: Yuanhan

first: Wei

last: Dong


title: Melatonin affects endothelial cells-derived exosomal miRNA expression

city: Shenyang

first: Xiangshan

last: Xu

state: Liaoning

address: Fourth Chongshan Road, Huanggu District

institute: fourth affiliated hospital of China Medical University

summary:
The proper response to shear stress involves multiple cell components of the vascular wall including endothelial cells (ECs), smooth muscle cells (SMCs), and the intercellular communications between them. Mounting evidence indicates that retinoid acid receptor-related orphan receptor-α (RORα) mediates many biological activities of melatonin including transactivation of transcriptional factors, and anti-inflammation. In this study we investigated the effect of melatonin on endothelial cells-derived exosomal miRNA expression.
We used microarrays to detail the global program of gene expression underlying melatonin treatment and identified distinct classes of dys-regulated genes during this process.

overall_design: Exosomes were isolated and purified from the ECs monoculture supernatants subjected to low shear stress together with or without melatonin treatment for 24 hours. Three replicated samples for each group were performed.

first: Yuanzhe

last: Jin


title: The wild rice locus CTS-12 mediates ABA-dependent stomatal opening modulation to limit water loss under severe chilling stress

city: Nanning

first: Jijing

last: Luo

state: Guangxi

address: 100 Daxue Rd.(East)

institute: Guangxi University

summary: To understand how CTS-12 the ABA-dependent multi-levels of regulation, the integration of transcriptomic and metabolomic profiling using the two-way orthogonal projections to latent structures (O2PLS) and OPLS discriminant analysis (OPLS-DA) modeling was performed to investigate the mechanisms underlying chilling tolerance. Our results revealed that metabolic flux shifts, including the activation of stachyose biosynthesis, amino acid metabolism pathways, phenylpropanoid/flavonoid biosynthesis, and ABA biosynthesis, and inhibition of glycolysis, occurred under chilling treatment, and in the recovery period, the differentially expressed genes/metabolites (DEGs/DEMs) that mapped to glutamate-related pathways, β-alanine biosynthesis and degradation, and serotonin biosynthesis pathways were differentiated between 9311 and DC90. Particularly, the differential alterations of the DEMs/DEGs, including galactinol, β-alanine, glutamate, naringenin, serotonin, abscisic acid (ABA), and LOC_Os03g44380 (OsNCED3), might be involved in the chilling stress phenotype variation of 9311 and DC90. The involvement of ABA pathway was validated by CRISPR/Cas9-edited of discriminatory DEGs OsNCED3 which impaired chilling tolerance of japonica rice. In addition, chilling tolerance of rice was associated with the balance of water uptake and loss that was modulated by stomatal movement under chilling stress. Therefore, we speculated that the CTS-12-mediated ABA signaling pathway leads to transcriptional regulation of chilling-responsive genes and, in turn, triggers metabolic shifts to coordinately regulate the stomatal movement of guard cells. The results of this study improve our understanding of the multilevel regulation of wild rice in response to chilling stress.

overall_design: Examination of transcriptomic profiling in 5 timepoints of chilling and recovery treatment in two rice materials, 9311 and DC90, respectively.

first: Weijian

last: Cen

first: Wenlong

last: Zhao


title: DOT1L-controlled cell-fate determination and transcription elongation are independent of H3K79 methylation

city: Chicago

first: Ali

last: Shilatifard

state: IL

address: 320 E Superior St

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University Feinberg School of Medicine

