title: Longitudinal shear stress response in human aortic endothelial cells to atheroprone and atheroprotective conditions

first: Shankar

last: Subramaniam

address: 9500 Gilman Dr MC 0412

department: Bioengineering

summary: The goal of this study was to find longitudinal transcriptional response of human aortic endothelial cells (HAECs) to pulsatile shear (PS) and oscillatory shear (OS) and compare them with the responses in human umbilical vein endothelial cells (HUVECs) [GSE103672]. PS is associated with an atheroprotective endothelial phenotype, while OS is associated with an atheroprone endothelial phenotype. Using RNASeq method (single-ended 75-bp sequencing on Illumina Hi-seq 4000 instrument), we measured the transcriptional response at 3 time-points (1, 4, and 24 hrs) under PS and OS conditions. Measurements were also taken under static condition (ST, no flow) at t = 0 hr. Three replicates were used for each condition/time-point. Our results indicate that the responses of HAECs and HUVECs are qualitatively similar for endothelial-function relevant genes and several important pathways with a few exceptions, thus demonstrating that HUVECs can be used as a model to investigate the effects of shear on arterial ECs, with some reservations. Our findings show that HAECs exhibit an earlier response or faster kinetics as compared to HUVECs. The comparative analysis in this study offers new insights into the mechanisms of common and disparate stress responses across these two endothelial cell types.

overall_design: HAECs (Cell Applications Cat# 03405A) were cultured in endothelial growth medium (Cell applications Cat#211-500) with 100 U/mL penicillin-streptomycin. Pulsatile shear (PS) and oscillatory shear (OS) flows were applied to ECs with shear stresses of 12 ± 4 dyn/cm2 and 0.5 ± 4 dyn/cm2, respectively. Samples for RNA sequencing analysis were collected at 1, 4, and 24 hrs after exposure to shear. For static condition (ST), samples were collected at t = 0 hr under no flow. Three biological replicates were collected for each time point and for each shear condition. Total RNA was extracted from ECs, processed and sequenced on the Illumina HiSeq 4000 75-bp single-end reads platform.

first: Shu

last: Chien

first: Julie

middle: Y

first: John

last: Shyy

first: Zhen

last: Chen

first: Mano

middle: R

last: Maurya

first: Shakti

last: Gupta


title: Serine biosynthesis is a metabolic vulnerability in FLT3-ITD-driven acute myeloid leukaemia [iFLT3-ITD-Depletion_In VIVO]

status: Public on Jan 13 2021

city: Boston

first: Stefan

last: Bjelosevic

state: MA

address: 450 Brookline Ave

department: Department of Pediatric Oncology

country: USA

institute: Dana-Farber Cancer Institute

type: Expression profiling by high throughput sequencing

summary: We performed 3'-RNA-Sequencing on RNA isolated from murine MLL-AF9/iFLT3-ITD cells where the inducible iFLT3-ITD transgene had been depleted for 48 hours compared to non-depleted conditions in vivo to perform differential gene expression analysis.

overall_design: 1 million MLL-AF9/iFLT3-ITD cells were transplanted into NSG mice via tail vein IV injection. Mice were fed doxycycline-containing chow and water to maintain inducible iFLT3-ITD transgene expression, and leukaemias were allowed to engraft for 96 hours. Half of the cohort of mice (n=3) was then placed on regular chow and water to deplete inducible iFLT3-ITD transgene expression for 48 hours, while the other half of the cohort (n=3) remained on doxycycline-containing chow and water to maintain inducible iFLT3-ITD transgene expression as a control. Mice were all culled after 48 hours and leukaemic cells sorted from the bone marrow. Total RNA was isolated from cells and 3'-RNA libraries prepared for next-generation sequencing.

first: Emily

last: Gruber

first: Kristin

middle: K

last: Brown

first: Ricky

middle: W

last: Johnstone


title: Germline inherited small RNAs facilitate the clearance of untranslated maternal mRNAs in C. elegans embryos (Degradome-seq)

city: Paris

first: Germano

last: Cecere

address: Institut Pasteur, 28 Rue Du Docteur Roux, Batiment Monod, 4eme Etage

department: Development and stem cell biology

country: France

institute: Institut Pasteur

summary: Inheritance and clearance of maternal mRNAs are two of the most critical events required for animal early embryonic development. However, the mechanisms regulating this process are still largely unknown. Here, we show that together with maternal mRNAs, C. elegans embryos inherit a complementary pool of small non-coding RNAs that facilitate the cleavage and removal of hundreds of maternal mRNAs. These antisense small RNAs are loaded into the maternal catalytically-active Argonaute CSR-1 and cleave complementary mRNAs no longer engaged in translation in somatic blastomeres. Induced depletion of CSR-1 specifically during embryonic development leads to embryonic lethality in a slicer-dependent manner and impairs the degradation of CSR-1 embryonic mRNA targets. Given the conservation of Argonaute catalytic activity, we propose that a similar mechanism operates to clear maternal mRNAs during the maternal-to-zygotic transition across species.

overall_design: 4 samples were analyzed by Degradome-seq, including replicates (sample name contains 'rep').

first: Piergiuseppe

last: Quarato

first: Meetali

last: Singh

first: Eric

last: Cornes

first: Blaise

last: Li

first: Celine

last: Didier

first: Loan

last: Bourdon


title: Germline inherited small RNAs facilitate the clearance of untranslated maternal mRNAs in C. elegans embryos (sRNA-IP-Seq)

type: Non-coding RNA profiling by high throughput sequencing

overall_design: 12 samples (6 IP-input paired samples) were analyzed by sRNA-seq.

first: Florian

last: Mueller


title: Germline inherited small RNAs facilitate the clearance of untranslated maternal mRNAs in C. elegans embryos (Ribo-Seq)

overall_design: 7 samples were analyzed by Ribo-seq, including replicates (sample name contains 'rep').


title: Germline inherited small RNAs facilitate the clearance of untranslated maternal mRNAs in C. elegans embryos (RNA-seq)

overall_design: 33 samples were analyzed by RNA-seq, including replicates (sample name contains 'rep').


title: Germline inherited small RNAs facilitate the clearance of untranslated maternal mRNAs in C. elegans embryos

overall_design: Refer to individual Series


title: Germline inherited small RNAs facilitate the clearance of untranslated maternal mRNAs in C. elegans embryos (GRO-Seq)

overall_design: 6 samples were analyzed by GRO-seq, including replicates (sample name contains 'rep').


title: Germline inherited small RNAs facilitate the clearance of untranslated maternal mRNAs in C. elegans embryos (sRNA-Seq)

overall_design: 4 samples were analyzed by sRNA-seq, including replicates (sample name contains 'rep').


title: Single cell(sc) RNAseq of wild type (WT) mouse lung

status: Public on Jan 12 2021

city: Kansas City

first: Venkatesh

last: Sampath

state: MO

address: 2401 Gillham Road

department: Neonatology

institute: Children's Mercy Hospital

summary: We report the application of scRNA sequencing for high-throughput profiling of gene epxression in 40 unsupervised lung cell clusters

overall_design: scRNAseq was done on 5677 lung cells from three WT mouse

first: Sheng

last: Xia

first: Heather

last: Menden

first: Jeffrey

last: Johnston

first: Daniel

last: Louiselle


title: In vivo evaluation of the 94 GHz-chronic exposure effect on Hairless rat skin gene expression [C2]

city: Fougeres

first: Denis

last: Habauzit

address: 10 B rue Claude Bourgelat

department: TC unit

institute: ANSES

summary: Millimeter waves are new broadband frequencies that start to be used in several applications such as future wireless communications, medical, and also non-lethal weapons (the 94-GHz band in Active Denial Systems). However, low information are available on their potential effects on human. These frequencies belong to radiofrequency and they have the property to be stopped by the first layer of the skin. Therefore our studies have aimed to evaluate 94 GHz effects on skin cells whole gene expression. Chronic long term 94 GHz MMW exposures were performed on two rat populations constituted by 17 young animals and 14 adult ones. Each group of animals were equally split in Exposed and Sham exposed subgroup. This two independent exposure experiments were conducted during 5 months, with rats exposed 3 h per day, 3 days per week, to an incident power density of 10 mW/cm², which correspond to twice the ICNIRP limit of exposure for professional. At the end of the experiment, skin explant were taken and RNA were extracted. Then, the modification of the whole gene expression were analyzed with gene expression microarray. Without modification of the animal’s temperature, long term chronic 94 GHz-MMW exposure did not significantly modify the rat’s skin gene expression on both young and adult rats.

overall_design: Animals were exposed (6 animals) or Sham exposed (8 animals) to 94 GHz exposure 3h/d, 3 days/w during 5.5 months. At the end of the exposure, animals were sacrified and skin explants were recovered and conserved in RNA later up to the analysis.

first: Guillaume

last: Nugue

first: Frédéric

last: Bourbon

first: Catherine

last: Martin

first: Flavia

last: Del Vecchio

first: Séverine

last: Maunoir-regimbal

first: Thomas

last: Poyot

first: Marco

last: Valente

first: Rachid

last: Jaoui

first: David

last: Crouzier

first: Yves

last: Le Dréan

first: Jean

middle: C

last: Debouzy


title: In vivo evaluation of the 94 GHz-chronic exposure effect on Hairless rat skin gene expression [C1]


title: Gene expression difference between normal and Pten-deficient 5-HT neurons

city: Shanghai

first: ling

last: chen

address: 1239 siping Rd

country: China

institute: Tongji university

summary: Purpose: compare the gene expression diffenrence between wildtype and Pten-deficient 5-HT neurons. Methods: 5-HT neurons cell body are collected from acut brain slice with patch electrodes.  Pten-deficient 5-HT neurons are obtained from Pet1-Cre; Ptenflox/flox mice. Normal 5-HT neurons are obtained from littermate wildtype mice. Reverse transcription was performed using a SMARTer Ultra Low RNA Kit (Clontech, Japan). Single-cell cDNA was amplified using an Advantage 2 PCR Kit (Clontech, Japan). Purified cDNA from individual cells were sequenced using the Illumina HiSeq 2000 platform. Sequence reads were aligned to the mouse mm9 genome using TopHat. Differential expression analysis was performed using DESeq (Bioconductor) followed by Student t-test. Genes were considered differentially expressed if p<0.05. Results: The somatic compartments of individual YFP-labeled 5-HT neurons in acute brain slices were harvested for RNA sequencing. Each of the cells had  2 X 107 reads mapping back to the mouse genome when analyzed using the RUM mapping program. We found 859 genes (381 up-regulated, 478 down-regulated) were differentially expressed in Pten-deficient 5-HT neurons. Gene ontology of differentially expressed genes revealed the biological processes up-regulated and down-regulated in Pten-deficient 5-HT neurons. The absence of Pten in 5-HT neurons was associated with significantly increased expression of genes involved in the regulation of axonal extension and cell size, cell morphogenesis and cytoskeleton organization

overall_design: mRNA profile and comparison between wildtype and Pten-deficient 5-HT neurons.

