title: Single-cell RNA-sequencing of tdTomato-labeled c-Kit lineage cells in aortic allografts of mouse models

first: Jiacheng

last: Deng

summary: We reported the transcriptional profiling of tdTomato-labeled c-Kit lineage cells in a mouse model of transplant arteriosclerosis by single-cell RNA sequencing.

overall_design: This entry contains 10X Single-cell RNA-seq data of  tdTomato-labeled c-Kit lineage cells of 2-week and 4-week aortic allografts.

first: Wenduo

last: Gu

first: Zhichao

last: Ni

first: Qingbo

last: Xu


title: An essential role for MEF2C in the cortical response to loss of sleep

status: Public on Sep 08 2020

city: Dallas

first: Genevieve

last: Konopka

state: TX

address: 5323 Harry Hines Blvd.

department: Neuroscience

country: USA

institute: UT Southwestern Medical Center

type: Expression profiling by high throughput sequencing

summary: Insight on molecular and cellular responses to sleep loss provides a clearer understanding of sleep function.

overall_design: Neurobiological changes in the mouse frontal cortex in response to sleep loss, involving transcriptomic architecture, regulation of synaptic strength in excitatory cortical neurons and SWS-SWA, correlated with sleep need.

first: T

middle: E

last: Bjorness

first: A

last: Kulkarni

first: V

last: Rybalchenko

last: Suzuki

first: C

last: Bridges

middle: W

last: Cowan

first: J

middle: S

last: Takahashi

first: G

first: R

last: Greene


title: Local translation highlights novel properties of perisynaptic astrocytic processes, and is modulated by behavior involving the dorsal hippocampus

city: PARIS

first: Stephane

last: Le Crom

address: 46 rue Ulm

department: Genomic Core Facility

country: France

institute: IBENS

summary: Local translation is a conserved molecular mechanism. It allows a cell with a complex shape to bypass somatic protein synthesis and transport, and thus to respond quickly to a local stimulus. Local translation also contributes to the establishment of molecular and functional polarity. In the brain, it has been extensively studied in neurons and has also been described more recently in astrocytes - a type of glial cell with specialized extensions that contact vessels and synapses. Here, we studied perisynaptic astrocytic processes (PAPs)  in the dorsal hippocampus and showed that they contain RNAs, ribosomes, the endoplasmic reticulum-Golgi intermediate compartment, particles of the Golgi apparatus, and protein synthesis events. We used our recently refined polysome immunoprecipitation technique to characterize the pool of polysomal mRNAs in PAPs (which we refer to as the “PAPome”) from the dorsal hippocampus and compared it with the polysomal mRNAs found in the astrocyte as a whole. The polysomal transcripts that were enriched in the PAPome encoded mostly cytoplasmic proteins and defined an unexpected molecular repertoire with the most enriched transcripts coding for proteins involved in iron homeostasis, translation, cell cycle and cytoskeleton. Interestingly, among them Erz, Fth1, and Rplp1 were  enriched in PAPs compared to perivascular astrocytic processes indicating that local translation differ at these two interfaces and may sustain distinct molecular properties. The PAP-enriched transcripts Flt1, Fth1, Ccnd2, Mdm2, Gnb2l1 and Eef1a1 code for proteins involved in memory and learning mechanisms. We therefore studied their local translation in the context of fear conditioning (i.e. behavior involving the dorsal hippocampus). We observed changes in the density and/or distribution of these mRNAs in astrocytes processes as well as a drop of their translation specifically in PAPs. Our results highlight unexpected molecular properties of hippocampal PAPs and suggest for the first time that local translation in this perisynaptic compartment is linked to fear-related memory.

overall_design: Perisynaptic Astrocytic Processes (PAPs) versus full astrocytes polysomal transcriptome from dorsal hippocampus in Aldh1l1:L10a-eGFP mice

first: Noémie

last: Mazaré

first: Marc

last: Oudart

first: Julien

last: Moulard

first: Giselle

last: Cheung

first: Romain

last: Tortuyaux

first: Philippe

last: Mailly

first: David

last: Mazaud

first: Alexis

last: Bemelmans

first: Anne-Cécile

last: Boulay

first: Corinne

last: Blugeon

first: Laurent

last: Jourdren

first: Stéphane

first: Nathalie

last: Rouach

first: Martine

last: Cohen-Salmon


title: Single-cell RNA-sequencing of aortic allografts in mouse models

city: London

first: Jiacheng

last: Deng

state: Not Applicable

address: James Black Centre, 125 Coldharbour Lane

country: United Kingdom

institute: King's College London

summary: We reported the transcriptional profiling of cells in a mouse model of transplant arteriosclerosis by single-cell RNA sequencing.

overall_design: This entry contains 10X Single-cell RNA-seq data of cells of 2-week and 4-week aortic allografts.

first: Jingjing

last: Cai

first: Wenduo

last: Gu

first: Qingbo

last: Xu


title: Pygopus2-Histone interaction is important for de-differentiation and breast cancer progression [ATAC-Seq]

city: Basel

first: Ravi Kiran Reddy

last: KALATHUR

state: Basel

address: Mattenstrasse 28

department: Departement Biomedizin

country: Switzerland

institute: University of Basel

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Pygopus2 (Pygo2), a co-activator of Wnt/ β-catenin signaling can also bind H3K4me2/3 and participate in chromatin reading and writing. To investigate the functional relevance of histone binding activity of Pygo2 in malignant progression of breast cancer, we generated a knockin mouse model wherein binding of Pygo2 to H3K4me2/3 was rendered ineffective. Loss of Pygo2-histone binding resulted in smaller, differentiated and less metastatic tumors, at least in part due to a decrease in Wnt/ b-catenin signaling. RNA and ATAC-Seq analyses of tumor-derived cell lines revealed a downregulation of TGFb signaling and an upregulation of differentiation pathways like PDGFR. This also correlated with an increase in luminal cell fate which could be rescued by inhibition of the PDGFR pathway. Mechanistically, we find that Pygo2-histone interaction is important for expression of mir-29 family members to potentiate Wnt/ b-catenin signaling by suppressing Wnt signaling antagonists expression in addition to suppression of PDGFR expression. Collectively, our results demonstrate that histone binding function of Pygo2 is important for driving de-differentiation and malignancy of breast tumors and loss of this binding activates differentiation pathways that lead to decreased tumor growth and metastases. Ablating the Pygo2-H3K4me3 interaction hence might serve as an attractive therapeutic target for breast cancer.

overall_design: ATAC-Seq of Pygo2+/+ (WT) and Pygo2 AE/AE mutant cell lines on treatment with TGFb or Wnt3a was performed in biological duplicates

first: Meera

last: Saxena

first: Ravi

middle: K

last: Kalathur

first: Gerhard

last: Christofori


title: Direct Formalin Fixation Induces Widespread Transcriptomic Effects in Archival Tissue Samples

city: RTP

first: Susan

last: Hester

state: NC

address: 109 TW Alexander Dr

institute: US EPA

summary: Archival formalin-fixed paraffin-embedded (FFPE) tissue samples hold a wealth of transcriptomic information; however, little is known about potential artifacts. Previously, we identified a consistent shift in global RNA-sequencing profiles between matching frozen and FFPE samples. We hypothesized that this shift was from fixing fresh tissue in formalin. To test this idea, RNA-sequencing was performed on liver samples collected from male mice treated with 600 ppm of a reference chemical (phenobarbital, 600 ppm phenobarbital) or vehicle control for 7 days. Samples were divided into: (1) fresh-frozen (FR); (2) directly fixed in 10% buffered formalin for 18 hours followed by paraffin embedding (FFPE); (3) frozen then fixed as FFPE (FR>FFPE); or (4) frozen then fixed in 70% ethanol followed by paraffin embedding (FR>OH)  (n=6/group/condition). Direct fixation resulted in 2946 differentially expressed genes (DEGs), 98% of which were down-regulated. Freezing prior to fixation resulted in ≥95% fewer DEGs vs. FR, indicating that most formalin-derived transcriptional effects occurred with fixation. This was supported by follow-up studies, which identified consistent enrichment in oxidative stress, mitochondrial dysfunction, and transcription elongation pathways with formalin fixation. Notably, formalin fixation in the parent study did not significantly impact chemical response profiles, which were consistent with CAR/PXR activation and 600 ppm phenobarbital exposure. Our results demonstrate distinct transcriptional effects of formalin fixation that could impact gene expression studies using FFPE samples.

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus

first: Leah

last: Wehmas

middle: D

first: Charles

last: Wood


title: Comparison between ethylene signaling regulated transcriptome with glucose-TOR  or glucose-HXK1 regulated  transcriptome

city: Shanghai

first: Renyi

last: Liu

state: Shanghai

address: 3888 Chenhua Road

department: Bioinformatics

country: China

institute: Shanghai Center for Plant Stress Biology

summary: There are two principal mechanisms underlie sugar perception and signal transduction in biological systems. Direct sensing and signaling is triggered by sugar direct binding sensors such as Hexokiase 1 (HXK1) with a broad range of affinity and specificity, and indirect sensing signaling via a variety of energy and metabolite sensor, such as Target of Rapamycin (TOR). To investigate the intertwined interaction between glucose and plant hormones ethylene, the transcriptome regulated by ethylene, glucose-TOR, and glucose-HXK1 were analyzed and compared. Our results unexpectedly revealed a deep antagonistic interaction between glucose-TOR-energy signaling and the plant stress hormone ethylene.

overall_design: For ethylene related mRNA-seq, 4-day-old Col-0 seedlings were treated with 10 μM ACC for 2 hours and 4 hours. Non-treated seedlings were used as a control. Three biological replicates were set for each treatment, and one technical replicate was set for each biological replicate. For glucose-TOR related mRNA-seq, Wild-type (Col-0) and inducible tor mutant were grown in liquid 1/2 MS medium without glucose for 4 days, and then were treated with or without 15 mM glucose for 2 hours. Three biological replicates were set for each treatment, and one technical replicate was set for each biological replicate. For glucose-HXK1 related mRNA-seq, Wild-type (Ler-0) and the gin2 mutant seedlings were grown in liquid 1/2 MS medium without glucose for 4 days, and then were treated with or without 15 mM glucose for 2 hours. Three biological replicates were set for each treatment, and one technical replicate was set for each biological replicate.

first: Liwen

last: Fu

first: Yanlin

first: Guochen

last: Qin

first: Ping

last: Wu

first: Shuhui

last: Yang

first: Yi-min

last: She

first: Sang-Dong

last: Yoo

first: Yan

last: Xiong


title: Pygopus2-Histone interaction is important for de-differentiation and breast cancer progression

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

overall_design: Refer to individual Series


title: Identification of a novel LysR-type transcriptional regulator in Staphylococcus aureus that is crucial for secondary tissue colonization during metastatic bloodstream infection

city: Wuerzburg

first: Martin

last: Fraunholz

address: Theodor-Boveri-Weg

department: Chair of Microbiology

country: Germany

institute: University of Wuerzburg

summary:
Purpose: The goal of this study was to identify Staphylococcus aureus genes which are important during establishment of metastatic infections in bloodstream infections.
Methods: Pooled mariner transposon mutant libraries were generated as previously described (PMID: 27185949). The libraries were sequenced on the Illumina HiSeq 2500 platform with the transposon-specific oligonucleotide primer Himar1-Seq.  Illumina adapter sequences were removed via cutadapt version 1.2.1. The reads were filtered for size (>16 bp) and to contain the signature transposon inverted repeat. Reads were mapped to the Staphylococcus aureus 6850 genome (RefSeq accession NC_022222.1) by Bowtie2 v2.1.0.  Identification of depleted and enriched mutants was performed via DESeq2 version 1.6.2.
Results: S. aureus genes were identified which were upregulated or downregulated upon iduced expression of the LysR-type transcriptional regulator RSAU_000852 in Staphylococcus aureus 6850.
Conclusions: Our study identifies a gene for a LysR-type transcription regulator, which is involved in metabolic adaptation of the pathogen  during colonization of secondary infection sites in a bloodstream infection model.