summary: Actively transcribed genes in mammals are decorated by H3K79 methylation, which is correlated with transcription levels and is catalyzed by the histone methyltransferase DOT1L. DOT1L is required for mammalian development, and the inhibition of its catalytic activity has been extensively studied for cancer therapy; however, the mechanisms underlying DOT1L’s functions in normal development and cancer pathogenesis remain elusive. To dissect the relationship between H3K79 methylation, cellular differentiation, and transcription regulation, we systematically examined the role of DOT1L and its catalytic activity in embryonic stem cells (ESCs). DOT1L is dispensable for ESC self-renewal but is required for establishing the proper expression signature of neural progenitor cells, while catalytic inactivation of DOT1L has a lesser effect. Furthermore, DOT1L loss, rather than its catalytic inactivation, causes defects in glial cell specification. Although DOT1L loss by itself has no major defect in transcription elongation, transcription elongation defects seen with the super elongation complex inhibitor KL-2 are exacerbated in DOT1L knockout cells, but not in catalytically dead DOT1L cells, revealing a role of DOT1L in promoting productive transcription elongation that is independent of H3K79 methylation. Taken together, our study reveals a catalytic-independent role of DOT1L in modulating cell-fate determination and in transcriptional elongation control.

overall_design: Studying the functions of methyltransferase DOT1L in neural differentiation and transcription elongation.

first: Kaixiang

last: Cao

first: Patrick

last: Ozark


title: Vascular Disease and Thrombosis in SARS-CoV-2 Infected Humans and Rhesus Macaques

city: Atlanta

first: Gregory

last: Tharp

state: GA

address: 954 Gatewood Dr

department: Developmental and Cognitive Neuroscience

institute: Yerkes National Primate Research Center

summary: The COVID-19 pandemic has led to extensive morbidity and mortality throughout the world. Clinical features that drive SARS-CoV-2 pathogenesis in humans include inflammation and thrombosis, but the mechanistic details that underlie these processes remain to be determined. In this study, we demonstrate endothelial disruption and vascular thrombosis in histopathologic sections of lungs from both humans and rhesus macaques infected with SARS-CoV-2. To define key molecular and cellular pathways associated with SARS-CoV-2 pathogenesis, we performed transcriptomic analyses of bronchoalveolar lavage (BAL) samples and peripheral blood, and proteomics analyses of serum from infected rhesus macaques. We observed upregulation of macrophage signatures, complement cascade pathways, platelet activation, and markers of thrombosis in BAL and peripheral blood as well as extensive macrophage infiltrates in lung. These observations coincided with robust induction of interferon and proinflammatory markers, including C-reactive protein, MX1, IL-6, IL-1, IL-8, TNFa and NF-κB as well as downstream signaling pathways. These findings suggest a model in which critical interactions between inflammatory and thrombosis pathways lead to SARS-CoV-2 induced vascular disease. Our findings also suggest potential novel therapeutic targets for COVID-19 disease.

overall_design: Nine adult rhesus macaques (6 to 12 years of age) with a total of 1.1 × 10^6 plaque-forming units (PFU) (Group Gr_I; N = 3), 1.1×10^5 PFU (Group Gr_II;N=3), or 1.1×10^4 PFU (Group Gr_III; N = 3) of SARS-CoV-2. PBMC were collected pre-infection, and 1, 2, 4, 7, 10 and 14 days post-infection. Bronchoalveolar lavage (BAL) was collected 1, 2, 4, 7, 10 and 14 days post-infection, and from 6 additional rhesus macques who were not infected.

first: Malika

last: Aid

first: Amanda

last: Martinot

first: Jacob

last: Estes

first: Dan

last: Barouch


title: Identification of fetal unmodified and 5-hydroxymethylated CGs in maternal cell-free DNA for non-invasive prenatal testing

city: Vilnius

first: Giedrius

last: Vilkaitis

address: Sauletekio av. 7

department: Department of Biological DNA Modification, Institute of Biotechnology

country: Lithuania

institute: Vilnius University

summary: Massively parallel sequencing of maternal cell-free DNA (cfDNA) is widely used to test fetal genetic abnormalities in non-invasive prenatal testing (NIPT). However, sequencing-based approaches are still of high cost. Building upon previous knowledge that placenta, the main source of fetal circulating DNA, is hypomethylated in comparison to maternal tissue counterparts of cfDNA, we propose that targeting either unmodified or 5-hydroxymethylated CG sites specifically enriches fetal genetic material and reduces numbers of required analytical sequencing reads thereby decreasing cost of a test.