first: Kun-Shan

last: Zhang

first: Li-Qiang

last: Zhou

first: Si-Guang

last: Li


title: Melanoma subpopulations that rapidly escape MAPK pathway inhibition incur DNA damage and rely on stress signalling

city: Boulder

first: Chengzhe

last: Tian

state: Colorado

address: 3415 Colorado Ave, JSC Biotech Building, Spencer Lab

department: Biochemistry

institute: University of Colorado Boulder

summary: Despite the increasing number of effective anti-cancer therapies, successful treatment is limited by the development of drug resistance. While the contribution of genetic factors to drug resistance is undeniable, little is known about how drug-sensitive cells first evade drug action to proliferate in drug. Here we track the responses of thousands of single melanoma cells to BRAF inhibitors and show that a subset of cells escapes drug via non-genetic mechanisms within the first three days of treatment. Cells that escape drug rely on ATF4 stress signalling to cycle periodically in drug, experience DNA replication defects leading to DNA damage, and yet out-proliferate other cells over extended treatment. Together, our work reveals just how rapidly melanoma cells can adapt to drug treatment, generating a mutagenesis-prone subpopulation that expands over time.

overall_design: Single-cell RNA-sequencing of A375 melanoma cells treated with and without 1 μM dabrafenib for 72 hr

first: Chen

last: Yang

first: Timothy

middle: E

last: Hoffman

first: Nicole

last: Jacobsen

first: Sabrina

middle: L

last: Spencer


title: The role of XIST in X-inactivation maintenance in B cells

city: Stanford

first: Howard

last: Chang

state: California

address: 269 Campus Drive

institute: Stanford University

type: Genome binding/occupancy profiling by high throughput sequencing

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


title: The role of XIST in X-inactivation maintenance in B cells (CRISPRi screen)

summary: The long noncoding RNA (lncRNA) XIST establishes X chromosome inactivation (XCI) in female cells in early development and thereafter is thought to be largely dispensable. Here we show XIST is continually required in adult human B cells to silence a subset of X-linked immune genes such as TLR7.  XIST-dependent genes lack promoter DNA methylation and require continual XIST-dependent histone deacetylation. XIST RNA-directed proteomics and CRISPRi screen reveal distinctive somatic cell-specific XIST complexes, and identify TRIM28 that mediates Pol II pausing at promoters of X-linked genes in B cells. XIST dysregylation, reflected by escape of XIST-dependent genes, occurs in CD11c+ atypical memory B cells across single-cell transcriptome data in patients with female-biased autoimmunity and COVID-19 infection. XIST inactivation with TLR7 agonism suffices to promote isotype-switched atypical B cells. These results suggest cell-type-specific diversification of lncRNA-protein complexes increase lncRNA functionalities, and expand roles for XIST in sex-differences in biology and medicine.

overall_design: CRISPRi screen for top XIST co-factors in dCas9-Krab+ GM B cells on allelic expression of TLR7. Targeted TLR7 amplicons containing SNP were amplified by nested PCR from cDNA of individual gRNA-perturbed cells and indexed with different barcodes and mixed for miseq sequencing.

first: Bingfei

last: Yu

first: Yanyan

last: Qi

first: Rui

first: Quanming

last: Shi

first: Ansuman

middle: T

last: Satpathy


title: The role of XIST in X-inactivation maintenance in B cells (ChIP-seq, Cut&Run))

overall_design: ChIP-seq of H3K27ac in XIST_WT and XIST KO GM12878 (GM) B cells. ChIP-seq of Trim28 in GM B cells. Cut&Run of H3K27me3 in XIST_WT and XIST KO GM B cells. Cut&Run of H3K9me3 in GM B cells.


title: JNK1 Signaling Downstream of the EGFR Pathway Contributes to Imiquimod-induced Skin Inflammation

city: Gosselies

first: Stanislas

last: Goriely

address: Rue Adrienne Bolland 8

country: Belgium

institute: Université Libre de Bruxelles


title: JNK1 Signaling Downstream of the EGFR Pathway Contributes to Imiquimod-induced Skin Inflammation (ATAC-seq)

summary: c-Jun N-terminal protein kinase 1 (JNK1) is involved in multiple biological processes but its implication in inflammatory skin diseases is still poorly defined. Herein, we studied the role of JNK1 in the context of imiquimod-induced skin inflammation. We observed that constitutive ablation of JNK1 reduced imiquimod-induced acanthosis and expression of inflammatory markers. Conditional deletion of JNK1 in myeloid cells led to reduced skin inflammation, a finding that was associated with impaired Aldara®-induced inflammasome activation in vitro. Next, we evaluated the specific role of JNK1 in epidermal cells. We observed a reduced imiquimod-induced acanthosis and keratinocytes proliferation despite similar levels of inflammatory markers. Transcriptomic and epigenomic analysis of keratinocytes revealed the potential involvement of JNK1 in the EGFR signaling pathway. Finally, we show that inhibition of the EGFR pathway reduced imiquimod-induced acanthosis. Taken together, these data indicate that JNK1 plays a dual role in the context of psoriasis by regulating the production of inflammatory cytokines by myeloid cells and the sensitivity of keratinocytes to EGFR ligands. These results suggest that JNK1 could represent a valuable therapeutic target in the context of psoriasis.

overall_design: 6 keratinocytes mouse samples:  2 conditions (Mapk8-WT/Mapk8-KO) x 2 replicates

first: Aurore

last: Le

first: Abdulkader

last: Azouz

first: Séverine

last: Thomas

first: Nicolas

last: Istaces

first: Muriel

last: Nguyen


title: JNK1 Signaling Downstream of the EGFR Pathway Contributes to Imiquimod-induced Skin Inflammation (RNA-seq)

overall_design: 6 keratinocytes mouse samples:  2 conditions (Mapk8-WT/Mapk8-KO) x 3 replicates


title: Developmental trajectory of oligodendrocyte progenitor cells in the human brain revealed by single cell RNA sequencing

city: Montreal

first: Jack

last: Antel

state: Quebec

address: 3801 Rue University

department: Neurology and Neurosurgery

country: Canada

institute: McGill University

summary: Characterizing the developmental trajectory of oligodendrocyte progenitor cells (OPC) is of great interest given the importance of these cells in the remyelination process. However, studiescof human OPC development remain limited by the availability of whole cell samples and material that encompasses a wide age range, including time of peak myelination. In this study, we apply single cell RNA sequencing to viable whole cells across the age span and link transcriptomic signatures of oligodendrocyte-lineage cells with stage-specific functional properties. Cells were isolated from surgical tissue samples of second-trimester fetal, 2 year old pediatric, 13 year old adolescent, and adult donors by mechanical and enzymatic digestion, followed by percoll gradient centrifugation. Gene expression was analyzed using droplet-based RNA sequencing (10X Chromium). Louvain clustering analysis identified 3 distinct cellular subpopulations based on 5613 genes, comprised of an early OL precursor cell (e-OPC) group, a late OPC group (l-OPC), and a mature OL (MOL) group. Gene ontology terms enriched for e-OPCs were cell cycle and development, for l-OPCs were extracellular matrix and cell adhesion, and for MOLs were myelination and cytoskeleton. The e-OPCs were mostly confined to the premyelinating fetal group, and the l-OPCs were most highly represented in the pediatric age group, corresponding to the peak age of myelination. Cells expressing a signature characteristic of l-OPCs were identified in the adult brain in situ using RNAScope. These findings highlight the transcriptomic variability in OL-lineage cells before, during, and after peak myelination and contribute to identifying novel pathways required to achieve remyelination.

overall_design: Comparison of 3 sub-populations of oligodendrocyte-lineage cells derived from single-cell RNA sequencing of human tissue from 4 different age groups (fetal, pediatric, adolescent, adult)

first: Kelly

last: Perlman

first: Charles

middle: P

last: Couturier

first: Yaqubi

last: Moein

first: Tanti

last: Arnaud

first: Qiao

last: Cui

first: Florian

last: Pernin

first: Jo-Anne

last: Stratton

first: Jiannis

last: Ragoussis

first: Luke

last: Healy

first: Kevin

last: Petrecca

first: Roy

last: Dudley

first: Myriam

last: Srour

first: Jeffrey

middle: A

last: Hall

last: Kennedy

first: Naguib

last: Mechawar


title: PDA Indolylmaleimides Induce Anti-Tumor Effects in Prostate Carcinoma Cell Lines through Mitotic Death

city: Hannover

first: Jan Torben

last: Schille

address: Buenteweg 9

department: Small Animal Clinic

country: Germany

institute: University of Veterinary Medicine Hannover

summary:
Introduction: Castrate resistant prostate cancer in men shares several characteristics with late-stage canine prostate cancer. Consequently, dogs became a model organism for the study of the human disease. Due to current insufficient therapies, evaluation of novel therapeutic agents for late-stage prostate cancer is of considerable interest for both species.
Methods: In this interdisciplinary study we characterized the influence of two indolylmaleimides (PDA-66 and PDA-377) on human and canine prostate carcinoma cell lines. For this purpose, we analyzed morphology, proliferation and apoptosis, recorded live cell imaging movies and performed whole transcriptome sequencing.
Results: While PDA-377 showed only weak growth inhibition on the human prostate cancer cell lines, PDA-66 inhibited proliferation and induced apoptosis in all prostate carcinoma cell lines with concentrations in the low micromolar range. Morphological characterization and whole transcriptome sequencing revealed that PDA-66 induces mitotic death through its microtubule-depolymerizing ability.
Discussion: PDA-66 appears to be a worthwhile anti-mitotic agent for further evaluation. The similarities in cellular and molecular response observed in the cell lines of both origins form a solid basis for the use of canine PCa in vivo models to provide valuable transferable findings to the benefit of both species.

sample_organism:
Homo sapiens
Canis lupus familiaris

platform_organism:
Homo sapiens
Canis lupus familiaris

overall_design: Examination of the mediated effects by PDA-66 application on human and canine prostate carcinoma cell lines

first: Jan

first: Ingo

last: Nolte

first: Julia

last: Beck

first: Daria

last: Jilani

first: Catrin

last: Roolf

first: Anahit

last: Pews-Davtyan

first: Arndt

last: Rolfs

first: Larissa

last: Henze

first: Matthias

last: Beller

first: Bertram

last: Brenig

first: Christian

last: Junghanss

first: Ekkehard

last: Schütz

first: Hugo

last: Murua Escobar


title: The role of XIST in X-inactivation maintenance in B cells (RNA-seq)

first: Howard

last: Chang

overall_design: RNA-seq of dCas9-Krab+ GM B cells transduced with guide RNA and treated with DNMT and EZH2 epigenetic inhibitors (sgNT+DMSO, sgXIST+DMSO, sgNT+inhibitors, sgXIST+inhibitors), Trim28_KO GM B cells, Spen_deltaRRM GM B cells, sgNT and sgXIST cells that are overexpressed with anti-CRISPR protein ( sgNT+ACRIIA4, sgXIST+ACRIIA4), dCas9-Krab+ IMR90 cells transduced with guide RNA (IMR90_sgNT, IMR90_sgXIST) .