overall_design: S. aureus RNAseq was performed on wild-type S. aureus 6850, an RSAU_000852 deletion mutant, and a strain carrying anhydrous tetracycline-induced RSAU_000852 ORF.  Experiments were performed in triplicates.

first: Maximilian

last: Klepsch


title: A novel antiviral lncRNA, EDAL, shields a T309 O-GlcNAcylation site to promote EZH2 lysosomal degradation

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.


title: Lipid stores and lipid metabolism associated gene expression in porcine and bovine parthenogenetic embryos revealed by immunostaining and RNA-seq [Ss]

status: Public on Sep 07 2020

city: Poznań

first: Arkadiusz

last: Kajdasz

state: wielkopolskie

address: ul. Z. Noskowskiego 12/14

country: Poland

institute: Institute of Bioorganic Chemistry PAS

summary: Compared to other mammalian species, porcine oocytes and embryos are characterized by large amounts of lipids stored mainly in the form of droplets in the cytoplasm. The amount and the morphology of LD change throughout the preimplantation development however, relatively little is known about expression of genes involved in lipid metabolism of early embryos. We compared porcine and bovine blastocyst stage embryos as well as dissected inner cell mass (ICM) and trophoblast (TE) cell populations with regard to lipid droplet storage and expression of genes functionally annotated to selected lipid Gene Ontology terms using RNA-seq. Comparing the number and the volume occupied by LD between bovine and porcine blastocysts, we have found significant differences both at the level of single embryo and a single blastomere. Aside from different lipid content we found that embryos regulate the lipid metabolism differentially at the gene expression level. Out of 125 genes, we have found 73 to be differentially expressed between entire porcine and bovine blastocyst, and 36 and 51 to be divergent between ICM and TE cell lines. We noticed significant involvement of cholesterol and ganglioside metabolism in preimplantation embryos as well as possible shift towards glucose rather than pyruvate dependence in bovine embryos. A number of genes like DGAT1, CD36 or NR1H3 may serve as lipid associated markers indicating distinct regulatory mechanisms while upregulated PLIN2, APOA1, SOAT1 indicate significant function during blastocyst formation and cell differentiation in both models.

overall_design: 4  blastocyst samples, 3 inner cell mass (ICM) samples, 3 trophectoderm (TE) samples

middle: P

first: Ewelina

last: Warzych

first: Natalia

last: Derebecka

first: Zofia

last: Madeja

first: Dorota

last: Lechniak

first: Joanna

last: Wesoły

first: Piotr

last: Pawlak


title: Lipid stores and lipid metabolism associated gene expression in porcine and bovine parthenogenetic embryos revealed by immunostaining and RNA-seq

sample_organism:
Sus scrofa
Bos taurus

platform_organism:
Sus scrofa
Bos taurus


title: Genome-wide binding of SEPALLATA3 (SEP3) and AGAMOUS (AG) complexes determined by sequential DNA-affinity purification sequencing

city: Grenoble

first: Jérémy

last: LUCAS

address: 17 avenue des Martyrs

institute: CNRS

summary:
In this study, we compare the DNA binding specifify and affinity of  SEPALLATA3 and AGAMOUS complexes
The MADS transcription factors, SEPALLATA3 (SEP3) and AGAMOUS (AG), are required for floral organ identity and determinacy of the floral meristem in Arabidopsis. Dimerization is obligatory for their DNA binding, however SEP3 and SEP3-AG also form tetrameric complexes. The goal of this study is to understand how homo and hetero-dimerization and tetramerization of MADS TFs affect genome-wide DNA-binding patterns. Using a modified sequential DNA affinity purification sequencing protocol (seq-DAP-seq), we selectively purified SEP3 homomeric and SEP3-AG heteromeric complexes, including the dimeric SEP3   tet-AG complex and the tetrameric SEP3-AG complex, and determined their genome-wide binding.

overall_design: We use a in vitro genome-wide DNA binding assay, termed sequential DNA affinity purification sequencing, to determine the genome-wide binding of SEP3-AG tetramer, SEP3∆tet-AG dimer, and SEP3 homocomplex.

first: Xuelei

last: Lai

first: Arnaud

last: Stigliani

last: Lucas

first: François

last: Parcy

first: Chloe

last: Zubieta


title: Pygopus2-Histone interaction is important for de-differentiation and breast cancer progression [RNA-Seq]

first: Ravi Kiran Reddy

last: KALATHUR

overall_design: RNA-Seq of Pygo2+/+ ( WT) and Pygo2 AE/AE mutant cell lines on treatment with TGFb or Wnt3a was performed in biological duplicates

first: Meera

last: Saxena

first: Ravi-Kiran-Reddy

last: Kalathur

first: Gerhard

last: Christofori


title: Lipid stores and lipid metabolism associated gene expression in porcine and bovine parthenogenetic embryos revealed by immunostaining and RNA-seq [Bt]

last: Kajdasz

overall_design: 2  blastocyst samples, 3 inner cell mass (ICM) samples, 3 trophectoderm (TE) samples


title: The role of Kdm5c

city: Los Angeles

first: Karen

last: Reue

state: CA

address: 695 Charles E. Young Drive South

institute: University of California, Los Angeles


title: LncRNA RVRL1 restricts rabies virus infection by disrupting PRC2 complex [RNA-seq]

first: Dong

last: Chen

summary: Rabies is an ancient infectious disease but still lacking efficient therapeutic approach despite of vaccine. In this study, we have identified a novel cytoplasmic lncRNA, namely rabies virus related lncRNA 1(RVRL1), whose expression in neuronal cells is up-regulated upon the infection of the causative agent of rabies, the neurotropic virus rabies virus (RABV). RVRL1 effectively inhibits RABV infection both in neuronal cells and in a mouse model. RVRL1 binds to EZH2 and disrupts the PRC2 complex, which is consistent with the inverse relationship between RVRL1 expression and the cellular H3K27me3 level. RVRL1 expression positively regulates the expression of PCP4L1 encoding a 10 kD peptide, which is shown to inhibit RABV replication. These findings highlight a novel mechanism for lncRNAs to upregulate the expression of antiviral genes, and define two potential anti-rabies reagents including an antiviral lncRNA and an antiviral peptide.

overall_design: Neuron cell and tissue mRNA profile of three time points after RABV infection,and control samples were generated by deep sequencing, in triplicate.  Neuron cell mRNA profile of two time points after RABV infection and RVRL1+/- were generated by deep sequencing, in duplicate.

first: Ling

last: Zhao

first: Yi

last: Zhang

first: Baokun

last: Sui

first: Jing

last: Hou


title: LncRNA RVRL1 restricts rabies virus infection by disrupting PRC2 complex [ChIP-seq]

overall_design: H3K27me3 marker profile of Neuron cells after RABV infection and RVRL1+/- were generated by deep sequencing, in duplicate.Input control samples were generated by deep sequencing, in duplicate.


title: Cell-of-origin and Developmental Trajectories Cooperate to Determine Chromatin Landscapes in Histone-Mutant Diffuse Midline Gliomas

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

summary: Diffuse midline gliomas (DMGs) are universally fatal pediatric brain tumors associated with mutations in genes encoding either histone H3.1 or histone 3.3, often substitution of methionine for lysine 27 (H3K27M). H3K27 is a critical determinant of chromatin state via methylation by Enhancer-of-Zeste-Homolog-2 (EZH2). Previous reports have suggested that the pathologically low levels of H3K27me3 found in histone-mutant DMGs result primarily from H3K27M inhibiting EZH2 directly, but recent reports have called this model into question. To better understand the chromatin landscape of DMGs, we applied CUT&RUN to patient-derived DMG cell lines. Remarkably, we find that the PRC2 activity is similar in DMGs and embryonic stem cells, suggesting a primitive cell-of-origin, despite transcriptionally active regions maintaining markers of both stem cells and differentiated cells. We also show that exogenous expression of H3.3M at physiological levels has little effect on H3K27me3 levels, that H3K27M can colocalize with H3K27me3 in vivo and that the H3.3K27M oncohistone does not show evidence of sequestering PRC2 components. Our results suggest that chromatin landscapes in DMGs are a consequence of a stem-like chromatin state that is retained despite activation of differentiation programs. Our findings have implications for understanding DMG gliomagenesis and therapeutic approaches centered on epigenome-modifying agents.

overall_design: We used Cleavage under targets and Release using nuclease (Cut-and-Run), a chromatin profiling strategy in which antibody-targeted controlled cleavage by micrococcal nuclease releases specific protein-DNA complexes into the supernatant for paired-end DNA sequencing.

first: Jay

middle: F

last: Sarthy

first: Michael

last: Meers

first: Eric

last: Ferguson

first: Derek

middle: H

last: Janssens

first: Nicholas

last: Vitanza

first: Kami

last: Ahmad

first: James

middle: M

last: Olson

first: Steven


title: RNA-seq analyses of endothelial-like cells from human embryonic stem cells

city: Changsha

first: hao

last: zhao

state: Hunan

address: 8 Luyun Road

department: School of Basic Medical Science

institute: Central South University

summary:
Rationale-Endothelial cells (ECs) play important roles in various regeneration processes and can be used in a variety of therapeutic applications, such as cardiac regeneration, gene therapy, tissue-engineered vascular grafts and prevascularized tissue transplants. ECs can be acquired from pluripotent and adult stem cells.
Objective-To acquire ECs from human embryonic stem cells (hESCs) in a fast, efficient and economic manner.
Methods and Results-We established a conditional overexpression system in hESCs based on 15 transcription factors reported to be responsible for hematopoiesis lineage. Among them, only overexpression of FLI1 could induce hESCs to a hematopoietic lineage. Moreover, simultaneous overexpression of FLI1 and activation of PKC rapidly and efficiently induced differentiation of hESCs into induced endothelial cells (iECs) within 3 days, while neither FLI1 overexpression nor PKC activation alone could derive iECs from hESCs. During induction, hESCs differentiated into spindle-like cells that were consistent in appearance with ECs. Flow cytometric analysis revealed that 92.2%-98.9% and 87.2-92.6% of these cells were CD31+ and CD144+ respectively. Expression of vascular-specific genes dramatically increased, while the expression of pluripotency genes gradually decreased during induction. iECs incorporated acetylated low-density lipoproteins, strongly expressed vWF and bound UEA-1. iECs also formed capillary-like structures both in vitro and in vivo. RNA-seq analysis verified that these cells closely resembled their in vivo counterparts.
Conclusions-Our results showed that co-activation of FLI1 and PKC could induce differentiation of hESCs into iECs in a fast, efficient and economic manner.

overall_design: RNA-seq analyses of endothelial-like cells(D1,D2,D3) from human embryonic stem cells (D0)and epc(control).