overall_design: We employed covalent derivatization of CG sites combined with lower cost next generation sequencing, or quantitative PCR for analysis of fetal chorionic villi tissue samples, and cfDNA of pregnant and non-pregnant women.

first: Juozas

last: Gordevičius

first: Milda

last: Narmontė

first: Povilas

last: Gibas

first: Kotryna

last: Kvedaravičiūtė

first: Vita

last: Tomkutė

first: Priit

last: Paluoja

first: Kaarel

last: Krjutškov

first: Andres

last: Salumets

first: Edita

last: Kriukienė


title: PacBio long-read RNA-sequencing to identify differentially expressed genes and repetitive elements between Wild Type RBM4 KO HAP1 cell line samples

city: Bethesda

first: Stefan

last: Muljo

state: MD

address: 10 CENTER DR RM 11N306

institute: NIH/NIAID

summary: Since short reads from Illumina RNA-seq data are challenging to map to repetitive elements , we wanted to confirm the bulk RNA-seq findings using an orthogonal method, namely, using the long read technology of Pacific Biosciences (PacBio) full-length transcriptome sequencing. This dataset provided around 1.1 (WT) and 1.3 (RBM4 KO) million sequence reads of 2.6 kb average length mapping to the human genome.

overall_design: Single replicate provided, each, for Wild Type and RBM4 KO (the 1B6 clone as prefixed in the bulk experiment)

first: Amir

last: Foroushani

first: Bryan

last: Chim

first: Kent

last: Barbian

first: Craig

last: Martens


title: Investigation of impact of DNA synthesis during Break-induced replication on transcription

city: Iowa City

first: Anna

last: Malkova

state: Iowa

address: 29 E. Jefferson St.

department: iBio

institute: University of Iowa

summary: We discovered that during rPolII is enriched in the TES region due colision between transcription and BIR synthesis. This enrichment is specific to genes that are located in head-on orientation to BIR.

overall_design: rPolII distribution on genes located in BIR template in the time frame of BIR.

first: Liping

last: Liu

first: Luyang

last: Sun

first: Zhenxin

last: Yan

first: Grzegorz

last: Ira

first: Weiwei

last: Dang

first: Daniel

last: Kraushaar


title: SWI/SNF inactivation in the endometrial epithelium leads to loss of epithelial integrity [Mouse EPCAM]

city: Grand Rapids

first: Ronald

middle: L.

last: Chandler

state: MI

department: Obstetrics, Gynecology and Reproductive Biology

institute: Michigan State University

summary: Although ARID1A mutations are a hallmark feature, mutations in other SWI/SNF (SWItch/Sucrose Non-Fermentable) chromatin remodeling subunits are also observed in endometrial neoplasms. Here, we interrogated the roles of Brahma/SWI2-related gene 1 (BRG1, SMARCA4), the SWI/SNF catalytic subunit, in the endometrial epithelium. BRG1 loss affects more than one-third of all active genes and highly overlaps with the ARID1A gene regulatory network. Chromatin immunoprecipitation studies revealed widespread subunit-specific differences in transcriptional regulation, as BRG1 promoter interactions are associated with gene activation, while ARID1A binding is associated with gene repression. However, we identified a physiologically relevant subset of BRG1 and ARID1A co-regulated epithelial identity genes. Mice were genetically engineered to inactivate BRG1 specifically in the endometrial epithelium. Endometrial glands were observed embedded in uterine myometrium, indicating adenomyosis-like phenotypes. Molecular similarities were observed between BRG1 and ARID1A mutant endometrial cells in vivo, including loss of epithelial cell adhesion and junction genes. Collectively, these studies illustrate overlapping contributions of multiple SWI/SNF subunit mutations in the translocation of endometrium to distal sites, with loss of cell integrity being a common feature in SWI/SNF mutant endometrial epithelia.