first: Bingfei

last: Yu

first: Yanyan

last: Qi

first: Rui

last: Li

first: Quanming

last: Shi

first: Ansuman

last: Satpathy


title: Next Generation Sequencing Facilitates Quantitative Analysis of E.coli DH5a Transcriptomes under triclosan exposure

city: Brisbane

first: Jianhua

last: Guo

address: Research Road, Level 4 Gehrmann Building

department: Australian Centre for Water and Environmental Biotechnology

country: Australia

institute: University of Queensland

summary: The goals of this study are to use Next-generation sequencing (NGS)to detect bacterial mRNA profiles of E.coli DH5a in response to 0, 20ug/L and 2 mg/L triclosan for 2 h, in triplicate, using Illumina HiSeq 2500.The NGS QC toolkit (version 2.3.3) was used to treat the raw sequence reads to trim the 3’-end residual adaptors and primers, and the ambiguous characters in the reads were removed. Then, the sequence reads consisting of at least 85% bases were progressively trimmed at the 3’-ends until a quality value ≥ 20 were kept. Downstream analyses were performed using the generated clean reads of no shorter than 75 bp. The clean reads of each sample were aligned to the E. coli reference genome (NC_000913) using SeqAlto (version 0.5). Cufflinks (version 2.2.1) was used to calculate the strand-specific coverage for each gene, and to analyze the differential expression in triplicate bacterial cell cultures. The statistical analyses and visualization were conducted using CummeRbund package in R (http://compbio.mit.edu/cummeRbund/). Gene expression was calculated as fragments per kilobase of a gene per million mapped reads (FPKM, a normalized value generated from the frequency of detection and the length of a given gene.

overall_design: Bacterial mRNA profiles of E.coli DH5a in response to  0, 20ug/L and 2 mg/L triclosan for 2 h, in triplicate, using Illumina HiSeq 2500

first: Ji

last: Lu


title: Peptidylarginine deiminase IV (PADI4) has a novel tumor cell-autonomous suppressor role in regulating breast cancer stem cells [RNA-Seq_2]

status: Public on Jan 11 2021

city: Bethesda

first: Maxwell

last: Lee

state: MD

address: Building 37, Room 5120

department: CCR

institute: National Cancer Institute, NIH

summary: Peptidylarginine deiminases (PADIs) catalyze post-translational modification of many target proteins and have been suggested to play a role in carcinogenesis. Since citrullination of histones by PADI4 was recently implicated in regulating embryonic stem and hematopoietic progenitor cells, here we investigated a possible role for PADI4 in regulating breast cancer stem cells. We showed by genetic and pharmacologic approaches that PADI4 activity limits the number of cancer stem cells (CSCs) in vitro and in vivo in multiple breast cancer models. A gene signature reflecting tumor cell-autonomous PADI4 inhibition is associated with poor outcome in human breast cancer datasets, consistent with a tumor suppressive role for PADI4. Mechanistically, PADI4 inhibition resulted in a global redistribution of histone H3 with accumulation around transcriptional start sites. Interestingly, epigenetic effects of PADI4 on the bulk tumor cell population did not explain the CSC phenotype. However, in sorted tumor cell populations, PADI4 down-regulated expression of the master transcription factors of stemness, NANOG and POU5F1, specifically in the cancer stem cell compartment, by reducing the transcriptionally activating H3R17me2a histone mark at those loci. This effect was not seen in the non-stem cells. Our findings reveal a novel role for PADI4 as a tumor suppressor in regulating breast cancer stem cells, and provide insights into context-specific effects of PADI4 in epigenetic modulation.

overall_design: MCF10Ca1h (M3) human breast cancer cells were treated with PADI4 inhibitor (GSK484) or control compound (GSK107) for 72 hours. Some samples were FACS sorted for SORE6-/+ populations.

first: Nellie

last: Moshkovich

first: Humberto

middle: J

last: Ochoa

first: Binwu

last: Tang

middle: H

last: Yang

first: Yu-an

first: Jing

last: Huang

first: Lalage

last: Wakefield


title: AtacWorks: A deep convolutional neural network toolkit for epigenomics

city: Cambridge

first: Jason

middle: Daniel

last: Buenrostro

address: 7 Divinity Ave

department: Stem Cell and Regenerative Biology

institute: Harvard University

summary: We introduce AtacWorks (https://github.com/clara-genomics/AtacWorks), a method to denoise and identify accessible chromatin regions from low-coverage or low-quality ATAC-seq data. AtacWorks uses a deep neural network to learn a mapping between noisy ATAC-seq data and corresponding higher-coverage or higher-quality data. We apply this approach to as few as 50 hematopoietic stem cells to identify regulatory elements that are differentially-accessible between rare lineage-primed cell subpopulations.

overall_design: dscATAC-seq of FACS-isolated hematopoietic stem cells

first: Zachary

middle: D

last: Chiang

first: Fabiana

middle: M

last: Duarte


title: Therapeutic modulation of nemo-like kinase in primary and acquired endocrine-resistant breast cancer

city: Pittsburgh

first: Xiaosong

last: Wang

state: Pennsylvania

address: 5150 Centre Ave

department: Pathology

institute: University of Pittsburgh

summary:
Purpose: To assess if NLK modulates ER target gene expression, we performed transcriptome sequencing following NLK inhibition or tamoxifen treatment in the BT483 or T47D-TamR cells under estrogen-deprived condition.
Methods: BT483 and T47D-TamR cells were treated with NLK-specific siRNAs, or VX-702, or Tamoxifen under estrogen-deprived condition to examine the expression profile changes of ER target genes. NovaSeq 6000 S2 Reagent Kit (Illumina) was used for paired-end reads (2×150 bp) sequencing reactions. The expression changes of ER target genes following NLK inhibition are compared to their changes following tamoxifen treatment.
Results: Our results show that the ER target gene expression changes following NLK silencing or VX-702 treatment correlate with their expression changes following tamoxifen treatment.

overall_design:
For RNA sequencing, total RNAs from the BT483 and T47D-TamR cells treated as below were extracted using the standard procedure of RNAzol (Molecular Research Center). Following estrogen deprived (ED) condition for 48hours (phenol red-free medium with 5% charcoal-dextran-stripped FBS), BT483 and T47D-TamR cells are seeded and treated with siCtrl +/- 0.5uM Tamoxifen, siNLK#1, and siNLK#2 for 72 hours. For siRNA treatment, the cells were seeded following 48h ED and reverse transfected with the respective siRNA (final concentration 10 nM). T47D-TamR cells are also treated with vehicle, 0.5uM Tamoxifen, or 0.5uM VX-702 following ED for 48hours.
The NovaSeq 6000 library for DNA sequencing was prepared using TruSeq Stranded mRNA Library Prep Kit (Illumina) following the protocol provided by the manufacturer. The final libraries were normalized by quantification with LightCycler 480 II (Roche Applied Science, Indianapolis, IN, USA) and quantification with Bioanalyzer (Agilent, Palo Alto, CA, USA). Final loading concentration was adjusted to 10 pM following the NovaSeq 6000 loading protocol and NovaSeq 6000 S2 Reagent Kit (Illumina) was used for paired-end reads (2×150 bp) sequencing reactions.

first: Suet

last: Loo

first: Chia

last: Liu

first: Sanghoon


title: Peptidylarginine deiminase IV (PADI4) has a novel tumor cell-autonomous suppressor role in regulating breast cancer stem cells [ChIP-Seq]

overall_design:
MCF10Ca1h cells were cultured in DMEM/F12 media supplemented with 5% horse serum.
Cells were treated with PADI4 inhibitor (GSK484) or control compound (GSK107) for 72 hours.

first: Howard


title: The histone demethylase KDM2B regulates human primordial germ cell-like cells specification

status: Public on Jan 10 2021

city: Munster

first: Veeramohan

last: Veerapandian

state: North Rhine-Westphalia

address: Röntgenstr. 20

department: Department Tissue Morphogenesis

institute: Max Planck Institute for Molecular Biomedicine

summary: Germline specification is a fundamental step for human reproduction and this biological phenomenon possesses technical challenges to study in vivo as it occurs immediately after blastocyst implantation. The establishment of in vitro human primordial germ cell-like cells (hPGCLCs) induction system allows sophisticated characterization of human primordial germ cells (hPGCs) development. However, the underlying molecular mechanisms of hPGCLC specification are not fully elucidated. Here, we observed particularly high expression of the histone demethylase KDM2B in male fetal germ cells (FGCs) but not in male somatic cells. Besides, KDM2B shared similar expression pattern with hPGC marker genes in hPGCLCs, suggesting an important role of KDM2B in germ cell development. Although deletion of KDM2B had no significant effects on human embryonic stem cell (hESC)’s pluripotency, loss of KDM2B dramatically impaired hPGCLCs differentiation whereas ectopically expressed KDM2B could efficiently rescue such defect, indicating this defect was due to KDM2B’s loss in hPGCLC specification. Mechanistically, as revealed by the transcriptional profiling, KDM2B suppressed the expression of somatic genes thus inhibited somatic differentiation during hPGCLC specification. These data collectively indicate that KDM2B is an indispensable epigenetic regulator for hPGCLC specification, shedding lights on how epigenetic regulations orchestrate transcriptional events in hPGC development for future investigation.

overall_design: We analyzed the time course expression profiles from human embryonic stem cells (hESCs) to human primordial germ cell like cells (hPGCLCs) in Wild type comparing to KDM2B-/-  (knockout) cell lines.

first: Xinyan

first: Fang

last: Luo

first: Xiao-yang

last: Zhao


title: Transcriptional profiling of Arabidopsis flower development of plants overexpressing ABAP1 gene

status: Public on Jan 09 2021

city: Rio de Janeiro

first: Adriana

middle: S

last: Hemerly

state: RJ

address: Bloco L, Subsolo, Salas 26,27 e 29 – CCS/UFRJ

department: Instituto de Bioquímica Médica Leopoldo de Meis

country: Brazil

institute: Universidade Federal do Rio de Janeiro

summary: Gene expression profile of flower buds at stage 13, open flowers at stage 14 and siliques at stages 15/16, according to Smyth et al., 1990) of ABAP1 overexpressing (ABAP1OE) plants compared to wild type flower buds, open flowers and siliques (Col-0) using a a whole-genome oligonucleotide array (Operon) (platform accession number accession number GPL1077). ABAP1 is a negative regulator of the cell cycle that binds to transcription factors and represses their target gene expression, including pre-replication complex genes. All experiments were performed in triplicate.