first: Liang

last: Hu

first: Hao


title: Oncogenes overexpressed in metastatic oral cancers from patients with pain: potential pain mediators released in exosomes

city: New York

first: Igor

last: Dolgalev

state: New York

address: 550 1st Ave

institute: NYU Grossman School of Medicine

summary: Oral cancer patients experience pain at the site of the primary cancer.  Patients with metastatic oral cancers report greater pain.  Lack of pain identifies patients at low risk of metastasis with sensitivity = 0.94 and negative predictive value = 0.89.  In the same cohort, sensitivity and negative predictive value of depth of invasion, currently the best predictor, were 0.95 and 0.92, respectively.  Cancer pain is attributed to cancer-derived mediators that sensitize neurons and is associated with increased neuronal density.  We hypothesized that pain mediators would be overexpressed in metastatic cancers from patients reporting high pain.  We identified 40 genes overexpressed in metastatic cancers from patients reporting high pain (n=5) compared to N0 cancers (n=10) and normal tissue (n=5).  The genes are enriched for functions in extracellular matrix organization and angiogenesis.  They have oncogenic and neuronal functions and are reported in exosomes.  Hierarchical clustering according to expression of neurotrophic and axon guidance genes also separated cancers according to pain and nodal status.  Depletion of exosomes from cancer cell line supernatant reduced nociceptive behavior in a paw withdrawal assay, supporting a role for exosomes in cancer pain.  The identified genes and exosomes are potential therapeutic targets for stopping cancer and attenuating pain.

overall_design: Identification of genes overexpressed in node positive (N+) cancers from patients who reported high levels of pain at the site of the cancer by comparison with node negative (N0) cancers and normal oral tissue

first: Donna

last: Albertson


title: Comprehensive analysis of soluble RNAs in human embryo culture media

city: Aarhus N

first: Morten

middle: T

last: Veno

state: DK

address: Aabogade 15

country: Denmark

institute: Omiics ApS

type: Non-coding RNA profiling by high throughput sequencing

summary: Spent culture media was collected on day 5 after in vitro fertilization from 5 human blastocysts that were subsequently transferred or frozen. Corresponding blank culture media from the same lot were cultured alongside the embryos and used as negative controls along with PBS. The samples were prepared for total RNA sequencing using SMARTer® Stranded Total RNA-Seq Kit v2 - Pico Input Mammalian (Takara) and for small RNA sequencing using the QIAseq RNA Library Prep kit (Qiagen). Prepared libraries were sequenced as 100 bp paired end on an Illumina HiSeq 4000 sequencer for total RNA and Illumina NextSeq500 sequencer for small RNA. We found that total RNA mapped only 3% of the reads to gene regions in spent culture media and control and only 2% in PBS samples mapped to gene regions. For the small RNA analysis, human annotated miRNAs constituted from 0.1% to 1.4% of the mapped reads and 40% of small RNA reads mapped to the human genome. tiRNAs derived from 5 different mature tiRNA were differentially expressed when comparing conditioned to unconditioned media. We conclude,  that in  spite of applying state-of-the-art sensitive detection methods no miRNAs were found to be reliably present in the spent culture medium. In contrast, tiRNA fragments appear to be overexpressed in cultured IVF media samples.

overall_design: examination of total RNA and small RNA in spent culture medium from human in vitro fertilized blastocysts

first: Kirstine

last: Kirkegaard

last: Venø

first: Jørgen

last: Kjems


title: Reduced representative bisulfite sequencing (RRBS) analysis for a genome-wide comparison of DNA methylation in wild-type, USP7-KO-1, DNMT3A/3B-DKO and DNMT3A/DNMT3B/USP7-TKO HeLa cell lines.

first: Jialun

last: Li

address: N0.500, Dongchuan road

department: School of life science

institute: ECNU

type: Methylation profiling by high throughput sequencing

summary: To better characterize the effects of USP7 on DNA methylation, we performed reduced representative bisulfite sequencing (RRBS) analysis for a genome-wide comparison of DNA methylation in wild-type, USP7-KO-1, DNMT3A/3B-DKO and DNMT3A/DNMT3B/USP7-TKO HeLa cell lines. RRBS analysis showed increased DNA methylation in USK7-KO cells and loss of USP7 elevates DNA methylation on pre-existing sites and de novo methylation.

overall_design: Examination of DNA methylation in 4 cell types.

first: Jiemin

last: Wong


title: Identification of genes regulated by the MADS transcription factor (TF), SEPALLATA3, in the context of the double sep1sep2 and triple sep1sep2sep3 mutant background, by RNA-Seq analysis

first: Veronique

last: Hugouvieux

address: 17 rue des Martyrs

department: IRIG

institute: CEA Grenoble

summary: These analyses allowed the identification of more than 2000 genes regulated by the MADS TF, SEPALLATA3 (SEP3). Although the involvement of SEP3 in all flower organ development is well established,  the single sep3  mutant shows no flower phenotype, due to redundancy with other SEPALLATA genes. The triple mutant, sep1sep2sep3, shows no floral organ development except sepals, while the sep1sep2 mutant showed characteristic WT flower with, from the outer to the inner whorl of the flower, 4 sepals, 4 petals, 6 stamens and one gynoecium. We performed RNA-seq analysis on sep1sep2, sep1sep2sep3 and on sep1sep2sep3 lines complemented with SEP3 to highlight genes involved in floral organ development and controlled by SEP3. For this purpose we compared gene expression between the triple and the double mutant, and between triple mutant lines complemented with SEP3 and the triple mutant.  Genes were considered regulated by SEP3 when the log FC was between 1 and -1 and the FDR <0.05. Using these criteria, we obtained a list of  >2000 genes regulated by SEP3.

overall_design: RNA seq analysis were performed on inflorescence meristem and flower buds up to stage 10-11 harvested from Arabidopsis plants grown in parralel in long days conditions. Experiments were performed on 2 replicates from sep1sep2 mutant, 2 replicates from sep1sep2sep3 triples mutant, and 2 independantes complemented lines expressing SEP3 in the sep1sep2sep3 background (considered as 2 replicats). These lines are described in Hugouvieux et al, Nucleic Acid Research, 2018:1;46(10):4966-4977. doi: 10.1093/nar/gky205. Libraries and sequencing were performed by Genewiz (Illumina High SEq, 2X150 bp configuration).

first: Véronique

first: Jeremy

last: Lucas

first: Chloé

last: Zubieta


title: High-throughput sequencing of 3T3-L1 cells with Kdm5c knockdown

first: Karen

last: Reue

summary: One day prior to confluency, non-targeting (scramble control) siRNA or Kdm5c siRNA were transfected into 3T3-L1 cells and gene expression was assessed by RNA-seq at 24hr, 48hr, and 72hr post transfection.

overall_design: 3T3-L1 preadipocytes were reverse-transfected with 50 nM non-targeting (scramble control) or Kdm5c siRNA with Lipofectamine RNAiMAX at differentiation day –3 (sub-confluent). For RNA isolation (transcriptome analysis), cells were collected at 24hr (day –2), 48hr (day –1), and 72hr (day 0) post-transfection.

first: Carrie

last: Wiese


title: Chromatin accessiblity of 3T3-L1 cells with Kdm5c knockdown

summary: One day prior to confluency, non-targeting (scramble control) siRNA or Kdm5c siRNA were transfected into 3T3-L1 cells and chromatin structure was assessed by ATAC-seq at 24hr, 48hr, and 72 hr post transfection.

overall_design: 3T3-L1 preadipocytes were reverse-transfected with 50 nM non-targeting (scramble control) or Kdm5c siRNA with Lipofectamine RNAiMAX at differentiation day –3 (sub-confluent). Nuclei were  collected at 24hr (day –2), 48hr (day –1), and 72hr (day 0) post-transfection and prepared for ATAC-seq.


title: Global gene expression pattern in human first trimester primary villous cytotrophoblast cells (vCTBs) in comparison with human first trimester placental villous mesenchymal cells

status: Public on Sep 06 2020

city: Kansas City

first: Sumedha

last: Gunewardena

state: Kansas

address: 2146 West 39th Avenue

department: Dept. of Molecular and Integrative Physiology

institute: University of kansas medical center

summary: vCTBs and mesenchymal cells were isolated from palcental villi of medically terminated and deidentified first trimester placenta (pulled samples from 4-6 placentas of 8-10 week gestation).

overall_design: Proliferating vCTBs and mesenchymal cells were isolated through enzymatic digestion followed by a percoll gradient separation and hence cultured in a specilized medium (TM, #7121, ScienCell Research Laboratories). RNA-seq analysis were performed using a illumina HiSeq2500 platform.

first: Soumen

last: Paul

first: Biswarup

last: Saha


title: Genome-wide analysis of PmTCP4 transcription factor binding sites by ChIP-Seq during pistil abortion in Japanese apricot

city: Nanjing

first: Shahid

last: Iqbal

state: Jiangsu

address: Nanjing, China

institute: Nanjing Agriculture University

summary: The TCP4 transcription factor plays an important role in plant growth and development, especially in flower development. PmTCP4 is involved in the process of pistil abortion in Japanese apricot, but its molecular mechanism, particularly the DNA binding sites and co-regulatory genes, are quite unknown. Therefore, to identify the genome-wide binding sites of PmTCP4 transcription factors and their co-regulatory genes, chromatin immunoprecipitation sequencing (ChIPSeq) was carried out. ChIP-Seq data produced the maximum enriched peaks in two Japanese apricot cultivars ‘Daqiandi’ (DQD) and ‘Longyan’ (LY), which showed that the majority ofDNA-protein interactions are relevant and have a significant function in binding sites. Moreover, 720 and 251 peak-associated genes regulated by PmTCP4 were identified in DQD and LY, respectively, and most of them were involved in the flower and pistil development process. Furthermore, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed that photosynthesis and oxidative phosphorylation were the most enriched pathways in both cultivars and all identified genes related to these pathways were down-regulated. This study will provide a reference for a better understanding of the PmTCP4 regulatory mechanism during pistil abortion in Japanese apricot.

overall_design: The samples (pistils of flower buds) of two cultivars "Daqiandi and Longyan" of Japanese Apricot were selected. The samples was prepared for ChIP-Seq Analysis as the method described by Kaufmann et al., 2010

first: Zhihong

last: Gao


title: scRNA-Seq of human iPSC derived lung alveolar epithelial type 2 cells with/without dox-induced activation of oncogenic KRAS G12D

city: Boston

first: Carlos

last: Villacorta-Martin

state: MA

address: 670 Albany St

institute: Center for Regenerative Medicine at Boston University

summary: Lung adenocarcinoma is responsible for significant global mortality with limited effective treatments. Although some studies suggest that these tumors arise from alveolar epithelial type 2 cells (AEC2s), there is scant information regarding the early events that might occur in human AEC2s at the inception of oncogenesis. This limitation, is partially due to a lack of human model systems that recapitulate the initiation of oncogenesis in AEC2s. Unfortunately, primary AEC2s from patients are difficult to access in vivo or stably maintain in cell cultures. Hence, we sought to develop an in vitro system to model the early stages of oncogenesis utilizing human induced AEC2s (iAEC2s) generated through the directed differentiation of induced pluripotent stem cells (iPSCs). To this end, we selected a normal human iPSC line we have previously engineered to carry fluorochrome reporters targeted to lung epithelial-specific loci, NKX2-1GFP and SFTPCtdTomato that enable monitoring and purification of alveolar lung epithelial cells.  To test the effects of adenocarcinoma oncogene induction in these cells, we targeted a third locus, AAVS1 using gene editing to engineer a doxycycline-inducible cassette encoding mutant KRASG12D, the most commonly found oncogene in lung adenocarcinomas. Successful induction of KRASG12D with doxycycline was demonstrated in both the targeted undifferentiated iPSCs as well as in the iAEC2s derived from these cells. We profiled the downstream effects of KRASG12D induction in iAEC2s, comparing dox vs vehicle exposed cells by cell counting, FACS for NKX2-1GFP/SFTPCtdTomato, RT-qPCR, deep proteomic and phosphoproteomic analyses, and scRNA-sequencing. Through this characterization, we found that induction of KRASG12D robustly activates MAPK signaling resulting in a shift of iAEC2s away from their mature alveolar program towards a distal lung epithelial progenitor phenotype, indicated by the upregulation of lung progenitor and proliferation markers (e.g. SOX9, ETV4, LEF1, TM4SF1, MKI67, and TOP2A) while maintaining NKX2-1 expression, at the expense of mature alveolar markers (e.g. SFTPC, SFTPB, NAPSA, and LPCAT1). Successful modeling of lung adenocarcinoma with this model system has a variety of future applications, including testing unknown mechanisms for oncogenesis, discovery of novel biomarkers of disease, or development of new effective treatment methods through drug screening.