overall_design: EPCAM+ endometrial epithelial cells from LtfCre^0/+; Brg1^fl/fl CD-1 mice were purified and assayed via RNA-seq.

first: Jake

last: Reske

first: Mike

middle: R

last: Wilson

middle: L


title: Expression data from different expressed genes(DEGs) in human esophageal squamous cell carcinoma(ESCC)  EC109 cells transfected with empty lentiviral vector(Lv-NC) or lentiviral vector carrying MEG3(Lv-MEG3)

city: Shantou

first: Li

middle: Ming

last: kai

address: Dong Xia

institute: Shantou University

summary:
Maternally expressed gene 3 (MEG3), a member of lncRNAs, has tumor-suppressor properties and is downregulated in the majority of malignancies. MEG3 was found to suppress some sorts of tumors through regulating epithelial mesenchymal transformation,inducing cell cycle arrest,triggering apoptosis and so on.
We used microarrays to investigate the possible regulatory mechanism of MEG3 in ESCC cell line.

overall_design: MEG3 was overexpressed with lentivirus.Hence,EC109 cells harvested from three dishes were divided into two groups: Lv-NC(control group, Control) and Lv-MEG3 (overexpressing MEG3 group, MEG3).A DNA microarray was applied to analyze DEGs,accompanied by bioinformatics analyses including Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway.

first: Ling-Fei

last: Wu

first: Ming-Kai

last: Li


title: Resolving transcriptional states and predicting lineages in the annelid Capitella teleta using single-cell RNAseq

city: Worcester

first: Néva

last: Meyer

state: MA

address: 950 Main Street

department: Biology

institute: Clark University

summary: Here we use a whole-organism, single-cell transcriptomic approach to map larval cell types in the annelid Capitella teleta at 24- and 48-hours post gastrulation (stages 4 and 5). We identified eight unique cell clusters (undifferentiated precursors, ectoderm, muscle, ciliary-band, gut, neurons, neurosecretory cells and protonephridia), thus helping to identify previously uncharacterized molecular signatures such as novel myogenic and neurosecretory cell markers. Analysis of coregulatory programs in individual clusters revealed gene interactions that can be used for comparisons of cell types across taxa.  We examined the neural and neurosecretory clusters more deeply and characterized a differentiation trajectory starting from dividing precursors to neurons using Monocle3 and velocyto. Pseudotime analysis along this trajectory identified temporally-distinct cell states undergoing progressive gene expression changes over time. Our data revealed two potentially distinct neural differentiation trajectories including an early trajectory for brain neurosecretory cells. This work provides a valuable resource for future functional investigations to better understanding neurogenesis and the transitions from neural precursors to neurons in an annelid.

overall_design: Whole organism single-cell RNAseq datasets using the 10X genomics platform on two C. teleta larval stages 24- (stage 4) and 48-hours post gastrulation (stage 5)

first: Abhinav

last: Sur

middle: P


title: SRSF6 -regulated alternative splicing of genes involved in DNA repair is associated with prostate cancer progression

city: Wuhan

first: Dong

last: Chen

state: Hubei

address: 388 GaoXin 2nd Road, East Lake Hi-Tech Development Zone

department: Center for Genome Analysis

institute: ABLife, Inc.

summary: Alternative splicing (AS) occurs in nearly all human genes and abnormal AS has a close connection with cancer. SRSF6, a canonical member of the serine/arginine-rich (SR) protein family, has been characterized as an important regulator of AS. However, the role of SRSF6 in regulating AS in prostate cancer remains unclear. In this study we analyzed SRSF6 expression levels in 550 prostate cancer tissue samples available in The Cancer Genome Atlas database and found that expression of SRSF6 was upregulated in all prostate cancer stages we examined. To investigate the biological consequences of SRSF6 overexpression, we constructed an SRSF6-overexpressing cell model and utilized RNA-seq to explore transcriptional changes. We also selected two groups of prostate cancer tumor samples in which SRSF6 was differentially expressed to analyze potential SRSF6-regulated AS. We hypothesized that overexpression of SRSF6 in cancer cells would induce large-scale changes in transcriptional expression levels and AS. We found that the pattern of SRSF6-regulated AS in cancer cells was similar to that observed in clinical samples and was enriched in pathways like DNA repair, double-strand break repair via homologous recombination and others. We also validated SRSF6-regulated AS events using RT-qPCR. Our findings highlighted how SRSF6 could regulate DNA repair-related pathways via regulation of AS to influence cancer progression in both cancer cells and prostate cancer tumor samples. These results greatly expand our current understanding of the mechanisms underlying SRSF6-mediated gene regulation and suggests the potential use of SRSF6 as a cancer therapeutic target.