overall_design: Two-condition experiment, WT vs. ABAP1OE flower buds, open flowers and siliques. Three independent biological replicates.

first: Hana

last: Masuda

first: Marcio

last: Alves-Ferreira


title: In vivo evaluation of the 94 GHz-chronic exposure effect on Hairless rat skin gene expression

first: Denis

last: Habauzit

overall_design: Refer to individual Series


title: Whole-transcriptome microarray analysis of different biovars of Pseudomonas syringae pv. actinidiae grown in rich medium or apoplast-like conditions

city: Verona

first: Elodie

last: Vandelle

state: Verona

address: Strada Le Grazie, 15

department: Department of Biotechnology

country: Italy

institute: University of Verona

summary: Purpose: Pseudomonas syringae pv. actinidiae (Psa) is a phytopathogen that causes devastating bacterial canker in kiwifruit. Among five biovars defined by genetic, biochemical and virulence traits, Psa3 is the most aggressive and is responsible for the most recent reported outbreaks, but the molecular basis of its heightened virulence is unclear. A custom P. syringae multi-strain whole-genome microarray platform, encompassing biovars Psa1, Psa2 and Psa3 and the well-established model P. syringae pv. tomato, was used to analyse early bacterial responses to an apoplast-like minimal medium. Conlusion: this work highlighted that diverse early responses to the host apoplast, even among bacteria belonging to the same pathovar, can lead to different virulence strategies and may explain the differing outcomes of infections.

sample_organism:
Pseudomonas syringae pv. tomato
Pseudomonas syringae pv. actinidiae

overall_design: mRNA profiles of Psa1, Psa2 and Psa3 cells incubated in rich medium (optima growth conditions) were compared to mRNA profiles of Psa1, Psa2 and Psa3 cells grown in apoplast-mimicking medium. All mRNA profiles were generated from three biological replicates.

first: Teresa

last: Colombo

first: Alice

last: Regaiolo

first: Vaness

last: Maurizio

first: Tommaso

last: Libardi

first: Maria

last: Puttilli

first: Davide

last: Danzi

first: Annalisa

last: Polverari


title: Genome-wide analysis of the transcriptional profile of astrocytes during EAE in Gfap-TdTomato mice by scRNA-seq.

city: Boston

first: Michael

last: Wheeler

address: 60 Fenwood Rd.

department: Neurology

institute: Brigham and Women's Hospital

summary: We identify astrocyte heterogeneity during EAE in B6 mice.

overall_design: Naïve mice (n=3), EAE peak phase (n=2)

first: Liliana

last: Sanmarco

first: Francisco

last: Quintana

first: Zhaorong

last: Li


title: Analysis of astrocyte interferon gamma signaling during EAE.

summary: We identify pathways regulated by interferon gamma in astrocytes during EAE

overall_design: Mice transduced with sgScrmbl (n=4), sgTnfsf10 (n=3), sgIfngr1 (n=4), sgIfngr2 (n=4), sgStat1 (n=4)


title: Analysis of microglia responses following astrocyte knockdown of interferon gamma signaling during EAE.

summary: We identify pathways in microglia regulated by astrocyte interferon gamma signaling during EAE

overall_design: Mice transduced with astrocyte-specific sgScrmbl (n=4), sgTnfsf10 (n=3), sgIfngr1 (n=4), sgIfngr2 (n=4), sgStat1 (n=4)


title: Peptidylarginine deiminase IV (PADI4) has a novel tumor cell-autonomous suppressor role in regulating breast cancer stem cells [Array]

first: Maxwell

last: Lee

overall_design: MCF10Ca1h cells were transduced with a lentiviral control shRNA (shCON) or PADI4 knockdown (sh80) and selected with puromycin.

first: Nellie

last: Moshkovich

first: Humberto

last: Ochoa

first: Binwu

last: Tang

first: Howard

last: Yang

first: Yu-an

first: Jing

last: Huang

first: Lalage

last: Wakefield


title: miRNA expression in differentiating primary human bronchial epithelial cells cultured at air-liquid-interface

city: San Francisco

first: Srilaxmi

last: Nerella

state: CA

address: HSE1355A, 513 Parnassus ave

department: Department of Medicine

institute: UCSF

summary: We performed microarray analysis of miRNA expression in differentiating primary human bronchial epithelial cells. The goal was to identify miRNAs that are dynamically expressed under airway epithelial development. Cells were cultured at air-liquid-interface and were harvested day 4, 6, 8, 11, 13, 15, 18, 20 and 22 for analysis.

overall_design:
Cells were cultured at air-liquid-interface and were harvested day 4, 6, 8, 11, 13, 15, 18, 20 and 22 for analysis.
Total RNA was labeled with Cy3-CTP and hybridized overnight at 65 deg C in Agilent hybridization buffer. Slides were scanned on an Agilent scanner immediately after washing and scanned images were analyzed with Feature Extraction Software 10.6 (Agilent) using default parameters. Log2 Normalized signal intensity (derived from replicate probes printed on the same array).

first: David

last: Erle

first: Prescott

middle: G

last: Woodruff


title: Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and Ccn2a-/- cardiac ventricular Transcriptomes

city: bad nauheim

first: Stefan

last: Günther

address: ludwigtr. 43

institute: MPI for heart and lung research

summary: Purpose: The goal of this study is to compare transcriptome level between 4 dpci wild-type and ccn2a-/- zebrafish cardiac ventricle. Methods: 4-days-post cryoinjured cardiac ventricular mRNA profiles of wild-type (WT) and cellular communication network factor 2a mutant (ccn2a−/−) zebrafish were generated by deep sequencing, in duplicate. Results: 7 genes showed differential expression between the WT and ccn2a−/− heart, with a fold change ≥1.5 and p value <0.05. Altered expression of all genes was confirmed with qRT–PCR.   Conclusions: Our study represents the detailed analysis of 4 dpci wild-type and ccn2a-/- cardiac ventricular transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that 7 genes were dysregulated in 4 dpci ccn2a-/- cardiac ventricle. We concluded, this analysis helped us to identify downstream signalling cascade of Ccn2a in injured zebrafish heart."

overall_design: Cardiac ventricular mRNA profiles of 4dpci wild type (WT) and ccn2a-/- zebrafish were generated by deep sequencing, in duplicate.

first: Kontarakis

last: Zacharias

first: Günther

last: Stefan

first: Patra

last: Chinmoy


title: Genome-wide analysis of the transcriptional profile of TRAIL+LAMP1+ astrocytes and LAMP1+ astrocytes during EAE in Aldh1l1CreERT2-TdTomato mice by scRNA-seq

first: Michael

last: Wheeler

overall_design: Naïve mice (n=3), EAE peak phase (n=3). Sorting of Aldh1l1CreERT2-TdTomato+ astrocytes with LAMP and/or TRAIL surface staining

first: Francisco


title: CELF6 5HT TRAP & Total RNA

city: St Louis

first: Joseph

last: Dougherty

state: Missouri

address: 4566 Scott Ave, box 8232

department: Genetics and Psychiatry

institute: Washington University

summary: Celf6 is an RNA binding protein expressed in a subset of neurons in the brain.  We conducted an analysis of splicing in RNAseq data derived from Celf6 knockout mice, compared to wildtype littermates. We profiled both total tissue RNAseq from a hindbrain dissection, as well as Translated Ribosome Affinity Purified (TRAP)seq specifically from serotonergic neurons – a population of cells we previously characterized as expressing Celf6. We used MISO for the splicing analysis (PMID: 21057496). MISO uses read evidence found in a sample in a Bayesian model in order to assign a percent spliced in (PSI) value, defining the expected frequency of an alternate event.    In the total RNA sample, the number of measurable PSI events was 1817. The most annotated events found were the Skipped Exons (35.5%) , Retained Introns (25 %), and Tandem 3’UTR (alternative poly-adenylation, 17.2%) categories. In general, the Skipped Exons category has the largest number of annotated events in MISO (Figure 1C). Out of 1817 assignable events, 95 showed nominal ANOVA p<0.05 between WT and Celf6 KO with 0 meeting Benjamini-Hochberg FDR <0.05. Alternative event categories had roughly the same distribution in nominally significant differentially abundant alternative events as to the data as a whole. 33/95 nominally significant events were in genes that were putative CELF6 targets, however only two (C1d, Rit2) were in genes matching both CLIP stringency criteria (significant enrichment in both CLIP input & antibody non-specific binding controls). As PSI describes the probability of being either in alternative form A (PSI = 0) or B (PSI = 1), large changes in PSI between one sample to another might indicate a splicing switch between predominant forms. Among the nominally significant events, the majority (88/95, 92.6%) were between -0.2 to 0.2  indicating only slight changes in splice form abundance. There were a few events with larger changes in PSI, with differences between KO and WT greater that 0.2, 5 in alternative polyadenylation, 1 in a Mutually Exclusive Exon event, and 1 in Alternative 5’ Splice Site usage. The largest change of these was in Acox1 (not a putative CELF6 target), peroxisomal straight-chain acyl-CoA oxidase, with a change of -0.62 in the KO (p = 0.037) which may indicate evidence of a switch in usage of a 897 nucleotide 3’UTR in the WT (WT median PSI = 0.89) to a 1660 nucleotide 3’UTR in the KO (KO median PSI = 0.26). However, none of these survive FRD correction.    In serotonergic cell mRNA accessed by TRAP, the total measurable events was 2,201 with a distribution of effect sizes similar to that found in total RNA.  112 of these events showed nominal p values < 0.05, with 2 events meeting Benjamini-Hochberg FDR < 0.05, which showed a similar distribution to the data as a whole. 40 nominally significant events were found in putative CELF6 CLIP targets, 5 genes meeting the most stringent criteria (Snap25, Tpm1, Rnf14, Shroom2, App). As with total RNA, most events showed small differences in PSI between WT and KO (-0.2 to 0.2, median ~.01), with only 4 events outside this range. The largest such difference was found in Nmnat2, nicotinamide nucleotide adenylyltransferase 2 (not a putative CELF6 target), again in an alternative 3’ UTR. The WT median PSI of 0.55 indicates two isoforms are similarly abundant: a 3.6 kb 3’UTR and a 400 bp UTR. In the KO, this is shifted towards the shorter UTR (KO median PSI 0.98).

overall_design: 5 WT and 5 Celf6 KO mouse brains were dissected at post-natal day 9 for changes to splicing in total RNA & serotonergic cell TRAP samples. We used MISO to calculate PSI values for alternate annotated events for each sample, and filtered events to those PSI values with credible intervals less than or equal to 0.20 in width in each sample. As MISO has no intrinsic ability to account for replicate design, we performed hypothesis testing between WT and Celf6 KO replicate sample PSI values using ANOVA on the logit transform (log2 (PSI/(1-PSI)), and required a minimum of 3 samples to have an assigned PSI value based on the filtering above.