overall_design: Human induced pluripotent stem cells (iPSCs; BU3 NGST line) carrying a GFP reporter targeted to the NKX2-1 locus (NKX2-1GFP) and a tdTomato reporter targeted to the SFTPC locus (SFTPCtdTomato) underwent gene editing to engineer a dox-inducible KRAS G12D coding sequence targeted to the AAVS1 locus (BU3 NGST KRAS G12D, clone 3).  iPSC-derived Alveolar epithelial type 2 cells (iAEC2s) were derived by directed differentiation of this line according to the protocol of Jacob/Kotton et al (Cell Stem Cell 2017) and were maintained as self-renewing epithelial spheres in 3D Matrigel culture. At passage 3, parallel wells of the iAEC2s were treated with either control vehicle (DMSO) or doxycycline (Dox; 1ug/mL) to induce expression of KRAS G12D. After 4 more passages and 69 days of exposure to Dox or DMSO (total differentiation time= 127 days), cells were dissociated from 3D Matrigel, and sorted for Calcein Blue+ live cells in preparation for 10X Chromium Genomics 3’ v3 scRNA-Seq analysis. (Note: cells were re-sorted for NKX2-1GFP+ cells 1 passage [2 weeks] prior to  final harvest to enrich for lung epithelial cells).

first: Marally

last: Vedaie

first: Darrell

last: Kotton


title: High-throughput sequencing of inguinal white adipose tissue fractions from female C57BL/6 Kdm5c+/+ and Kdm5c+/– mice

summary: RNA sequencing was performed on white adipose tissue fractions to determine the effects of having one vs. two copies of the Kdm5c gene in female mice.

overall_design: Inguinal white adipose tissue was fractionated into mature adipocytes and stromal vascular fraction for RNA sequencing


title: Duke DNase-seq in LNCaP

status: Public on Sep 05 2020

city: Chicago

first: Sebastian

last: Pott

state: IL

address: 920 E 58th St

institute: The University of Chicago

summary: We collected DNase-seq data in LNCaP cells (human prostate adenocarcinoma cell line) before and after 45 minutes and 4 hours of 1 nM R1881 treatment. We then used these DNase-seq data to predict the quantitative TF occupancy with our statistical method (TOP) and examine how their occupancy changes genome-wide across treatment conditions.

overall_design: Examination of chromatin accessibility (DNase-seq) in LNCaP cells before and after 45 minutes and 4 hours of 1 nM R1881 treatment, each with two replicates.

first: Kaixuan

last: Luo

first: Alok

last: Tewari

first: Lingyun

last: Song

first: Gregory

last: Crawford

first: Alexander

middle: J

last: Hartemink


title: Bulk RNA sequencing for sNinj1-mimetic peptide-treated classically activated macrophages

status: Public on Sep 04 2020

city: Seoul

first: GooTaeg

last: Oh

address: 52, Ewhayeodae-gil

department: Department of Life Sciences

country: South Korea

institute: Ewha Womans University

summary: We have applied bulk RNA sequencing as an unbiased transcriptomics to demonstrate the mechanism underlying actions of sNinj1 in macrophage inflammation. The anti-inflammatory role of Ninj1 was verified by treating macrophages with the peptide Ninj11-56 (ML56), which mimic the soluble form of Ninj1 (sNinj1).

overall_design: Peripheral blood samples from 10- to 16-week-old C57BL/6J (B6) mice (n=60, 1 ml per mouse) were collected in EDTA-coated tubes, pooled according to the experimental requirements. The CD11b-selected monocyte suspension was cultured for 6 or 7 days with RPMI/FBS plus 30 ng/ml M-CSF. For activation in vitro, mouse differentiated macrophages were stimulated with lipopolysaccharide (LPS, 500 ng/ml) and IFNγ (20 ng/ml) to M(LPS/IFNγ)s, which were classically activated macrophages (CAMs).

first: Sejin

last: Jeon

first: Tae

last: Kim

first: Sung

last: Park

first: Tae-Young

last: Roh


title: Impact of the long non-coding RNA CRNDE on the transcriptome of the multiple myeloma cell line KMS11

city: Paris

first: David

last: GARRICK

address: 1 Av. Claude Vellefaux

department: INSERM U976

institute: Institut de Recherche Saint Louis

summary: Multiple myeloma (MM) is a currently incurable malignancy of antibody-secreting plasma cells. Long non-coding RNAs (lncRNAs) have been recognised as an important class of regulatory molecules which are increasingly implicated in tumorigenesis. While recent studies have demonstrated changes in expression of lncRNAs in MM, the functional significance and molecular pathways downstream of these changes remain poorly characterised. In this study we have performed CRISPR-mediated deletion of the locus encoding the lncRNA Colorectal Neoplasia Differentially Expressed (CRNDE), a known oncogenic lncRNA that is overexpressed in plasma cells of MM patients and is a marker of poor prognosis. We found that CRISPR-mediated deletion of the CRNDE locus in MM cells decreases proliferation and adhesion properties, increases sensitivity to Dexamethasone and reduces tumour growth in an in vivo xenograft model. Transcriptomic profiling in CRNDE-deleted MM cells demonstrated that CRNDE activates expression of a number of genes previously implicated in the aetiology of MM, including IL6R. We further demonstrate that deletion of the CRNDE locus diminishes IL6 signalling and proliferative responses in MM cells. Altogether this study reveals the IL6 signalling pathway as a novel mechanism by which CRNDE impacts upon MM cell growth and disease progression.

overall_design: Comparison of the transcriptome in wild type KMS11 multiple myeloma clones (+/+), or clones which are either heterozygous (+/-) or homozygous (-/-) for a CRISPR-engineered deletion of exons 4-6 of the gene encoding the long non-coding RNA CRNDE. Two clones were analysed per genotype.

first: Antoine

last: David

first: Michele

last: Goodhardt

last: Garrick


title: Monocytic ontogeny of regenerated macrophages characterizes the mesotheliomagenic responses to carbon nanotubes

city: Woluwé

first: Jérôme

last: Ambroise

address: Avenue Hippocrate 54

department: Institut de Recherche Expérimentale et Clinique

country: Belgium

institute: Université catholique de Louvain

summary: Macrophages not only derive from circulating blood monocytes, but also originate from embryonic progenitors that can proliferate. This dichotomic origin has an impact on macrophage functions in steady state and pathological conditions. Long needle-like carbon nanotubes (CNT) damage serous membranes and induce mesothelioma like asbestos fibers. Macrophages predominantly infiltrate CNT-induced injured tissue and contribute to cancer development. Here, we revealed the exact ontogeny of macrophage populations after CNT exposure by comparing the response to needle-like mesotheliomagenic CNT-7 with tangled-like non-mesotheliomagenic CNT-T. In a rat peritoneal cavity model, both CNT induced a rapid and complete depletion of MHCIIlow resident macrophages generating an empty niche available for macrophage repopulation. Macrophage disappearance after mesotheliomagenic CNT-7 was followed by a substantial inflammatory reaction, and macrophage replenishment completed after 7 days. Fifteen days after non-mesotheliomagenic CNT-T, macrophage repopulation was incomplete and accompanied by a limited inflammatory reaction. Cell depletion experiments, flow cytometry and RNA-seq analysis demonstrated that, after CNT-7, resident macrophages were mainly replaced by an influx of monocytes, which differentiated locally into MHCIIhigh inflammatory macrophages. In contrast, the low inflammatory response induced by CNT-T was linked to the activation of self-renewing MHCIIlow macrophages. In conclusion, the mesotheliomagenic response to CNT specifically relies on macrophage niche recolonization by monocytic progenitors. The homeostasis returns after exposure to non mesotheliomagenic CNT-T involves, in contrast, a macrophage maintenance by proliferation. Macrophage depletion and repopulation are thus decisive events characterizing the carcinogenic activity of fibrous particles.

overall_design: RNA-sequencing analysis of peritoneal macrophages purified from mesotheliomagenic (CNT-7) and non mesotheliomagenic (CNT-T) carbon nanotube treated  rats. There are 4 rats for each treatment.

first: Micaela

last: Orsi

first: Mihaly

last: Palmai-Pallag

first: Yousof

last: Yakoub

first: Saloua

last: Ibouraadaten

first: Michèle

last: De Beukelaer

first: Caroline

last: Bouzin

first: Bertrand

last: Bearzatto

first: Jean-Luc

last: Gala

first: Davide

last: Brusa

first: Dominique

last: Lison

first: François

last: Huaux


title: Bulk RNA sequencing for classically activated macrophages (CAMs) from Ninj1+/+ and Ninj1-/- mice

first: GooTaeg

last: Oh

summary: We have applied bulk RNA sequencing as an unbiased transcriptomics to explore the association between Ninj1 deletion and macrophage activation.

overall_design: BM cells were isolated from the femurs and tibias of Ninj1+/+ and Ninj1-/- mice at 6 weeks of age and cultured in DMEM containing 20% FBS and 20 μM β-mercaptoethanol for 16 hours. The suspended cells were plated and treated with 20 ng/ml recombinant human/mouse macrophage colony-stimulating factor (M-CSF, 30ng/ml) for 7 days to differentiate macrophages. For activation in vitro, mouse differentiated macrophages were stimulated with lipopolysaccharide (LPS, 500 ng/ml) and IFNγ (20 ng/ml) to M(LPS/IFNγ)s, which were classically activated macrophages (CAMs).

first: Sejin

last: Jeon

first: Tae

last: Kim

first: Sung

last: Park

last: Roh


title: HIV cell-to-cell spread slows evolution of drug resistance

city: Durban

first: Alex

last: Sigal

state: KwaZulu-Natal

address: 719 Umbilo Road, K-RITH Tower building, Level 5

country: South Africa

institute: Africa Health Research Institute

summary:
Here we investigated whether a combination of cell-free infection and cell-to-cell spread confers a selective advantage in the evolution of resistance to an inhibitor relative to cell-to-cell spread alone due to the stronger selection pressure against drug sensitive virus.
We propagated HIV infection using coculture of infected with uninfected cells in the face of the reverse transcriptase inhibitor efavirenz (EFV), and compared the effect on drug resistance evolution of including one cycle of cell-free infection. In the presence of a single cell-free infection step, we obtained earlier evolution of resistance to EFV. When we increased selective pressure by adding emtricitabine (FTC as a second drug, infection with the cell-free step evolved multidrug resistance and was able to replicate, while infection without a cell-free step failed to evolve multidrug resistance.
In conclusion, our results suggest that, HIV cell-to-cell spread has a decreased capacity to rapidly evolve resistance to inhibitors, which is conferred by cell-free infection.