overall_design: transcrptional analysis of SRSF6 overexpression and control in Hela cells

first: Xiao

last: Yang

first: Mengdi

last: Wang


title: STAU1 selectively regulates the expression of inflammatory and immune response genes and alternative splicing of nerve growth factor receptor signaling genes

summary: To study the potential targets and functions of STAU1 which might be related to neuropathic pain, we obtained STAU1-regulated transcriptome in HeLa cells. High-throughput  RNA sequencing(RNA-seq) were performed for HeLa cells and control cells to define the comprehensive gene expression profiles and identify the genome-wide alternative splicing events regulated by STAU1.

overall_design: transcriptional analysis of STAU1 overexpression and control in Hela cells

first: Feng

first: Yu

last: Zhang


title: Differential Peptide Subtraction (DPS):  mass spectrometry for human cancer virus discovery

city: Frankfurt

first: Tuna

last: Toptan

state: Hessen

address: Institute of Medical Virology, Paul-Ehrlich-Straße 40

institute: University Hospital Frankfurt

summary: The challenge of discovering a completely new human tumor virus of unknown phylogeny or sequence depends on detecting viral molecules and differentiating them from host-derived molecules in the virus-associated neoplasm. We developed differential peptide subtraction (DPS) using differential mass-spectrometry (dMS) followed by targeted analysis to facilitate this discovery. To trace the non-human dMS identified peptides back to its genetic origin by next generation sequencing (NGS) cDNA libraries were generated using degenerate oligos corresponding to the identified peptides. In a pilot study DPS identified both viral and human biomarkers. Based on the identified peptides that are differentially expressed in the virus positive tumor sample, degenerate oligos were designed. Degenerate oligos or LNA modified degenerate oligos or a modified oligo(dT) SMART primer were used to facilitate reverse transcription from viral or viral and host RNA. NGS analysis revealed seven times more MCV reads in degenerate oligo primed RNAseq samples compared to polyA-based sequencing reads.

overall_design: Ribodepleted RNA from Merkel cell polyomavirus (MCV) positive Merkel cell carcinoma (MCC) tissue was used for RNA sequencing. Three different cDNA libraries were prepared using the same tissue RNA. Sample 1 (MCC-deg): a pool of degenerate oligos used to prime the cDNA synthesis. Sample 2 (MCC-LNAdeg): a pool of LNA modified degenerate oligos were used for cDNA synthesis. Sample 3 (MCC-polyA): modified oligo dT primer was used for cDNA synthesis prior to library generation. RNAseq data was analyzed to detect virus specific reads and counts were compared to evaluate the efficiency of library genertion.

first: Patrick

last: Moore

first: Yuan

last: Chang

first: Nathan

last: Yates


title: Epigenetic therapy induces transcription of Inverted SINEs and ADAR1 dependency

city: Toronto

first: Sajid

last: Marhon

state: ON

address: 101 College Street

institute: Princess Margaret Cancer Centre

summary: In this study we show that intronic and intergenic SINE elements, specifically inverted repeats (IR) Alus, are the major source of drug-induced immunogenic dsRNA. These IR-Alus are frequently located downstream of ‘orphan’ CpG Islands (CGIs). In mammals, the enzyme Adenosine Deaminases Acting on RNA (ADAR1) targets and destabilizes IR-Alu dsRNA, which prevents activation of MDA5. We found that ADAR1 establishes a negative feedback loop, restricting the viral mimicry response to epigenetic therapy. Depletion of ADAR1 in patient-derived cancer cells potentiates the efficacy of epigenetic therapy, restraining tumour growth and reducing cancer initiation. Thus, epigenetic therapies trigger viral mimicry by inducing a subset of IR-Alus, leading to an ADAR1 dependency. Our findings suggest that combining epigenetic therapies with ADAR1 inhibitors represents a promising new strategy for cancer treatment.