last: Rieger

first: Yating

last: Liu


title: Evaluating Capture Sequence Performance for Single-cell CRISPR-activation Experiments

status: Public on Jan 08 2021

city: Singapore

first: John

middle: F.

last: Ouyang

address: 8 College Road, Level 8 CVMD

country: Singapore

institute: Duke-NUS Medical School

summary: The combination of single-cell RNA sequencing with CRISPR inhibition/activation provides a high-throughput approach to simultaneously study the effects of hundreds if not thousands of gene perturbations in a single experiment. One recent development in CRISPR-based single-cell techniques introduces a feature barcoding technology which allows for the simultaneous capture of mRNA and gRNA from the same cell. This is achieved by introducing a capture sequence, whose complement can be incorporated into each gRNA, and which can be used to amplify these features prior to sequencing. However, with the technology in its infancy, there is little information available on how such experimental parameters can be optimised. To overcome this, we varied the capture sequence, capture sequence position and gRNA backbone to identify an optimal gRNA scaffold for CRISPR-activation gene perturbation studies. We provide a report on our screening approach along with our observations and recommendations for future use.

overall_design: single cell RNA sequencing of hESC overexpressing either ASCL1 or SOX17 via CRISPR activation, cells are collected 3 days, 5 days and 7 days after introduction of gRNA and pooled into a single single-cell library, which is the sequenced for gene expression and CRISPR gRNA capture

first: Xin

last: Choo

middle: F

first: Owen

last: Rackham


title: Next Generation Sequencing unveils the effect of caloric restriction on the splenic response to virulent H37rv Mycobacterium tubercolosis in DBA/2 mice

city: Naples

first: Giuseppe

last: Matarese

address: Via Pansini 5

department: Dipartimento di Medicina Molecolare e Biotecnologie Mediche

institute: Università di Napoli Federico II

summary:
Purpose: The goal of this study was the transcriptome high-throughput data analysis of spleen cells from mice infected with a virulent strain of mycobacterium (H37rv) and either fed ad libitum (AL) or caloric restricted (CR)

Methods: Spleen cell total RNA profile of 17 weeks old mice (1. MTB infected, Ad libitum fed; 2. MTB infected, Caloric restricted; 3. Not infected, Ad libitum fed; 4. Not infected, Caloric restricted; a pool of 5 animals per group) were generated by deep sequencing, in triplicate, using TruSeq Stranded Total RNA Illumina technology

Results: Principal component analysis and unsupervised clustering showed that the RNA-seq profiles of spleen cells from these four groups of mice were clearly distinct and infection was the major force affecting gene expression, with 67% of variance coming from the infection status and another 23% variance being dependent on caloric intake .   When directly comparing AL versus CR MTB infected spleens, 628 genes were found differentially expressed. Supervised hierarchical clustering based on the expression of these 628 genes was able to unveil the unique transcriptional profile of CR MTB infected mice compared to both AL MTB infected and Not infected mice.

Coclusions: Caloric restriction is able to affect the expression of a significant number of genes in spleen cells isolated from mice infected with MTB. The study of biological process enrichements highlights relevant immune-related regulation in these animals, than may be at the base of the higher capacity of caloric restricted, compared to ad libitum fed, animals to resist MTB infection.

overall_design: Spleen cell mRNA profiles of 17 weeks old DBA/2 mice (a pool of 5 animals per group) were generated by deep sequencing, in triplicate, using Illumina platform.

first: Carla

last: Palma


title: Genome-wide analysis of the transcriptional profile of WT and Lamp1 KD astrocytes during EAE in B6 mice

summary: We identify pathways regulated by Lamp1 in astrocytes during EAE.

overall_design: sgScrmbl-targeted (n=3) and sgLamp1-targeted (n=3) mice.

first: Zhaorong

last: Li

last: Quintana


title: Identification of NEDD9 regulated genes in VCaP prostate cancer cells

first: Dong

last: Han

address: 100 Morrissey Blvd

department: Biology-CPCT

institute: Umass Boston

summary: NEDD9 gene (Neural precursor Expressed, Developmentally Down-regulated 9) encodes a member of the Cas family of signaling transduction molecules, which is a non-catalytic focal adhesion protein phosphorylated by FAK and Src and functions as a signaling hub to regulate downstream signaling, such as the AKT pathway. In cancers, NEDD9 regulates cell migration, epithelial-mesenchymal transition (EMT), invasion, and metastasis. Gene amplification of NEDD9 is commonly found in the advanced metastatic castration-resistant PCa (~3%) and neuroendocrine PCa (~15%), suggesting NEDD9 may play an important role in PCa progression. In order to characterize the global functions of NEDD9 in PCa cells, we performed RNA-seq analyses in VCaP cells (containing NEDD9 gene amplification) stably infected with shRNA against NEDD9 or non-targeting control (NTC).

overall_design: RNA-seq analyses were performed to examine gene expressions altered by silencing NEDD9 in VCaP cells. The stable cells were generated by stably infecting VCaP cells with shRNA against NEDD9 (shNEDD9) versus non-targeting control (shNTC). These stable cells were grown in the medium containing 10% FBS. The experiments were done in biological triplicates.

first: Maryam

last: Labaf

first: Kourosh

last: Zarringhalam

first: Changmeng

last: Cai


title: Concerted roles of PTEN and ATM in controlling hematopoietic stem cell fitness and dormancy

city: Toronto

first: Tak

last: Mak

state: ON

address: 610 University Ave

department: The Campbell Family Institute for Breast Cancer Research

institute: Princess Margaret Cancer Centre, University Health Network

summary: In order to sustain proficient life-long hematopoiesis, hematopoietic stem cells (HSCs) must possess robust mechanisms to preserve their quiescence and genome integrity. DNA-damaging stress can perturb HSC homeostasis by affecting their survival, self-renewal and differentiation. Ablation of the kinase ATM, a master regulator of the DNA damage response, impairs HSC fitness. Paradoxically, we show here that loss of a single allele of Atm enhances HSC functionality in mice. To explain this observation, we explored a possible link between ATM and the tumor suppressor PTEN, which also regulates HSC function. We generated and analyzed a knock-in mouse line (PtenS398A/S398A), in which PTEN cannot be phosphorylated by ATM. Similar to Atm+/-, PtenS398A/S398A HSCs have enhanced hematopoietic reconstitution ability, accompanied by resistance to apoptosis induced by genotoxic stress. Single-cell transcriptomic analyses and functional assays revealed that dormant PtenS398A/S398A HSCs aberrantly tolerate elevated mitochondrial activity and the accumulation of reactive oxygen species, which are normally associated with HSC priming for self-renewal or differentiation. Our results unveil a molecular connection between ATM and PTEN, which couples the response to genotoxic stress and dormancy in HSC.

overall_design: Long-term hematopoietic stem cells (Lin-Sca1+cKit+CD48-CD150+) were sorted from pools of 3 male and 3 female mice per genotype (Pten+/+ and PtenS398A/S398A). Cells were sorted directly onto the cell loading inlet of the C1 chip. The raw data had a total of 280 cells. After quality control analyses, a total of 206 cells (116 Pten+/+ cells and 90 PtenS398A/S398A) were retained for further downstream analyses.

first: Jerome

last: Fortin

first: Parameswaran

last: Ramachandran

first: Christian

last: Bassi

first: Wanda


title: SUMOylation regulates the protein network and chromatin accessibility at glucocorticoid receptor-binding sites [array]

city: Kuopio

first: Ville

last: Paakinaho

address: Yliopistonranta 8

department: School of Medicine

country: Finland

institute: University of Eastern Finland

summary: Glucocorticoid receptor (GR) is an essential transcription factor (TF), controlling metabolism, development and immune responses. SUMOylation regulates chromatin occupancy and target gene expression of GR in a locus-selective manner, but the mechanism of regulation has remained elusive. Here, we identify the protein network around chromatin-bound GR by using selective isolation of chromatin-associated proteins and show that the network is affected by receptor SUMOylation, with several nuclear receptor coregulators and chromatin modifiers preferring interaction with SUMOylation-deficient GR and proteins implicated in transcriptional repression preferring interaction with SUMOylation-competent GR. This difference is reflected in our chromatin binding, chromatin accessibility and gene expression data, showing that the SUMOylation-deficient GR is more potent in binding and opening chromatin at glucocorticoid-regulated enhancers and inducing expression of target loci. Blockage of SUMOylation by a SUMO-activating enzyme inhibitor (ML-792) phenocopied to a large extent the consequences of GR SUMOylation deficiency on chromatin binding and target gene expression. Our results thus show that SUMOylation modulates the specificity of GR by regulating its chromatin protein network and accessibility at GR-bound enhancers. We speculate that many other SUMOylated TFs utilize a similar regulatory mechanism.

overall_design: Total RNA isolated from isogenic HEK293 cells stably expressing wild-type GR (HEK293flpGR) treated with non-targeting control (siNON) or UBC9 (siUBC9) siRNA for 72 h and exposed to either vehicle (EtOH) or 100 nM of dexamethasone (dex) for 6 h. All conditions are performed in triplicate.

first: Marjo

last: Malinen

first: Joanna

middle: K

last: Lempiäinen

first: Einari

last: Niskanen

first: Jorma

last: Palvimo


title: Asc-1 regulates white versus beige adipocyte fate in a subcutaneous stromal cell population

status: Public on Jan 07 2021

city: Neuherberg

first: Matthias

last: Heinig

address: Ingolstädter Landstrasse

institute: Helmholtz-Zentrum München

summary: Here we provide scRNAseq data from the stromal vascular fraction of adult (8 weeks) and adolescent (2 weeks)  murine inguinal subcutaneous white adipose tissue. This allows studying cellular composition and cellular heterogeneity within subcutaneous fat.