overall_design:
Mathenmatical modeling of evoltion of drug resistance by cell-free infection and cell-to-cell sread supported by sequencing of Reverse transciptase region of proviral DNA from infected cell populations over 48hr timepoints at 20nM EFV, 40nM EFV and a combination of 20nM EFV and 770nM FTC
E7 clone was generated by subcloning RevCEM cells at single cell density. Surviving clones were subdivided into replicate plates. One of the plates was screened for the fraction of GFP expressing cells upon HIV infection using microscopy, and the clone with the highest fraction of GFP positive cells was selected

first: Jessica

last: Hunter

first: Sandile

last: Cele

first: Laurelle

last: Jackson

first: Jennifer

last: Giandhari

first: Tulio

last: de Olivera

first: Gila

last: Lustig


title: Methylation profiles of benign and malignant bone tumorus

city: London

first: Adrienne

last: Flanagan

state: Middlesex

address: Huntley Street

institute: University College London

type: Methylation profiling by genome tiling array

summary: DNA methylation profiles or a range of primary bone tumours

overall_design: DNA methylation profiles were obtained from a range of primary bone tumours obtained from the Royal National Orthopaedic Hospital, Stanmore, UK


title: Expression data from the mouse brain after 90'-MCAO and sham-control surgery

city: Barcelona

first: Alba

last: Simats

state: Catalunya

address: Passeig Vall d'Hebron, 119-129

country: Spain

institute: Vall d'Hebron Institute of Research

summary: Stroke remains a major leading cause of death and disability worldwide. Despite continuous advances, the identification of key molecular signatures of ischemic stroke within the hyper-acute phase of the disease is still of primary interest for a real translational research on stroke diagnosis, prognosis and treatment. High-throughput -omics technologies are enabling large-scale studies on stroke pathology at different molecular levels. Data integration resulting from these -omics approaches is becoming crucial to unravel the interactions among all different molecular elements and highly contribute to interpret all findings in a complex biological context. Here, we have used advanced data integration methods for multi-level joint analysis of transcriptomics and proteomics datasets depicted from the mouse brain 2h after cerebral ischemia. By modeling net-like correlation structures, we identified a set of differentially expressed genes and proteins with a relevant association in stroke pathology. The ischemia-induced deregulation of 10 of these inter-correlated elements was successfully verified in a new cohort of ischemic mice and changes in their expression pattern were also evaluated at a later time-point after cerebral ischemia. Of those, CLDN20, GADD45G, RGS2, BAG5 and CTNND2 were highlighted as potential candidates as blood biomarkers of cerebral ischemia and hence they were evaluated in blood samples from ischemic and sham-control mice and in ischemic strokes and patients presenting stroke-mimicking conditions. Our findings indicated that CTNND2 and GADD45G levels in blood within the first hours after ischemic stroke might be potentially useful to discriminate ischemic strokes from mimics and to predict patients’ poor outcome after stroke, respectively. In summary, we have here used for the first time an integrative approach that enabled us to elucidate by means of biostatistical tools key elements of the initial stages of the stroke pathophysiology and highlight new outstanding proteins that might be further considered as blood biomarkers of ischemic stroke.

overall_design: Mice were submitted to 90'-middle cerebral artery occlusion (intraluminal model) or sham-control surgery. Thirty minutes later (2h from the induction of ischemia) animals were euthanized and transcardially perfused with cold-saline. Brains were collected and ipsilateral (IP) and contralateral (CL) brain hemispheres were separated. mRNA fraction was obtained from each sample with the mirVana Paris kit and kept frozen (-80ºC) until the transcriptomic array was conducted.

first: Teresa

last: García-Berrocoso

first: Ricardo

last: Gonzalo

first: Joan

last: Montaner


title: Dynamics of the cell-free DNA methylome of metastatic prostate cancer during androgen-targeting treatment

city: Toronto

first: Bharati

last: Bapat

state: Ontario

address: 60 Murray Street, Rm L6-304B

department: Lunenfeld-Tanenbaum Research Institute

country: Canada

institute: Sinai Health System

summary: Aim: We examined methylation changes in cell-free DNA (cfDNA) in metastatic castration resistant prostate cancer (mCRPC) during treatment. Patients and Methods: Genome-wide methylation analysis of sequentially collected cfDNA samples derived from mCRPC patients undergoing androgen-targeting therapy was performed. Results: Alterations in methylation states of genes previously implicated in prostate cancer progression were observed, and patients that maintained methylation changes throughout therapy tended to have a longer time to clinical progression (TTP). Importantly, we also report that markers associated with a highly aggressive form of the disease, Neuroendocrine-CRPC, were associated with a faster TTP. Conclusion: Our findings highlight the potential of monitoring the cfDNA methylome during therapy in mCRPC, which may serve as predictive markers of response to androgen-targeting agents.

overall_design: We performed methylated DNA immunoprecipitation followed by high-throughput sequencing (cfMeDIP-seq) of cfDNA derived from 16 patients with mCRPC at various stages of treatment with androgen-targerting agents. Visit A = Baseline/pre-treatment visit; Visit B = Week-12 (+/- 2 weeks) during treatment; Visit C = Clinical progression/change of treatment. Each cfDNA sample has a methylated DNA enriched sequencing file and an input control file.

first: Madonna

last: Peter

first: Misha

last: Bilenky


title: PTEN-dependent regulation of genome-wide transcription during metabolic stress [RNA-Seq]

city: Cleveland

first: Ata

last: Abbas

state: OH

address: 10900 Euclid Ave

department: Division of Hematology & Oncology

institute: Case Western Reserve University

summary: PTEN is implicated in a wide variety of pathophysiological conditions and traditionally studied in the context of the PIK3-AKT-mTOR axis. Recent studies from our group and others have reported a novel role of PTEN in the regulation of transcription at the genome-wide scale. This emerging role of PTEN on global transcriptional regulation is providing a better understanding of various diseases, including cancer. Since cancer progression is an energy-demanding process and PTEN is known to regulate metabolic processes, we sought to understand the role of PTEN in transcriptional regulation under metabolic stress, a condition often developing in the tumor microenvironment. In the present study, we demonstrate that PTEN modulates genome-wide RNA Polymerase II (Pol II) occupancy in cells undergoing glucose deprivation. The glucose-deprived PTEN null cells were found to continue global gene transcription, which may activate a survival mode. However, cells with constitutive PTEN expression slow transcription, an evolutionary mechanism that may save cellular energy and activate programmed cell death pathways, in the absence of glucose.  Interestingly, alternative exon usage by PTEN null cells is increased under metabolic stress compared to PTEN expressing cells. Overall, our study comprehensively demonstrates distinct mechanisms involved in PTEN-dependent genome-wide transcriptional control under metabolic stress. Our findings provide a new insight in understanding the tumor microenvironment and how PTEN loss of function, whether by genetic or non-genetic mechanisms, can contribute to a favorable transcriptional program employed by tumor cells to escape apoptosis, hence developing more aggressive and metastatic phenotypes.

overall_design: RNA-seq was performed on BT-549 control and PTEN over-expressing BT-549_PTEN cell.

first: Charis

last: Eng


title: Differential expression and bioinformatics analysis of circRNA expression in migraine patients

city: haerbin

first: lin

last: jinghan

address: 23 Postoffice Street, Nangang District

department: neurology

institute: The First Affiliated Hospital of Harbin Medical University

summary: Background：CircRNAs are non-coding RNA molecules that have recently been described and shown to regulate miRNA functionality. While recent studies have suggested such circRNAs to be associated with pain related diseases in humans, no comprehensive migraine-related circRNA profiles have been generated and there is currently no clear understanding of whether they can serve as regulators of migraine pathology. Methods:We initially conducted a circRNA microarray analysis of the plasma of migraine patients and healthy controls. Based upon these data, we then selected 8 differentially expressed circRNAs and confirmed their expression in more migraine patient plasma samples via real-time PCR. We then performed functional and pathway enrichment analyses. Lastly, using a robust rank aggregation approach, we constructed a ceRNA network according to predicted circRNA–miRNA and miRNA–mRNA pairs in these migraine patient samples. Results:we were able to detect 2039 circRNAs in our patient samples, with and 794 of 1245 these circRNAs being up- and down-regulated in migraine patients relative to controls, respectively (fold change ≥1.5, P < 0.01). A qRT-PCR analysis confirmed that the expression of hsa_circRNA_100236, hsa_circRNA_102413, and hsa_circRNA_000367 was significantly enhanced in migraine patients, whereas the expression of hsa_circRNA_103809, hsa_circRNA_103670, and hsa_circRNA_101833 was significantly reduced in these individuals relative to healthy controls. We found these differentially regulated circRNAs to be associated with numerous predicted biological processes, with enrichment analyses suggesting that they may modulate PI3K-Akt signaling so as to promote inflammation to drive migraine development. However, further research will be needed to formally test these mechanistic possibilities and to validate these circRNAs as potential biomarkers of migraine patients.Conclusions: our results offer new potential insights into the mechanistic basis of this condition, and suggest that hsa_circRNA_000367 and hsa_circRNA_102413 may offer value as regulators of migraine pathology.

overall_design: We obtained peripheral blood samples for 4 migraine patients and 3 healthy control patients.

first: Jinghan

last: Lin

first: Shanshan

last: Shi

first: Qihui

last: Chen

first: Yonghui

last: Pan


title: miCLIP-seq of postnatal day 0 (P0) normal mouse skin epithelial cells

first: Tom

middle: Samuel

last: Carroll

address: 1230 York Avenue

institute: The Rockefeller University

summary: We employed miCLIP-seq to profile the location and extent of m6A in the Poly(A)+ transcriptome. Skin epithelial cells were isolated from wild-type P0 neonates by FACS. Total RNA was extracted by TRIzol-LS and Poly(A)+ RNA was extracted with Dynabeads™ mRNA Purification Kit (Thermo-Fisher, 61006). Input and miCLIP libraries was prepared from 3 biological replicates with each containing RNA isolated from 3 litters of neonates. Libraries were sequenced on the Illumina Hi-seq platform to generate paired-ended 50 bp reads. Sequencing data was processed as described in (Geula et al., 2015).

overall_design: Examination of transcriptome-wide m6A profile in normal skin epithelial cells.

first: Linghe

last: Xi

first: Elaine

last: Fuchs


title: mouse skin epithelial cells

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

overall_design: Refer to individual Series


title: PTEN-dependent regulation of genome-wide transcription during metabolic stress [ChIP-Seq]

overall_design: RNA Pol II ChIP-seq was performed on BT-549 control and PTEN over-expressing BT-549_PTEN cell.


title: MicroRNA microarray analysis for Primary human nasal epithelial cells (HNECs) treated with or without P1

first: Daniela

last: Tudor

address: 22, rue Mechain

department: Infection, Immunity  and Inflammation

institute: INSERM U1016

summary:
We investigated the microRNAs profiles upon nasal epithelial HNECs stimulation by P1 after treatment with or in absence of P1 for 6 hours, comparatively by microRNA array analysis.
Three untreated and treated paired samples from three independent experiments were analyzed by GeneChip miRNA 4.0 arrays (Affymetrix, Thermo Fisher) containing probes for 2578 human mature microRNAs and 2025 pre-mature microRNAs

overall_design: microRNAs differentially expressed with p value<0.05 from paired student’s t-test, and a cut-off of fold change >1.3 are included

first: Lin

last: XU

first: Morgane

last: BOMSEL


title: Caulobacter crescentus mRNA half-lives by rifampicin transcription shutoff

city: Bloomington

first: Jared

middle: Michael

last: Schrader

state: IN

address: 212 S Hawthorne Dr

department: Biology

institute: Indiana University

summary: mRNA half-life profiling in the bacterium Caulobacter crescentus was preformed by shutting of transcription with the antibiotic rifampicin, and following mRNA abundance at 1, 2, 4, 8, and 15 minutes post rifampicin.  All RNA measurements were performed on cells grown to mid-log in M2G minimal growth medium.  Two biological replicates time coursees were collected from independent starter cultures.