overall_design: Identifying the source of immunogenic dsRNAs that can be used as a biomarker for response to epigenetic therapies in cancer cells and finding ADAR1 as a new target that can be used in combination with epigenetic therapies for cancer treatment

first: Parinaz

last: Mehdipour

first: Ilias

last: Ettayebi

first: Daniel

last: De Carvalho


title: Epigenome wide association study of post-mortem Alzheimer’s Disease samples

city: Titusville

first: Qingqin

state: NJ

address: 1125 Trenton-Harbourton Road

institute: Janssen R&D, LLC

summary:
Samples from 72 AD patients and 62 age-matched cognitively normal controls were assayed using Illumina© Infinium MethylationEPIC BeadChip. We performed an epigenome-wide association study (EWAS) to evaluate the epigenetic differences using post-mortem superior temporal gyrus (STG) and inferior frontal gyrus (IFG) samples.
We performed an epigenome-wide association study (EWAS) to evaluate the epigenetic differences using post-mortem superior temporal gyrus (STG) and inferior frontal gyrus (IFG) samples.


title: Fasting does not promote a metabolic memory effect  in mouse liver

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

country: Netherlands

institute: Wageningen University

summary: Background: Tissues may respond differently to a particular stimulus if they have been previously exposed to that same stimulus. Here we tested the hypothesis that a strong metabolic stimulus may elicit a memory effect in the liver. To that end, we examined whether prior fasting, which profoundly impacts hepatic nutrient metabolism, may influence the metabolic response to a subsequent fast. Methods: 24 mice were exposed to two 16h fasts over a 8 week period, each time being allowed to return to their normal growth trajectory, while another group of 24 mice was fed ad libitum throughout. Of each group, half of the mice were euthanized after a 16h fast, whereas the other half was euthanized in the ab libitum fed state. Results: An acute fast significantly increased plasma NEFA, glycerol, β-hydroxybutyrate and liver triglycerides, and decreased plasma glucose, triglycerides, and liver glycogen levels. An acute fast also markedly changed the liver transcriptome, upregulating genes involved in fatty acid catabolism and downregulating genes involved in cholesterol synthesis, and majorly impacted the liver metabolome, reducing the levels of numerous amino acids, glycolysis and TCA cycle intermediates, and nucleotides. However, none of the these changes were significantly influenced by prior fasting. The limited number of genes that were significantly altered by prior fasting are likely false positives. Finally, no significant effect was observed of prior fasting on glucose tolerance. Conclusion: We conclude that previous exposure to fasting in mice does not influence the metabolic response to a subsequent fast, thus arguing against the concept of metabolic memory in the hepatic response to fasting.

overall_design: Microarray analysis was performed on murine liver tissue after a 16 hour fast or in the fed state

first: Merel

last: Defour

middle: J

first: Michel

last: van Weeghel

first: Sander

last: Kersten


title: Next Generation Sequencing of Ezh2-cKO and Utx cKO transcriptomes of exostoses in mice

city: Xi'an

address: Changle west Road

department: Department of Pathology

institute: Fourth Military Medical University

summary: Ezh2-cKO or Utx-cKO in chondrocytes initiate  metachondromatosis which containing both exostoses and enchondromas. To figure out whether the exostoses of Ezh2-cKO and Utx-cKO were the similar tumore, we compared the transcriptional profiles using RNA-sequencing of exostoses from Ezh2-cKO and Utx-cKO mice.