overall_design: We utilize the stromal vascular fraction of subcutaneous white adipose tissue of  adult (8 weeks) and adolescent (2 weeks) old male C57Bl/6J mice

first: Lisa

last: Suwandhi

first: Irem

last: Altun

first: Ruth

last: Karlina

first: Viktorian

last: Miok

first: Tobias

last: Wiedemann

first: David

last: Fischer

first: Thomas

last: Walzthoeni

first: Christina

last: Lindner

first: Anika

last: Boettcher

first: Silke

last: Heinzmann

first: Andreas

last: Israel

first: Ahmed

last: Khalil

first: Alexander

last: Braun

first: Ines

last: Pramme-Steinwachs

first: Ingo

last: Burtscher

first: Philippe

last: Schmitt-Kopplin

first: Martin

last: Elsner

first: Heicko

last: Lickert

first: Fabian

last: Theis

first: Siegfried

last: Ussar


title: Ahmed_Shireman Brain 2021 ChIPSeq

city: Madison

first: Jack

last: Shireman

state: WI

address: 1500 Highland Ave

department: Neurological Surgery

institute: UW-Madison

summary: Glioblastoma is a primary brain cancer with a near 100% recurrence rate. Upon recurrence, the tumor is resistant to all conventional therapies, and because of this, 5-year survival is dismal. One of the major drivers of this high recurrence rate is the ability of GBM cells to adapt to complex changes within the tumor microenvironment. To elucidate the molecular mechanisms of this adaptation, specifically during chemotherapy, we employed ChIP-Sequencing and gene expression analysis. We identified a molecular circuit in which the expression of ciliary protein ALR13B is epigenetically regulated to promote adaptation to chemotherapy. Immuno-precipitation combined with Liquid Chromatography-Mass Spectrometry binding partner analysis revealed that that ARL13B interacts with the purine biosynthetic enzyme IMPDH2. Further, radioisotope tracing revealed that this interaction function as a negative regulator for purine salvaging. Inhibition of ARL13B-IMPDH2 interaction enhances temozolomide (TMZ)-induced DNA damage by forcing GBM cells to rely on the purine salvage pathway. Targeting the ARLI3B-IMPDH2 circuit can be achieved by using an FDA-approved drug, Mycophenolate Mofetil, that can block the IMPDH2 activity and enhance the therapeutic efficacy of TMZ. Our results suggest and support clinical evaluation of MMF in combination with TMZ treatment in glioma patients.

overall_design: Comparison of H3K27me3 marks in glioblastoma cells (proneural GBM 43) after 4 days of either DMSO or TMZ treatment

first: Atique

last: Ahmed

first: Cheol

last: Park


title: Heterogenous chemosensitivity of a panel of organoid lines derived from small cell neuroendocrine carcinoma of the uterine cervix

city: Kyoto

first: Masahiro

last: Inoue

state: Kyoto

address: Med-Pharm Collaboration Bldg 503, Shimoadachi-cho 46, Sakyou-ku, Kyoto, 606-8304, Japan

country: Japan

institute: Kyoto Univ Graduate School of Medicine

summary: Small cell neuroendocrine carcinoma (SCNEC) of the uterine cervix is a rare disease with a poor prognosis. The lack of established disease models has hampered therapy development. We established a panel of organoid from patients of SCNEC of the uterine cervix. Sensitivity assays against clinically used drugs revealed remarkable variations between the lines, and we successfully identified specific gene sets which likely contribute to the sensitivity to the tested drugs. Of note, we found one line which was exceptionally sensitive to irinotecan, and investigated the mode of action in the case. Chemotherapeutic drug sensitivity assays using SCNEC CTOS lines obtained from primary patient samples with a high success rate may provide useful information for treating SCNEC.

overall_design: One sample for each condition.


title: Hyperglycemic memory of innate immune cells promotes in vitro proinflammatory responses of human monocytes and murine macrophages

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

country: Netherlands

institute: Wageningen University

summary: It has been well established that the presence of diabetes is accompanied by a chronic inflammatory state promoting various diabetes-associated complications. One potential driver of this enhanced inflammatory state in patients with diabetes is hyperglycemia. Even after blood glucose control is achieved, diabetes-associated complications persist, suggesting the presence of a “hyperglycemic memory.” Innate immune cells, critically involved in various complications associated with diabetes, can build nonspecific, immunological memory (trained immunity) via epigenetic regulation. We examine the potential involvement of hyperglycemia-induced trained immunity in promoting inflammation. Our results show that hyperglycemia induces a trained phenotype in vivo in mice and in vitro in human monocytes, representative by an increased TNF-a secretion after ex vivo stimulation with LPS. These effects were largely mediated by epigenetic changes controlled by the mixed lineage leukemia (MLL) family because treatment with the MLL inhibitor menin-MLL during the process of trained immunity acquisition repressed the proinflammatory phenotype. Collectively, our results identify a novel link between hyperglycemia and inflammation in innate immune cells that might explain the increased proinflammatory state during diabetes potentially contributing to the development of various diabetes-associated complications.

overall_design: Microarray analysis was performed on CD14+ monocytes isolated from peripheral blood mononuclear cells obtained from patients with type 1 diabetes and healthy nondiabetic control subjects

first: Kathrin

last: Thiem

first: Rinke

last: Stienstra


title: Zc3h10 controls pre-mRNA profile in preadipocytes

city: Milan

first: Nico

last: Mitro

state: MI

address: Via Balzaretti 9

department: Pharmacological and biomolecular sciences

institute: Università degli studi di Milano

summary: NGS was performed to understand the impact of Zc3h10 on pre-mRNA expression profile in C3H10T1/2 preadipocytes.

overall_design: Cells were transduced after 24 hours they reached confluency with adenoviral vectors carrying Scramble and ShZc3h10 sequences. After 36h cells were labelled with 4-thiouridine 300μM for 10 minutes and then total RNA was isolated. Labelled pre-mRNA was subsequently isolated by biotin-streptavidin method.

first: M

last: Audano

first: N

first: F

last: Gualdrini


title: Total RNA sequencing for human induced pluripotent derived cortical neurons

first: Annie

last: Kathuria

address: 185 cambridge street

institute: Mass Gen Hospital

summary: Total RNA sequencing for human induced pluripotent derived neurons organoids from healthy controls and schizophrenia (SCZ) patients

overall_design: Comparing the disease state (SCZ) versus healthy control

first: Rakesh

last: Karmacharya


title: Gene expression profiles in skeletal muscle satellite cells lacking Dnmt1

city: Toon

first: Yuuki

last: Imai

state: Ehime

address: Shitsukawa

department: Proteo-Science Center

institute: Ehime University

summary: To identify the target genes of Dnmt1 in skeletal muscle satellite cells, we generated satellite cell specific Dnmt1 KO using Pax7-CreERT2 and Dnmt1 flox mice. After Tamoxifen (cKO) or vehicle (Control) treatment for 5 consecutive days, mice were euthanized and muscle satellite cells were collected by FACS sorting and total RNA were extracted.

overall_design: mRNA profiles of muscle satellite cells obtained from control and satellite cell specific Dnmt1 KO mice were generated by sequencing, in triplicates, using Illumina Miseq.

first: Hiroyuki

last: Iio


title: Relevance of Unfolded Protein Response to Spaceflight-Induced Transcriptional Reprogramming in Arabidopsis

city: East Lansing

first: Dae Kwan

last: Ko

address: 612 Wilson Rd.  Plant Biology RM 122

department: Plant Research Laboratories

institute: Michigan State University

summary: Plants are primary producers of food and oxygen on Earth and will likewise be indispensable to the establishment of large-scale sustainable ecosystems and human survival in space. To contribute to the understanding of how plants respond to spaceflight stresses, we examined the relevance of the unfolded protein response (UPR), a conserved signaling cascade that responds to a number of unfavorable environmental stresses, in the model plant species Arabidopsis thaliana.  To do so, we compared the transcriptional responses of wild type and UPR-defective seedlings to spaceflight during the SpaceX-CRS12 mission to the International Space Station. We established that orbital culture substantially altered the expression of hundreds of stress related genes compared to ground control conditions. Although many of these genes were differentially regulated in the UPR mutants in the ground control conditions compared to wild type, their expression was largely equalized in all genotypes by flight. Our results have yielded new information on how plants respond to growth in orbit and support the hypothesis that spaceflight induces the activation of signaling pathways that compensate for the loss of UPR regulators in the control of downstream transcriptional regulatory networks.

overall_design:
RNA-seq profiles of WT (Col-0) and various mutants of sensor genes for unfolded protein response in Arabidopsis seedlings in spaceflight compared to ground.
Please note that the *_5 samples are used for initial quality controls, but not used for further data processing (as indicated in the corresponding sample description field). Therefore there are only 4 biological replicates for eachh genotype/treatment and total 40 processed data columns in the 'fpkm.txt'.

first: DaeKwan

first: Evan

last: Angelos

first: Federica

last: Brandizzi


title: Colorectal cancer cells possess an equipotent capacity to enter a reversible diapause-like state to survive chemotherapy

first: Catherine

last: O'Brien

address: 101 College St., PMCRT, Suite 13-704

institute: University Health Network

summary: Cancer cells enter a reversible drug tolerant persister (DTP) state to evade death from chemotherapies and targeted agents. It is increasingly appreciated that DTPs are an important driver of therapy failure and tumor relapse. We combined cellular barcoding and mathematical modeling in patient-derived colorectal cancer models to identify and characterize DTPs in response to chemotherapy. Barcode analysis revealed no loss in clonal complexity of tumors that entered the DTP state and recurred following treatment cessation. Our data fits a mathematical model where all cancer cells, and not a small subpopulation, possess an equipotent capacity to become DTPs. Mechanistically, we determined that DTPs display remarkable transcriptional and functional similarities to diapause, a reversible state of suspended embryonic development triggered by unfavorable environmental conditions. Our study provides insight into how cancer cells use a developmentally conserved mechanism to drive the DTP state pointing to novel therapeutic opportunities to target DTPs.

overall_design: mRNA profiles of POP66 PDXs treated with Vehicle (DMSO, Saline) or CPT-11 (DTP-state, regrowth, and resistant)

first: Sumaiyah

last: Rehman

first: Jennifer

last: Haynes

first: Evelyne

last: Collignon

first: Kevin

middle: R

last: Brown

first: Yadong

last: Wang

first: Jeff

last: Bruce

first: Arvind

last: Mer

first: Edwyn

last: Lo

first: Cherry

last: Leung

last: Lima-Fernandes

first: Nicholas

last: Pedley

first: Sophie

last: McGibbon-Gardner

first: Aaron

last: Pollet

first: Trevor

last: Pugh

first: Benjamin

last: Haib-Kains

first: Miguel

last: Ramalho-Santos

first: Sidhartha

last: Goyal

first: Jason

last: Moffat

first: Allison

last: Nixon

first: Fraser

last: Soares

first: Housheng

last: He

first: Quaid

last: Morris


title: RNAseq of LepA knock-out in M. smegmatis

city: College Station

first: Thomas

last: Ioerger

state: TX

address: 3112 TAMUS

department: Department of Computer Science

institute: Texas A&M University

summary: Expression profiling in LepA knock-out in M. smegmatis

overall_design: 3 replicates each of wild-type (mc2 155), LepA knock-out, and complemented strain (LepA integrated in L5 site); BioProject PRJNA518044; SRA Study SRP183056

first: Skye

last: Fishbein


title: Next Generation Sequencing Facilitates Analysis of Fangchinoline Treatment and Control SamplesTranscriptomes

city: Shanghai

first: keting

last: bao

address: 130 Mei Long Road

institute: East China University of Science and Technology

summary: Conjunctival melanoma (CM) is a rare and potentially deadly ocular tumor with poor prognosis. Thus, effective drugs to kill CM are urgently needed. Herein, we reported the discovery of a novel potential therapeutic target for CM. Through phenotypic screening of our in-house library, fangchinoline was discovered to potently inhibit the growth of CM cell lines including CM-AS16, CRMM1, CRMM2 and CM2005.1. Further mechanistic experiments indicated that fangchinoline could suppress homologous recombination (HR)-directed DNA repair through binding to far upstream element binding protein 2 (FUBP2) and downregulating expression of the HR factors BRCA1 and RAD51. In vitro and in vivo antitumor experiments revealed that fangchinoline increased the efficacy of cisplatin by blocking HR factors and diminished the drug dose and toxicity. In summary, our work provides a promising therapeutic strategy for the treatment of CM that is worthy of extensive preclinical investigation.