overall_design: Total RNA samples were collected just prior to rifampicin addition, and after 1, 2, 4, 8, or 15 minutes incubation with 100ug/mL rifampicin.  Each total RNA sample was depleted of rRNA using the ribozero kit (Illumina) for gram negative bacteria, and subjected to total RNA-seq libraries as in (Ingolia et al. Science 2009).  RNA was left in each sample as an internal control for stable RNA.  Resulting DNA libraries  were pooled, and sequenced in a Hiseq 2500 (U. Texas - Austin).

first: Nadra

last: Al-Husini


title: Quantitative transcriptome analysis of human embryonic stem cells, human mesenchymal stem cells and human musculoskeletal stem cells

city: Jeonju-si

first: Myung-Kwan

last: Han

state: Jeollabuk-do

address: Baeje-daero 567

department: Department of Microbiology

institute: Jeonbuk National University Medical School

summary: We first derived human musculoskeletal stem cells from human embryonic stem cells. mRNA profiles of human embryonic stem cells, human mesenchymal stem cells and human musculoskeletal stem cells were generated by QuantSeq 3’ mRNA-sequencing, in triplicate, using NextSeq 500.

overall_design: QuantSeq 3’ mRNA-sequencing of human embryonic stem cells, human mesenchymal stem cells and human musculoskeletal stem cells.

first: Myung


title: Global studies of Genome-Wide DNA Methylation Patterns on porcine  longissimus dorsi muscle

city: hefei

last: Liu

address: No. 40 South Agricultural Road

institute: Anhui Academy of Agricultural Sciences

summary: In order to discover the systematic association between DNA methylation and meat quality, we analyzed the whole-genome DNA methylation patterns in longissimus dorsi muscle by using methylated DNA immunoprecipitation sequencing (MeDIP-seq) in the two breed pigs . Our results showed that a total of 1425 differentially methylated genes (DMGs) were identified (447 hyper-methylated and 978 hypo-methylated) between YY and WH. Gene ontological functional analysis identified 346 terms and 139 genes that may be potential key regulators of pig meat quality.

overall_design: Epigenetic modifications in two breed pigs

first: Lin-Qing

first: Hui

last: Dong

first: Jia-Jun

last: Yang

first: Xiao-Jin

last: Li

first: Kun

last: Qian

first: Jie

last: Zhou

first: Chong-Long

last: Wang


title: Genome-wide identification of differentially methylated promoters and enhancers associated with response to anti-PD-1 therapy in non-small cell lung cancer

first: JAE WON

last: CHO

address: 60 Fenwood Road

department: Neurology

institute: Brigham and Women's Hospital

summary: The purpose of this study is to compare transcriptional profiles of non-small-cell lung cancer between responder patients and non-responder patients during anti-PD-1 treatment.

overall_design: (1) Genomic DNA quantitation: DNA samples were checked for their quality using NanoDrop® ND-2000 UV-Vis Spectrophotometer. The samples were electrophoresed on agarose gels and those with intact genomic DNA, showing no smear upon agarose gel electrophoresis, were selected for subsequent experiment. Intact genomic DNA was diluted to 50 ng/㎕ based on Quant-iT PicoGreen (Invitrogen) quantitation. Concentrations were adjusted according to these results. All prepared samples were bisulfite-converted according to the EZ DNA methylation kit (Zymo Research) protocols. (2) Bisulfite conversion (Zymo EZ DNA methylation kit): For the bisulfite conversion, 600 ng of input gDNA was required. Conversion reagent was added, followed by subsequent incubation in a thermocycler to denature. CT-converted DNA was washed and de-sulfonated with de-sulfonation buffer, after which, the DNA was washed again and eluted with 12-ml elution buffer. (3) Sample amplification and hybridization for BeadChips: The whole-genome amplification process required 250 ng of input bisulfite-converted DNA, MA1, and created a sufficient quantity of DNA (1000× amplification) to be used on a single BeadChip in the Infinium methylation assay (Illumina RPM and MSM). After amplification, the product was fragmented using a proprietary reagent (FMS), precipitated with 2-propanol (plus precipitating reagent; PM1), and re-suspended in formamide-containing hybridization buffer (RA1). The DNA samples were denatured at 95 °C for 20 min, and placed in a humidified container for a minimum of 16 h at 48 °C, allowing CpG loci to hybridize with the 50-mer capture probes. (4) Allele-specific single-base extension and staining on BeadChips: Following hybridization, the BeadChip/Te-Flow chamber assembly was placed on a temperature-controlled Tecan flow-through chamber rack, and subsequent washing, extension, and staining were performed by the addition of reagents to the Te-Flow chamber. For the allele-specific single-base extension assay, primers were extended with a polymerase and labeled nucleotide mix (TEM), and stained with repeated application of STM (staining reagent) and ATM (anti-staining reagent). After completion of staining, the slides were washed with low-salt wash buffer (PB1), immediately coated with XC4, and then imaged in the iScan System. (5) Imaging the BeadChip and data analysis: The iScan System has a two-color (532 nm/658 nm) confocal fluorescent scanner with 0.54-μm pixel resolution. The scanner excited the fluorophores generated during signal amplification/ staining of the allele-specific (one color) extension products on the BeadChips. The image intensities were extracted using Illumina’s GenomeStudio Software.

first: Jae-Won

last: Cho

first: Min

last: Hong

first: Sang-Jun

last: Ha

first: Young-Joon

last: Kim

first: Byoung

middle: C

first: Insuk

last: Lee

first: Hye

middle: R


title: To determine the effects of non-cholesterol based sterol enriched diets on liver gene expression

city: Cincinnati

first: Shailendra

last: Patel

address: 231 Albet Sabin Way

department: Endorcrinology

institute: University of Cincinnati

summary: Xenosterol accumulation in mice deficient in sterolin function leads to significant toxicity, with infertility, decreased body fat accumulation, macrothrombocytopenia, cardiac fibrosis and premature death. The predominant xenosterols are phytosterols derived from the diet which includes a mixture of sitosterol (typically 70%), campesterol (~20%) and stigmasterol (~5-10%), and ergosterol found in fungi. To delineate whether ergosterol or stigmasterol could accumulate in Abcg8 knockout mice and demonstrate any signs of xenosterol toxicity, we designed two diets with one supplemented with ergosterol (>98% purity) and other with stigmasterol (>80% purity) and fed them to Abcg8 knockout mice. Over a 12-week period, both male and female Abcg8 knockout mice fed with either of the diets gained normal amounts of weight, body fat, showed no disturbances in tail-cuff measured blood pressure, and plasma analyses showed no abnormalities of platelet counts or volumes, blood glucose, or plasma cholesterol. Fertility testing showed no abnormalities. While stigmasterol accumulation was observed in the both plasma and tissues, elevated levels of ergosterol was not observed in plasma. However, there was a dramatic increase in plasma brassicasterol, with levels reaching 80 mg/dL in plasma.  Our data concludes that either stigmasterol or  ergosterol or brassicasterol accumulations in Abcg8 Knockout Mice does not account for xenosterol toxicity.

overall_design: Total RNA was isolated from liver and adrenal of 1 mouse in each group. Livers from HS, Ergosterol and Stigmasterol fed mice were compared with those of low sitosterol to determine the differentially expressed genes and underlying pathways responsible for these changes. While in case of adrenals the comparison was exclusively between HS and LS fed ones.

first: Babunageswararao

last: Kanuri

first: Vincent

last: Fong

first: Ranjuna

last: Weerasekera

first: Sithara

last: Ponny


title: Rat muscle derived stem cells treated with fatty acids or cholesterol

city: Torrance

first: Robert

middle: Allen

last: Gelfand

address: 1124 W Carson St

department: surgery/urology

institute: los angeles biomedical research institute

summary:
Rat skeletal muscle derived stem cells were grown in the presence of cholesterol or sodium palmitate in 1 mM glucose (low) or 4 mM glucose (high)
RNA was extracted using the RNeasy Plus Micro kit and analyzed at UCLA by the Affymetrix rat Gene 2.0 ST chip.

overall_design: Muscle derived stem cells from male rats were isolated and grown. Cells in six well plates were subjected to 100 mg/dl cholesterol or sodium palmitate. RNA was isolated using the Qiagen RNeasy Plus Micro kit and subjected to analysis by DNA microarray using the Affymetrix rat Gene 2.0 ST chip.

first: Maryam

last: Masouminia

first: Istvan

last: Kovanecz

first: Nestor

last: Gonzalez-Cadavid

first: Jacob

last: Rajfer

first: Kenny

last: Castro

first: Ruben

last: Salas

first: Leila

last: Loni

first: Dolores

last: Vernet

first: James

last: Tsao


title: Combined Calorie and Protein Restriction in Healthy Kidney Donors Improves Kidney Outcome in Donors and Kidney Transplant Recipients

city: Bilthoven

first: Paul

last: Wackers

address: Antonie van Leeuwenhoeklaan

country: Netherlands

institute: RIVM

summary: A preoperative short-term diet combining calorie and protein restriction diet given during five days before kidney donation and transplantation improves kidney outcome in live kidney donors and kidney transplant recipients, which shows that the benefits of dietary restriction on stress resistance can be induced rapidly in humans, and may be used for clinically relevant endpoints.

overall_design: Gene expression differences of 3 CCPR and 5 control (n=8) male donors and 6 CCPR and 6 control (n=12) female  donors were examined. Biopsies were taken during the transplantation procedure and before implantation after cold-ischemia. Making a total of n=40 samples.

first: Franny

last: Jongbloed

first: Ron

last: de Bruin

first: Piet

last: Beekhof

first: Dennis

last: Hesselink

first: Harry

last: van Steeg

first: Jan

last: Hoeijmakers

first: Martijn

last: Dollé

middle: N

last: Ijzermans


title: Single-cell RNA-seq of embryonic day 17 (E17) mouse skin epithelial cells with or without Mettl3 knockout

city: New York

first: Tom

last: Carroll

department: Bioinformatics

summary: We employed single-cell RNA sequencing to examine effects on steady-state RNA levels induced by METTL3 depletion in mouse skin epithelium during morphogenesis. YFP+ epidermal cells were isolated from E17 embryos with Mettl3 fl/+, Rosa16-YFP fl/+, K14Cre (Ctrl) and Mettl3 fl/fl, Rosa16-YFP fl/+, K14Cre (cKO) genotypes through FACS. Trypan blue staining indicates the ratio of living cells in each sample is >90%. 8000 single cells from each sample is processed with the Chromium™ Single Cell 3’ Library and Gel Bead Kit (V2) (10× Genomics, 120267) to prepare single-cell RNA-seq libraries, which is sequenced on Illumina NextSeq 500 sequenceras 26 × 57 × 8.

overall_design: Examination of transcriptome-wide steady-state RNA expression levels in Ctrl and cKO E17 epithelial cells.

first: Linghe

last: Xi

first: Elaine

last: Fuchs


title: RNA-seq data for lineage negative, Sca1 positive, and cKit positive hematopoietic stem/progenitor cells from H3.3K36M, H3.3K9M, or rtTA control mice

city: Boston

first: Justin

last: Brumbaugh

address: 185 Cambridge St.  CPZN4200

department: Molecular Biology

institute: Massachusetts General Hospital

summary: We analyzed gene expression patterns in LKS cells using RNA-seq after one week of mutant histone induction. We chose this time point to minimize secondary effects and because methylation at both H3K9 and H3K36 is maximally depleted by that time. Compared to rtTA control, 150 and 364 genes were differentially expressed in H3K9M and H3K36M expressing LKS cells, respectively. Principal component analysis indicated that biological replicates were highly consistent and expression profiles for H3K9M, H3K36M, and rtTA control were distinct. In general, gene expression changes following H3K9M induction were subtle, while induction of H3K36M led to more robust differences in expression patterns.