overall_design:
Examination of differential genes expression in 2 genotipic exostoses cells
Exostoses in Ezh2-cKO or Utx-cKO-M mice at the 4 weeks after tumors were visible and the wild type articular cartilage from the same aged control mice were collected. RNA was extracted, purified, and checked for integrity using an Agilent Bioanalyzer 2100 (Agilent Technologies). Libraries were generated for sequencing using the TruSeq RNA sample preparation kit (Illumina). Libraries were sequenced using an Illumina HiSEq. 2500 sequencer. All of the above processes were performed at Shanghai Biotechnology Corporation, Shanghai, China


title: TBK1-METTL3 axis facilitates antiviral immunity [RNA-seq: Mock or VSV treated WT and Mettl3 crispr iBMDM]

status: Public on Oct 19 2020

city: beijing

first: fuping

last: you

address: xueyuan street

institute: Peking University Health Science Center

summary: mRNA m6A modification is involved in regulation of immune system. However, its function in antiviral immunity is controversial, and how immune responses regulate m6A modification is unknown. We here found TBK1, a key kinase of antiviral pathways, phosphorylated the core m6A methyltransferase METTL3 at Serine 67. The phosphorylated METTL3 interacted with translational complex and enhanced proteins translation, including IRF3, and facilitated antiviral responses. TBK1 also promoted METTL3 activation and m6A modification, which is required for stabilizing IRF3 mRNA. Type I IFN induction was severely impaired in METTL3 deficient cells. Mettl3flfl-lyz2-Cre mice were significantly more susceptible to IAV-induced lethality than control mice. Consistently, Ythdf1—/— mice cannot control viral infection and showed higher mortality than control mice due to decreased IRF3 expression. Together, we demonstrated that innate signals activated METTL3 via TBK1, and METTL3 and m6A modification secured antiviral immunity by promoting mRNA stability and protein translation.

overall_design: mRNA profiles of wild type (WT)  and Mettl3 knockdown(KD) immortalized bone marrow-derived macrophage (iBMDM) treated with mock or vesicular stomatitis virus (VSV) for 9 hours

first: Jingxuan

first: Xuemei

last: Wei

first: Xiao

first: Fuping

last: You


title: Transcriptome profiling of Chlorella sorokiniana under normal and stress condition

city: Serdang

first: Siti Nor Ani

last: Azaman

state: Selangor

address: Jalan UPM

country: Malaysia

institute: Universiti Putra Malaysia

summary:
Purpose: The purpose of this study are to identify the genes involved in regulating the production of metabolites in Chlorella sorokiniana under stress-induced condition through RNA-sequencing technique.
Methods:Transcriptome profile from normal and stress sample of C. sorokiniana were generated, in triplicate, using Illumina Hiseq2000.  The sequence reads that passed quality filters were de-novo assembled using Trinity doftware, followed by analysis using  Burrows–Wheeler Aligner (BWA) and DESeq2. Lastly, RT-qPCR was used to validate the expression level of genes.
Results: A total of 200 million reads was obtained from both normal and stress induced samples of C. sorokiniana. The assembly program produced 18,310 assembled transcripts with a length ranging from 165 to 16,695 bp. Differential expression analysis revealed  933 transcripts were significantly differentially expressed between normal and stress with 431 were up-regulated and 502 were down-regulated. Pathway analysis revealed that the differentially expressed genes were associated with a variety of KEGG pathways.

overall_design: Transcriptome profile of C. sorokiniana under normal and stress conditions were generated in triplicates by deep sequencing using NEBNext Ultra Directional RNA for Illumina.