overall_design: Examination of fangchinoline treatment and control samples in CM-AS16 cells by RNA-seq, in triplicate.

first: Keting

last: Bao

first: Yongyun

last: Li

first: Jinlian

last: Wei

first: Jian

first: Fei

last: Mao


title: Cell-type specific analysis of physiological action of estrogen in mouse oviducts

status: Public on Jan 06 2021

city: Columbia

first: Wipawee

last: Winuthayanon

address: 1030 Hitt Street

department: OB/GYN & Women's Health

institute: University of Missouri

summary: The goal of this study is to determine the oviduct responds to 17beta-estradiol (E2) in an in vivo environment using mouse models

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Adult female mice (C57B6/J, 8-12 weeks old) were purchased from the Jackson (JAX) laboratory. Animals were ovariectomized and left for 14 days to recover and eliminate endogenous steroid hormones . Sesame oil (100 uL) was used as a negative (Vehicle or Veh) control group. 17beta-estradiol (E2) was diluted in 100 uL of 10% ethanol and sesame oil. Ovariectomized females were subcutaneously injected with Veh or E2. Twenty-four hrs after injection, the oviduct was collected for scRNA-seq analysis. Vaginal washes were collected from adult female mice (between 8-12 weeks old) to determine the stage of estrous cycle. At 11:00 h, oviducts from females in estrus were collected for subsequent scRNA-seq analysis.

first: Emily

last: McGlade

first: Gerardo

last: Herrera

first: Kalli

last: Stephens

first: Sierra

last: Olsen

first: Sarayut

first: Diana

last: Monsivais

first: Joie

last: Guner

first: Francesco

last: Demayo

first: John

last: Lydon


title: Transcriptome analysis of KM-H2 cells with different Rhodamin 123 eflux capacity

city: Halle (Saale)

first: Martin

middle: Sebastian

last: Staege

state: Sachsen-Anhalt

address: Ernst-Grube-Str. 40

department: Universitätsklinik u. Poliklinik für Kinder- u. Jugendmedizin

summary:
Rhodamin 123 is a dye which can be used to detect the activity of ABC transporters. We observed that after staining of KM-H2 Hodgkin lymphoma cells with Rhodamin 123, part of the cells rapidly eliminated the dye, while another part of the cells retained the dye for a longer time.
We compared the transcriptome of KM-H2 Hodgkin lymphoma cells with high Rhodamin 123 efflux capacity and KM-H2 cells with low Rhodamin 123 efflux capacity.

overall_design: KM-H2 cells were stained with Rhodamine 123, washed, and incubated for 4 hrs. Thereafter, cells with high Rhodamine 123 efflux capacity and low efflux capacity were separated by flow cytometry. Samples were analyzed using the Clariom D Human platform. Array data was processed by using Transcriptome Analysis Console.

first: Teresa

last: Kreinest

first: Ines

last: Volkmer


title: SUMOylation regulates the protein network and chromatin accessibility at glucocorticoid receptor-binding sites

first: Ville

last: Paakinaho

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

overall_design: Refer to individual Series


title: Single-Cell RNA Sequencing of Batch Chlamydomonas Cultures Reveals their Heterogeneity and Place Along the Diurnal Cycle

city: Los Angeles

first: Feiyang

last: Ma

address: 610, Charles Young Dr East, TLSB 3000C

department: Molecular Biology Institute

institute: UCLA

summary: We did single cell RNA sequencing on cultured chlamydomonas cells. We used two strains, one with cell wall, one without cell wall. The cells were cultured under three different conditions, with iron, without iron and without nitrogen.

overall_design: We used two strains: one with cell wall, one without cell wall. We compared cells cultured under three different conditions: with iron, without iron and without nitrogen.

first: Patrice

last: Salomé

first: Sabeeha

last: Merchant

first: Matteo

last: Pellegrini


title: IFN-gamma+ NK cells control TRAIL+ anti-inflammatory astrocytes

city: Boston

first: Michael

last: Wheeler

state: MA

address: 60 Fenwood Rd.

institute: Brigham and Women's Hospital


title: High-throughput screen in post-implantation haploid epiblast stem cells reveals Hs3st3b1 as a modulator for reprogramming

city: Tianjin

first: Ling

last: Shuai

address: 38 Tong Yan Road, Jinnan district

department: State Key Laboratory of Medicinal Chemical Biology and College of Pharmacy

institute: Nankai University

summary: Epiblast stem cells (EpiSCs) derived from post-implantation epiblast are pluripotent stem cells, epigenetically distinct from embryonic stem cells (ESCs), which are widely used in reprogramming studies. Recent achieved haploid cell lines in mammalian open a new era for high-throughput genetic screening, due to their homozygous phenotypes. Here, we report the generation of mouse haploid EpiSCs (haEpiSCs) from post-implantation embryos at embryonic day 6.5 (E6.5). These cells maintain one set of chromosomes, express pluripotent and EpiSC specific genes, and have potentials to differentiate into three germ layers. We also develop a massive mutagenesis protocol with haEpiSCs, and subsequently perform reprogramming genetic screening using this mutation library. Multiple mutants related to various pathway modules are implicated. The validation experiment proves that Knock-out (KO) of Hst3st3b1 (One of the candidates) can promote reprogramming of EpiSCs to ground state efficiently. Our results open the possibility of utilizing haEpiSCs to uncover fundamental biological processes including cell fate alternations.

overall_design: Comparation of the transcriptomes between haEpiSCs, WT-diploid EpiSCs, p53-KO diploid EpiSCs,  p53-KO haEpiLCs, WT-EpiLCs, WT-ESCs, haESCs and MEFs by RNA-seq analysis, in triplicate

first: Qian

last: Gao

first: Wenhao

last: Zhang

first: Yiding

last: Zhao


title: Identification of a cancer driver lncRNA in DNA damage response

city: Gothenburg

first: Subazini

last: TK

address: Medicineragatan 9A

country: Sweden

institute: Gothenburg university

summary: We report that the lncRNA SCAT7 (ELF3-AS1) is involved in the activation of the DNA damage response in cisplatin resistant LUAD cells, akin cisplatin in sensitive cells. Therefore SCAT7 is a potential therapeutic target for drug resistant cancers

overall_design: Effects of cisplatin treatment were assessed for cisplatin resistant (A549/cDDP) cells and their parental cell line by RNA-Seq. Evaluation of the role of lncRNA SCAT7 in a therapeutic approach alternative to cisplatin in A549/cDDP cells was assessed by RNA-Seq following lncRNA knockdown, in presence or absence of drug treatment.

first: Chandrasekhar

last: Kanduri

first: Luisa

last: Statello

first: Mohamad

last: Moustafa Ali

first: Vijay

last: Suresh Akhade

last: Thankaswamy Kosalai


title: Gene expression profile of primary human monocyte subsets

city: Bethesda

first: Alice

last: Muglia Thomaz da Silva Amancio

address: Building 33, room 1W10.A3

department: Immunobiology Section

institute: National Institutes of Health (NIH)

summary:
Human primary monocytes are composed of a minor, more mature CD16+(CD14low/neg) population and a major CD16neg(CD14+) subset. The specific functions of CD16+ vs. CD16neg monocytes in steady state or inflammation remain poorly understood. In previous work we found that IL-12 is selectively produced by the CD16+ subset in response to the protozoan pathogen, Toxoplasma gondii. Here we demonstrated that this differential responsiveness correlates with the presence of an IFN-induced transcriptional signature in CD16+ monocytes already at baseline. Consistent with this observation, we found that in vitro IFN-γ-priming overcomes the defect in IL-12 response of the CD16neg subset.
We have anayzed the gene expression profile of primary human CD16+ and CD16neg monocyte subsets cultured for  without or with IFN-gamma

overall_design: Monocytes from three healthy donors were purified from elutriated preparations iwith MACS MicroBeads (Miltenyi Biotec, Bergisch Gladbach, Germany).  For monocyte subset isolation, elutriated preparations were first depleted of granulocytes and NK cells with a mixture of anti-CD15 and CD56 MicroBeads, followed by CD16+ monocytes purification with a CD16pos Monocyte Isolation Kit, and CD16neg monocytes isolation from flow throw using CD14 MicroBeads. Purity of all populations was >95%. Total RNA was extracted from monocytes cultured for 8 hours with or without IFN-γ, and with or without stimulation with Toxoplasma gondii tachyzoytes, by adding Trizol reagent (Life Technologies) and employing the Direct-zol RNA Miniprep kit (Zymo Research, Irvine, CA) according to the manufacturer’s instructions. For broad assessment of gene expression, we used the nCounter® Myeloid Innate Immunity Panel (NanoString Technologies, Seatttle, WA). Total RNA (170 ng) from each sample was hybridized with the probes for 770 genes. Data were processed with nSolver Analysis software (NanoString Technologies) which included assessment of quality of the runs and differential gene expression assessed. The false discovery rate (FDR) was calculated using Benjamini-Yekutieli procedure.