overall_design: There are 9 total samples that include three biological replicates for rtta control, K9M, and K36M.  Each biological replicate represents a single, independent mouse.

first: Hanno

last: Hock

first: Konrad

last: Hochedlinger


title: Time-resolved RNA atlas of early human T cell activation (miRNA)

city: Saarbrücken

first: Tim

last: Kehl

address: Campus E2.1, P.O. box 151150

department: Center for Bioinformatics

institute: Saarland University

summary: MicroRNAs are central regulators of the T cell function. We explored RNA expression profiles over the initial 24 h of human CD4+ T cell activation. We found high similarity in time-resolved miRNA expression courses comparing independent activations and different donors. The detected miRNA expression patterns could be grouped into six classes only, each with a defined time course. MiR-155-5p known for its role in T cell immunity showed the most prevalent expression changes, quantified with an hourly increase of about 60 molecules/cell. As demonstrated for miRNA-155-5p, the analysis of time-resolved miRNA and mRNA expression data allowed to increase the validation rate of predicted miRNA targets to close to 90 %. Combining our time-resolved expression analysis with an absolute quantification of miRNA expression changes, gives new insights into miRNA regulatory networks and indicates the functional dominance of specific miRNAs within the early T cell activation.

overall_design: Early human T cell activation

first: Caroline

last: Diener

first: Martin

last: Hart


title: Single cell analysis in the adult bone marrow vascular niche after irradiation in Vegfc deleted mice

status: Public on Sep 03 2020

city: Nanjing

first: Shentong

last: Fang

address: Longmian Avenue

institute: China Pharmaceutical University

summary: To analyze the role of  VEGF-C in the adult bone marrow vascular niche after irradiation

first: Kari

last: Alitalo


title: Single cell RNA sequencing of aortic CD45+ cells reveals aortic macrophage heterogeneity in mouse atherosclerosis

city: Seoul

first: GooTaeg

last: Oh

address: 52, Ewhayeodae-gil

institute: Ewha Womans University

summary: We have applied single cell RNA sequencing (scRNA-seq) as an unbiased transcriptomics to characterize aortic immune cells in atherosclerosis. The scRNA-seq analysis revealed macrophage heterogeneity at the single-cell level in mouse atherosclerotic aorta.

overall_design: Single cell RNA sequencing of aortic CD45+ leukocytes from atherosclerotic Apoe-/- mice (n=6, western diet for 13 weeks). To characterize myeloid cells, we first extracted cell clusters with expression of canonical macrophage (Adgre1, Csf1r, Fcgr1, and Lyz2) or dendritic cell (Zbtb46, Itgax, and Flt3) markers and confidently defined 3,002 myeloid cells.

first: Sejin

last: Jeon

first: Tae

last: Kim

first: Sung

last: Park

first: Woong-Yang

first: Hae-Ock

last: Lee

first: Nayoung


title: PTEN-dependent regulation of genome-wide transcription during metabolic stress

first: Ata

last: Abbas


title: Angiotensin IV attenuates diabetic cardiomyopathy via suppressing FoxO1-induced excessive autophagy, apoptosis and fibrosis

city: jinan

first: meng

last: zhang

address: Jinan 250012, Shandong, China

institute: Shandong University

summary:
It is well known that rennin-angiotensin-aldosterone system (RAAS) plays a critical role in the development of diabetic cardiomyopathy. The present study was aimed to clarify the role of a novel member of RAAS, angiotensin IV (Ang IV) and its downstream mediator forkhead box protein O1 (FoxO1), in the pathogenesis of diabetic cardiomyopathy. In vivo, diabetic mice were treated with low-, medium- and high-dose Ang IV, AT4R antagonist divalinal, FoxO1 inhibitor AS1842856 (AS), or their combinations. In vitro, cardiomyocytes and cardiofibroblasts were treated with different concentrations of glucose, low-, medium- and high-dose Ang IV, divalinal, FoxO1-overexpression plasmid (FoxO1-OE), AS or their combinations.
The results showed that Ang IV treatment dose-dependently attenuated left ventricular dysfunction, fibrosis, and myocyte apoptosis in diabetic mice. Besides, enhanced autophagy and FoxO1 protein expression by diabetes were dose-dependently suppressed by Ang IV treatment. However, these cardioprotective effects of Ang IV were completely abolished by divalinal administration. Bioinformatics analysis revealed that the differentially expressed genes were enriched in autophagy, apoptosis, and FoxO signaling pathways among control, diabetes, and diabetes+high-dose Ang IV groups. Similar to Ang IV, AS treatment ameliorated diabetic cardiomyopathy in mice. In vitro, high glucose stimulation increased collagen expression, apoptosis, overactive autophagy flux and FoxO1 nuclear translocation in cardiomyocytes, and upregulated collagen and FoxO1 expression in cardiofibroblasts, which were substantially attenuated by Ang IV treatment. However, these protective effects of Ang IV were completely blocked by the use of divalinal or FoxO1-OE, and these detrimental effects were reversed by the additional administration of AS. In summary, Ang IV treatment dose-dependently attenuated left ventricular dysfunction and remodeling in a mouse model of diabetic cardiomyopathy, and the mechanisms involved stimulation of AT4R and suppression of FoxO1-mediated fibrosis, apoptosis, and overactive autophagy.

overall_design: Myocardial tissues were isolated from mice of negative control, diabetes mellitus, and diabetes mellitus+high-dose angiotensin IV groups;  four mice in each group

first: Meng

last: Zhang

first: Yun


title: Structure of LINC00511-siRNA-conjugated nanobubbles and improvement of cisplatin sensitivity on triple negative breast cancer

city: Harbin

first: Bolin

last: Wu

state: Heilongjiang Province

address: No.150, Haping Road, Nangang District

institute: Harbin Medical University Cancer Hospital

summary: In the present study, we prepared a novel theranostic agent loaded with LINC00511-siRNA, which not only can serve as the siRNA delivery platform but also as the UCA for contrast enhanced ultrasound (CEUS). Combined with low-frequency ultrasound exposure, the novel theranostic agent achieves the controlled release of LINC00511-siRNA and facilitates the transfection of LINC00511-siRNA. Furthermore, in order to investigate how LINC00511 affects drug resistance of TNBC, we conducted the RNA-Seq to gain new insights into the mechanisms of drug sensitivity.

overall_design: MDA-MB-231 cells and MDA-MB-231/CDDP cells were used for the research. The cells were transfected using the nanoparticles with the aid of low-frequency ultrasound(LFUS) exposure. Two groups of pre-transfection cells and two groups of post-transfection cells were analyzed using high-throughput RNA sequencing.

first: Yanchi

last: Yuan

first: Erjing

last: Li

first: Hui

last: Jing

first: Wen

last: Cheng


title: Landscape of open chromatin regions in Arabidopsis upon exposure to abiotic stresses

city: Brno

first: Vivek

middle: Kumar

last: Raxwal

address: building A26, Kamenice 753/5

department: Plant Molecular Biology

country: Czech Republic

institute: Central European Institute of Technology

summary: The goal of the study is to illustrate how various abiotic stresses reshape the chromatin architecture of the Arabidopsis genome to fine tune gene expression.

overall_design: 10 day old  Arabidopsis seedlings were subjected to heat (38C for 6h), cold (4C for 24h), salt (250mM Nacl for 6h) and drought(1h) and open chromatin was extracted using FAIRE-seq and Dnase-seq. RNA-seq was also performed from same tissue.

first: Manu

last: Agarwal


title: Global gene expression profiles of GATA6-AS1 knockdown cultures and control shRNA cultures at cardiomyocyte differentiation day 2 and day 5

city: ATLANTA

first: Dong

state: GEORGIA

address: 2015 UPPERGATE DR

department: Pediatrics

institute: Emory University

summary:
Methods: RNA-seq libraries were prepared using the Illumina TruSeq technology. The libraries were quantified and samples were multiplexed in each lane of the flowcell. Cluster generation was performed and then sequenced on the Illumina HiSeq2500 system. Reads were mapped on the Human Genome Reference (GRCh38) and normalized expression table was generated.
Results: At day 2, GATA6-AS1 knockdown resulted in 499 down-regulated and 1086 up-regulated differentially expressed genes (DEGs, set at -log10 p-value = 1.996; log2 changes <-1 or >1). At day 5, GATA6-AS1 knockdown resulted in 1457 down-regulated and 1166 up-regulated DEGs
Conclusions: Data demonstrate GATA6-AS1 knockdown inhibits mesoderm induction and cardiomyocyte differentiation in hPSCs.

overall_design: RNA-sequencing analysis was performed to compare the genome-wide gene expression profile of GATA6-AS1 knockdown cultures with that of control shRNA cultures at cardiomyocyte differentiation day 2 and day 5.

first: Rajneesh

last: Jha

first: Katherine

middle: E

last: Ferguson

first: Gregory

last: Gibson

first: Chunhui

last: Xu


title: Transcriptome sequencing of strong, middle and weak gluten strength wheat

city: Zhengzhou

first: Jinshui

last: Wang

state: Henan

address: No. 100 Lianhua street

institute: Henan university of technology

summary: Here is reported the first study of transcriptome analyses using the Illumina HiSeq 4000 platform for three kinds of wheat (G represents Strong gluten wheat, Z represents middle gluten wheat,R represents weak gluten wheat). The variation of wheat varieties with different gluten content is mainly shown in the content of gluten, flour is divided into high gluten powder ( > 30%), medium gluten powder (26%-30%) and low gluten powder ( < 20%), according to the wet gluten content. In total, over 102.6 Gb clean reads were produced and 114, 621 unigenes were assembled; more than 59,085 unigenes had at least one significant match to an existing gene model. Differentially expressed gene analysis identified 2339 and 2600 unigenes which were expressed higher or lower among strong gluten, middle gluten and weak gluten wheat. After functional annotation and classification, three dominant pathways including protein isomerase, antioxidase activity and energy metabolism, and 410 unigenes related to gluten strength polymerization of wheat were discovered. In strong-gluten wheat, low molecular weight subunit content is higher than weak-gluten wheat, and the activity of cysteine synthase and isomerase is increased, which may promote the cross-linking of low molecular weight protein to high molecular weight protein. Meanwhile, POD enzyme strengthens gluten network and CAT enzyme affects gluten polymerization, along with higher ATPase activity, which will provides energy for protein polymerization reaction in comparison of strong-gluten wheat and weak-gluten wheat. The accuracy of these RNA-seq data was validated by qRT-PCR analysis. These data will extend our knowledge of quality characteristics of wheat and provide a theoretical foundation for molecular mechanism research of wheat.

overall_design: Different gluten strength wheat mRNA profiles was generated by RNA-seq, with three biological repeats, using Illumina 4000 platform

first: Qi

first: Guiling

first: Feng

last: Jia


title: Investigating the impact of chronic stress on behavior, brain plasticity and hippocampal transcriptome in the ewe.

status: Public on Sep 02 2020

city: Nouzilly

first: Hugues

last: Dardente

address: INRA

department: INRAE - Centre Val de Loire

institute: INRAE

summary: Using NGS, we report the impact of chronic stress on the transcriptome of the dorsal and ventral hippocampus in the ewe. This study identifies specific transcriptional signatures of the dorsal VS ventral hippocampus (>220 DEG) and demonstrates a very limited transcriptional impact of stress, exclusively restricted to the dorsal hippocampus (<10 DEG).