first: Siti

middle: N

first: Swee

last: Yeap

first: Sheau

last: Tan

first: Darren

middle: C

last: Wong

first: Dilan

last: Satharasinghe

first: Woan

last: Liew

first: Choong

last: C Wean

first: David

last: Hornby

first: Qaiser

middle: I

last: Sheikh

first: Fatimah

last: Mohd Yusoff

first: Norio

last: Nagao

first: Hyunh

last: Ky


title: RNAseq experiment to identify differentially expressed genes by AP2 induction in the Shoot Apical Meristem (SAM) of Arabidopsis

city: Valencia

first: Vicente

last: Balanzà

state: Valencia

address: Campus de la Univ. Politécnica de Valencia. CPI Edif. 8E. Av de los Naranjos s/n

institute: Instututo de Biologia Molecular y Celular de Plantas (CSIC-UPV)

summary: Trough a two components inducible system, we expressed ectopically in the SAM a resistant version of AP2 to the regulation of miR172 (AP2m3).  The differentially expressed genes by AP2 induction allows the identification of differents groups of genes that could mediate AP2 functions in the SAM.

overall_design: Two weeks after bolting, SAM of Arabidopsis plants were treated with dexamethasone, that allows the AP2m3 expression, or mock. 6-8 hours after treatment SAM were collected removing all visible flower primordia. RNA was extracted and sequenced on a MiSeq (Illumina),

first: Cristina

last: Ferrándiz


title: DNA methylation analysis of methamphetamine abusers with different addiction susceptibility

city: Changsha

first: Wei

last: Hao

state: Hunan

address: 139 Renmin Middle Road, Furong district

institute: The second xiangya hospital of Central South University

summary: Genome wide DNA methylation profiling of normal and methamphetamine (MA) abusers with different addiction susceptibility. The Illumina Infinium HumanMethylation450 Beadchip was used to obtain DNA methylation profiles in peripheral blood lymphocytes (PBLs). Samples included 8 health controls, 8 high MA addiction susceptibility (HMAS) abusers, and 8 high MA addiction susceptibility (LMAS) abusers.

overall_design: Bisulphite converted DNA from the 24 samples were hybridised to the Illumina Infinium HumanMethylation450 Beadchip

first: Liang

last: Liu


title: SWI/SNF inactivation in the endometrial epithelium leads to loss of epithelial integrity [12Z RNA-seq]

first: Ronald

last: Chandler

overall_design: Human 12Z endometriotic epithelial cells were transfected with either siBRG1 or non-targeting siRNA control and assayed 72 hours post-transfection via RNA-seq.

first: Jake

last: Reske

first: Mike

last: Wilson


title: Mitochondrial Potentiation Ameliorates Age Related Heterogeneity in Hematopoietic Stem Cell Function

city: Lund

first: Shamit

last: Soneji

address: Solvegatan 19

institute: BMC

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

overall_design: Refer to individual Series


title: Mitochondrial Potentiation Ameliorates Age Related Heterogeneity in Hematopoietic Stem Cell Function [HSCs treated]

summary:
introduction: Ageing is associated with reduced fitness and increased myeloid bias of the hematopoietic stem cell (HSC) compartment, causing increased risk of immune compromise, anemia and malignancy
results: We show that mitochondrial membrane potential (MMP) can be used to prospectively isolate chronologically old HSCs with transcriptional features and functional attributes characteristic of young HSCs, including a high rate of transcription and balanced lineage-affiliated programmes. Strikingly, MMP is a stronger determinant of the quantitative and qualitative transcriptional state of HSCs than chronological age and transcriptional consequences of manipulation of MMP in HSCs within their native niche suggest a causal relationship.  Accordingly, we show that pharmacological enhancement of MMP in old HSCs in vivo increases engraftment potential upon transplantation and reverses myeloid-biased peripheral blood output at steady state.
conclusion: Our results demonstrate that MMP is a source of heterogeneity in old HSCs and its pharmacological manipulation can alter transcriptional programs with beneficial consequences for function.

overall_design: RNA seq of Old HSCs compared to Young HSCs as well as Old HSCs from Mito-Q treated mice and young HSCs from CCCP treated mice

first: Els

last: Mansell

first: Chela

last: James

first: Elitza

last: Deltcheva

first: John

last: Brown

first: T

last: Enver