overall_design: Comparison of the transcriptomes of dorsal and ventral hippocampus (groups thereafer denoted "DG for dorsal" and "vHip for ventral"), in control ewes and ewes submitted to chronic stress (groups thereafter denoted "C for Control" and "St for Stress"). 6 ewes per group (C & St) / RNA individually extracted from DG and vHip of all 12 ewes / Equimolar mixes of n=2 per condition (C & St) and structure (Dg & vHip) --> Triplicates for  each condition and structure.

first: Matthieu

last: Keller


title: To understand the effects of global Cyp27a1 and Cyp3a4 gene deletions on bile acid synthesis

first: Shailendra

last: Patel

summary: The overall goal of the project is to generate a potential mouse model for Cerebrotendinous Xanthomatosis (CTX), a rare autosomal recessive bile synthesis disorder. Patients with this disorder were found to have mutations in Cyp27a1 gene, which results in deficiency of the enzyme sterol 27-hydroxylase. This loss leads to inhibition of cholesterol being converted into bile acids. Global deletion of Cyp27a1 in mice showed none of the CTX phenotype and do not accumulate bile alcohols. As livers of these mice show tremendous upregulation of Cyp3a4, so we hypothesized that it may have "detoxified’ the intermediates and therefore presented a normal phenotype. For answering this, we made double KO mice with Cyp27a1-/- and Cyp3a cluster (Cyp3a13/57/16/41/44/11/25/59) and found no phenotype till 6 months of age.

overall_design: Total RNA was isolated from liver of one mouse in each group. Cyp27a1 KO, Cyp3a4 KO, Cyp27a1-Cyp3a4 DKO mouse livers were compared to WT mouse.

first: Babunageswararao

last: Kanuri

first: Vincent

last: Fong

first: Ranjuna

last: Weerasekera

first: Sithara

last: Ponny


title: Genome-wide maps of chromatin state in buffalo fibroblasts with different cloning efficency

city: Nanning

first: Zhiqiang

address: No. 6 TaoYuan Rd.

institute: Guangxi Academy of Medical Sciences and the People’s Hospital of Guangxi Zhuang Autonomous Region

summary:
We report the distribution of histone acetylation at global genomic level in donor cells with different cloning efficiency. By obtaining over 40 million clean reads from immunoprecipitated DNA, we generated genome-wide chromatin-state maps of two buffalo fibroblast cell lines. We find that high cloning efficiency BFFs also had more regions enriched with H3K9ac sites near to the upstream of genes related to glycolysis. This study provides a framework for the application of comprehensive chromatin profiling towards donor cells with different cloning efficiency.
We report the landscape of chromatin accessibility in donor cells with different cloning efficiency by transposase-accessible chromatin sequencing (ATAC-seq). We found that the landscape of chromatin accessibility was similar in both BFF1 and BFF2. Most loci defined by ATAC-seq was abundant just on the upstream of transcription start sites (TSS), but the signals in BFF2 were stronger than BFF1. GO analysis showed that most differential loci annotated genes were involved in developmental process, cellular process and metabolic process. This study provides a framework for the application of landscape of chromatin accessibility towards donor cells with different cloning efficiency.

overall_design: Examination of H3K9Ac modification and chromatin accessibility  in buffalo fibroblasts with different cloning efficency

first: Deshun

last: Shi

first: Chan

last: Luo

first: Fenghua

last: Lu


title: Expression Data from fracture calli of alpha-CGRP deficient mice

city: Hamburg

first: Thorsten

last: Schinke

address: Martinistr. 52

department: Department of Osteology and Biomechanics

institute: University Medical Center Hamburg-Eppendorf

summary:
alpha-CGRP is a neuropeptide that is also expressed in the fracture callus during bone regeneration. Our aim was to evaluate the role of alpha-CGRP in the context of fracture healing. Therefore we investigated the effect of alpha-CGRP deficiency on fracture callus formation.
We used microarray analysis to compare the global gene expression of fracture calli from alpha-CGRP deficient mice and WT mice.

overall_design: 2 sample sets with 3 independent repetitions for each sample set were compared. 1st sample set: Fracture callus from WT mice. 2nd sample set: Fracture callus from alpha-CGRP deficient mice.

first: Timur

middle: A

last: Yorgan

first: Jessika

last: Appelt

first: Lana

last: Rosenthal

first: Johannes


title: Hemolysis in the spleen facilitates erythrocyte turnover

city: Amsterdam

first: Benjamin

last: Nota

state: Noord Holland

address: Plesmanlaan 125

department: Research Facilities

institute: Sanquin

summary: Red pulp macrophages of the spleen mediate daily turnover of billions of senescent erythrocytes. However, the exact molecules and mechanisms involved in sequestration of senescent erythrocytes, their recognition and ultimately their turnover remain unclear and are currently subject to debate. In this study we provide evidence that the splenic environment is of substantial importance in facilitating erythrocyte turnover through induction of haemolysis. Detailed characterization of human spleen and its red pulp macrophages lead to the identification of a population of erythrocytes devoid of haemoglobin without fully disintegrating, so-called erythrocyte ghosts. By in-vivo imaging and transfusion experiments we established that senescent erythrocytes are subject to haemolysis specifically within the spleen. We show that aged erythrocytes are captured by the extracellular matrix within the red pulp of the spleen and that their retention under low shear conditions is key in driving haemolysis.  In contrast to senescent erythrocytes, the erythrocyte ghost shells were found to be prone to recognition and breakdown by red pulp macrophages. As such, these data put forward haemolysis as an efficient mechanism for the turnover of senescent erythrocytes which alters our current understanding on how erythrocyte turnover is regulated.

overall_design: RNA was isolated from 3 different populations (blood monocytes, spleen monocytes,and spleen macrophages) from 3 donors.

first: T

last: Klei

first: B

first: R

last: van Bruggen


title: Intracellular Cryptococcus neoformans Disrupts the Transcriptome Profile of M1- and M2-Polarized Host Macrophages

city: Murfreesboro

first: Rebecca

middle: L

last: Seipelt-Thiemann

state: TN

address: 1301 E Main St Box 60

institute: Middle Tennessee State University

summary: Macrophages serve as a first line of defense against infection with the facultative intracellular pathogen, Cryptococcus neoformans (Cn). However, the ability of these innate phagocytic cells to destroy ingested Cn is strongly influenced by polarization state with classically (M1) activated macrophages better able to control cryptococcal infections than alternatively (M2) activated cells. While earlier studies have demonstrated that intracellular Cn minimally affects the expression of M1 and M2 markers, the impact on the broader transcriptome associated with these states remains unclear. To investigate this, we used an in vitro cell culture model of intracellular infection together with RNA sequencing-based transcriptome profiling to measure the impact of Cn infection on gene expression in both polarization states. The gene expression profile of both M1 and M2 cells was extensively altered to become more like naive (M0) macrophages. Gene ontology analysis suggested that this involved changes in the activity of the Janus kinase-signal transducers and activators of transcription (JAK-STAT), p53, and nuclear factor-κB (NF-κB) pathways. Analyses of the principle polarization markers at the protein-level also revealed discrepancies between the RNA- and protein-level responses. In contrast to earlier studies, intracellular Cn was found to increase protein levels of the M1 marker iNos. In addition, we identified common gene expression changes that occurred post-Cn infection, independent of polarization state. This included upregulation of the transcriptional co-regulator Cited1, which was also apparent at the protein level. These changes constitute a transcriptional signature of macrophage Cn infection and provide new insights into how Cn impacts gene expression and the phenotype of host phagocytes.

overall_design: Naïve macrophage (naïve, M0, overall control), M1-polarized mock-infected (M1 control), M1-polarized pathogen-infected, M2-polarized mock-infected (M2 control), M1-polarized pathogen-infected were compared in a pairwise manner.  Samples were used in triplicate, but analyses supported removal of 1 replicate for all treatments except M2-polarized pathogen-infected

first: David

last: Nelson

first: Erin

last: McClelland


title: Myosin 7b is a regulatory long noncoding RNA (lncMYH7b) in the human heart [adenovirus either expressing GFP or MYH7b_sp]

city: Boulder

first: Lindsey

middle: Joe

last: Broadwell

state: CO

address: 3415 Colorado Avenue, Apt, Suite, Bldg. (optional)

department: Biochemistry

institute: University of Colorado Boulder

summary: We have discovered that the alternative splice form of MYH7b is a regulatory non-coding RNA in the human heart. This dataset contains a forced expression of a peptide encoded by a short ORF at the beginning of the alternate splice form (MYH7b_sp) in cardiomyocytes.

overall_design: Human induced pluripotent stem cells differentiated to cardiomyocytes and infected with an adenovirus either expressing GFP or MYH7b_sp.


title: Gene expression analysis of fibroblasts of systemic sclerosis patients silenced for lncRNA H19X

city: Schlieren

first: Elena

last: Pachera

address: Wagistrasse 14

institute: USZ

summary: LncRNA H19X was silienced in dermal fibroblats of systemic sclerosis patients with antisense oligonuclotides.  The hypothesis tested in the present study was that H19X is an important factor in the development of TGFb-driven fibrosis. Results provide important information about the role H19X in fibroblasts in particolar on extracellular matrix production and cell cycle regulation.

overall_design: Total RNA obtained from dermal fibroblssts of systemic sclerosis patients transfected with antisense oligonucleotide targeting H19X for 120h compared to fibroblasts transfected with scrambled control. H19X silenced fibroblasts and scramble fibroblasts were treated with TGFb for the last 72h of transection.

first: Oliver

last: Distler


title: Plasma protein-profiling to predict distant metastasis in locoregionally advanced nasopharyngeal carcinoma (LA-NPC)

city: Guangzhou

first: Jun

last: Ma

address: 651 Dongfeng Road East

institute: Sun Yat-sen University Cancer Center

summary: Plasma protein expression patterns can be used as prognostic biomarkers in various types of cancer. We aimed to identify a protein-based signature for distant metastatic risk assessment in patients with locoregionally advanced nasopharyngeal carcinoma (LA-NPC).

overall_design: Using Raybiotech Quantibody Human Kiloplex Proteomics Array (QAH-CAA-X00), we detected 1000 plasma protein expression between 8 LA-NPC plasma samples with distant metastasis after radical treatment  and 8 LA-NPC plasma samples without distant metastasis after radical treatment to identify distant metastasis-related proteins.

first: Y

last: Liang

first: N

last: Liu

middle: Q

last: Li

first: J


title: Gene expression profiling of colorectal cancer cell lines after treatment with talazoparib

city: Oslo

first: Anita

last: Sveen

address: P.O.Box 4953, Nydalen

department: Department of Molecular Oncology

country: Norway

institute: Institute for Cancer Research, Oslo University Hospital

summary: We have performed post-treatment gene expression profiling of cell lines to analyze response mechanisms to PARP inhibition.

overall_design: Four colorectal cancer cell lines were treated with talazoparib at two different concentrations or DMS for control.

first: Ina

last: Eilertsen

first: Ragnhild

last: Lothe


title: The differential expression of microRNA in human ESCC cells after JMJD3 knockdown

first: Shuman

address: No. 651, Dongfeng Road East

summary: Microarrays were used to screen differentially expressed microRNAs in JMJD3 silencing cells and their control cells  .

platform_organism:
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design: In the experiments，we knock down JMJD3 expression in ESCC cell line K30 by shRNA, and screen for microRNAs that were up- or down-regulated simultaneously in these cells compared to the control cells.