first: Dragana

last: Jankovic

first: Alan

last: Sher


title: Genome-wide occupancy of Arabidopsis SWI/SNF chromatin remodeler SPLAYED provides insights into its interplay with its close homolog BRAHMA and Polycomb proteins

city: London

first: YUHAI

last: CUI

address: 1391 Sandford Street

department: London Research and Development Centre

institute: Agriculture and Agri-Food Canada

summary: SPLAYED (SYD) is a SWItch/Sucrose Non-Fermentable (SWI/SNF)-type chromatin remodeler identified in Arabidopsis thaliana (Arabidopsis). It is believed to play both redundant and differential roles with its closest homolog BRAHMA (BRM) in diverse plant growth and development processes. To better understand how SYD functions, we profiled the genome-wide occupancy of SYD and its impact on the global transcriptome and deposition of trimethylation of histone H3 on lysine 27 (H3K27me3). To map the global occupancy of SYD, we generated a GFP-tagged transgenic line and used it for ChIP-seq experiments, by which more than 6,000 SYD target genes were identified. Through integrating SYD occupancy and transcriptome profiles, we found that SYD preferentially targets to nucleosome-free regions of expressed genes. Further analysis revealed that SYD occupancy peaks exhibit five distinct patterns, which were also shared by BRM and BAF60, a conserved SWI/SNF complex component, indicating the common target sites of these SWI/SNF chromatin remodelers and the functional relevance of such distinct patterns. To investigate the interplay between SYD and Polycomb-group (PcG) proteins, we performed a genome-wide analysis of H3K27me3 in syd-5. We observed both increases and decreases in H3K27me3 levels at a few hundred genes in syd-5 compared to wild-type (WT). Our results imply that SYD can act antagonistically or synergistically with PcG at specific genes. Together, our SYD genome-wide occupancy data and the transcriptome and H3K27me3 profiles provide a much-needed resource for dissecting SYD’s crucial roles in the regulation of plant growth and development.

overall_design: Examination of SYD genome-wide binding, and H3K27me3 profiles in WT and syd-5 14-day-old seedlings.

first: Jie

last: Shu

first: Yuhai

last: Cui


title: Multiple annealing and looping based amplification cycles for digital transcriptomics (MALBAC-DT) of single cells from the developing mouse cortex and hippocampus

city: Cambridge

first: Longzhi

last: Tan

address: 12 Oxford St

department: Chemistry and Chemical Biology

institute: Harvard University

summary: To study the dynamics of gene expression during postnatal development of the mouse forebrain, we performed MALBAC-DT on 3,517 single cells from the mouse cortex and hippocampus at different ages throughout the first 6 postnatal months.

overall_design: We performed MALBAC-DT on 3,517 single cells.


title: A CRISPR/Cas9-engineered ARID1A-deficient human gastric cancer organoid model reveals essential and non-essential modes of oncogenic transformation

status: Public on Jan 05 2021

city: Stanford

first: Andrey

last: Krokhotin

address: 279 Campus Drive W

institute: HHMI

summary: Mutations in ARID1A, encoding a subunit of the BAF chromatin remodeling complex, rank amongst the most common molecular aberrations in human cancer. However, oncogenic consequences of ARID1A mutation in human cells remain poorly defined due to lack of forward human genetic model systems. Here, CRISPR/Cas9 knockout of ARID1A in primary TP53-/-  human gastric organoids induced morphologic dysplasia, tumorigenicity and mucinous differentiation but impaired Wnt/b-catenin signaling. Genetic Wnt pathway activation rescued mucinous differentiation, but not hyperproliferation, suggesting alternative essential pathways of ARID1A-mediated transformation. ARID1A loss induced transcriptional regulatory modules characteristic of MSI and EBV subtype human gastric cancer, including FOXM1 regulation of multiple mitotic regulatory genes and BIRC5/survivin. Convergently, high-throughput compound screening indicated selective vulnerability of ARID1A-deficient organoids to BIRC5/survivin inhibition, functionally implicating BIRC5/survivin as an essential mediator of proliferation following initial ARID1A loss. However, CRISPR inactivation of ARID1A in established gastric cancer cell lines did not elicit sensitivity to BIRC5/survivin inhibitors, suggesting that this pathway may be selectively required during the early establishment of tumorigenesis. Overall, we define distinct pathways downstream of oncogenic ARID1A mutation, with non-essential Wnt-inhibited mucinous differentiation in parallel with essential transcriptional FOXM1/BIRC5-stimulated proliferation, indicating the general utility of organoid-based forward genetic cancer analysis in human cells.

overall_design: mRNA profiles of TP53 KO and TP53/ARID1A DKO gastric organoids

first: Yuang-Hung

last: Lo


title: Cellular response to spinal cord injury in regenerative and non-regenerative stages in Xenopus laevis.

city: Santiago

first: Juan

last: Larraín

address: Portugal 49

department: Cellular and Molecular Biology

country: Chile

institute: Pontificia Universidad Catolica de Chile

summary:
Background: The efficient regenerative abilities at larvae stages followed by a non-regenerative response after metamorphosis in froglets makes Xenopus an ideal model organism to understand the cellular responses leading to spinal cord regeneration.
Methods: We compared the cellular response to spinal cord injury between the regenerative and non-regenerative stages of Xenopus laevis. For this analysis, we used electron microscopy, immunofluorescence and histological staining of the extracellular matrix. We generated two transgenic lines: i) the reporter line with the zebrafish GFAP regulatory regions driving the expression of EGFP, and ii) a cell specific inducible ablation line with the same GFAP regulatory regions. In addition, we used FACS to isolate EGFP + cells for RNAseq analysis.
Results: In regenerative stage animals, spinal cord regeneration triggers a rapid sealing of the injured stumps, followed by proliferation of cells lining the central canal, and formation of rosette-like structures in the ablation gap. In addition, the central canal is filled by cells with similar morphology to the cells lining the central canal, neurons, axons, and even synaptic structures. Regeneration is almost completed after 20 days post injury. In non-regenerative stage animals, mostly damaged tissue was observed, without clear closure of the stumps. The ablation gap was filled with fibroblast-like cells, and deposition of extracellular matrix components. No reconstruction of the spinal cord was observed even after 40 days post injury. Cellular markers analysis confirmed these histological differences, a transient increase of vimentin, fibronectin and collagen was detected in regenerative stages, contrary to a sustained accumulation of most of these markers, including chondroitin sulfate proteoglycans in the NR-stage.   The zebrafish GFAP transgenic line was validated, and we have demonstrated that is a very reliable and new tool to study the role of neural stem progenitor cells (NSPCs). RNASeq of GFAP::EGFP cells has allowed us to clearly demonstrate that indeed these cells are NSPCs. On the contrary, the GFAP::EGFP transgene is mainly expressed in astrocytes in non-regenerative stages. During regenerative stages, spinal cord injury activates proliferation of NSPCs, and we found that are mainly fated to form neurons and glial cells. Specific ablation of these cells abolished proper regeneration, confirming that NSPCs cells are necessary for functional regeneration of the spinal cord.
Conclusions: The cellular response to spinal cord injury in regenerative and non-regenerative stages is profoundly different between both stages. A key hallmark of the regenerative response is the activation of NSPCs, which massively proliferate to reconstitute the spinal cord, and are differentiated into neurons. Also very notably, no glial scar formation is observed in regenerative stages, but a transient, glial scar-like structure is formed in non-regenerative stage animals.

overall_design: Characterization of cell population GFAP::EGFP from the transgenic line Xla.Tg(Dre.gfap:EGFP)Larra in regenerative stages of Xenopus laevis

first: Gabriela

last: Edwards-Faret

first: Karina

last: González-Pinto

first: Arantxa

last: Cebrián-Silla

first: Johany

last: Peñailillo

first: José

last: García-Verdugo


title: RNA-seq analysis of WT and bdt1 in Arabidopsis.

city: Beijing

first: Xin-Jian

last: He

address: No. 7, zhongguancun life science park road, changping district

institute: National Institute of Biological Sciences

summary: Trimethylated histone H3 lysine 27 (H3K27me3) is a repressive histone marker that regulates a variety of developmental processes, including those that determine flowering time. However, relatively little is known about the mechanism of how H3K27me3 is recognized to regulate transcription. We identified BAH domain-containing transcriptional regulator 1 (BDT1) as an H3K27me3 reader. BDT1 is responsible for preventing flowering by suppressing the expression of flowering genes. Mutation of the H3K27me3 recognition sites in the BAH domain disrupted the binding of BDT1 to H3K27me3, leading to de-repression of H3K27me3-enriched flowering genes and an early-flowering phenotype.

overall_design: We perfomed RNA seq on wild type and bdt1 grown for 10 days in long-day conditions. Two biological replicates per genotype were sequenced.


title: Gene expression data from formalin-fixed paraffin-embedded (FFPE) human uveal melanoma tumors; subset early metastasis vs subset no metastasis

city: OSLO

first: Agate

last: Noer

address: Kirkeveien 166

department: Ophthalmology

country: Norway

institute: Oslo University Hospital, Ullevaal

summary:
Uveal melanoma (UM) is an aggressive malignancy, in which nearly 50% of the patients die from metastatic disease. Formalin-fixed paraffin-embedded (FFPE) samples represent a valuable source of tumor tissue. Our aim was to investigate differential DNA methylation correlated to gene expression in relation to survival data.We sought to identify aberrant DNA methylation of genes that could be linked to metastatic disease and poor survival.
Formalin-fixed parafin-embedded human uveal melanoma tumors; subset early metastasis (n=4) vs subset no metastasis (n=4) were examined by differential gene expression analysis (One-Way ANOVA) to identify 1536 transcripts (1394 up- and 142 down-regulated genes) (p<0.05, FC ≥1.5) .

overall_design: 8 FFPE human uveal melanoma samples; subset early metastasis (n=4) and subset no metastasis (n=4) were used to investigate differential DNA methylation correlated to gene expression in relation to histopathological classification and survival data of human uveal melanoma.We sought to identify aberrant DNA methylation of genes that could be linked to metastatic disease and poor survival.

first: Charlotte

last: Ness

first: Kirankumar

last: Katta

first: Øystein

last: Garred

first: Theresa

last: Kumar

first: Ole

last: Olstad

first: Goran

last: Petrovski

first: Morten

middle: C

last: Moe


title: Gene expression profiles of liver sinusoidal Vγ9+Vδ2+ T cells from healthy donors and patients with hepatitis B virus-related chronic liver disease

city: Seoul

first: Min-Seok

last: Rha

address: 50-1 Yonsei-ro, Seodaemun-gu

department: Otorhinolaryngology

country: South Korea

institute: Yonsei University College of Medicine

summary: We report the gene expression profiles of  liver sinusoidal Vγ9+Vδ2+ T cells from healthy donors and patients with hepatitis B virus-related chronic liver disease.

overall_design: Examination of gene expression profiles in liver sinusoidal Vγ9+Vδ2+ T cells from 5 healthy donors and 5 patients with hepatitis B virus-related chronic liver disease.

first: Eui-Cheol

last: Shin


title: A Grad-seq resource of RNA and protein complexes of Pseudomonas aeruginosa including transcripts of phage ΦKZ

city: Würzburg

first: Jörg

last: Vogel

address: Josef-Schneider-Straße 2

department: Institute for Molecular Infection Biology

institute: Würzburg University

summary: Grad-seq of Pseudomonas aerugionosa PAO1 at OD 0.3 with and without infection with ΦKZ bacteriophage

sample_organism:
Phikzvirus phiKZ
Pseudomonas aeruginosa PAO1

overall_design: lysate preparation of Pseudomonas aerugionosa PAO1 at OD 0.3 with and without infection with ΦKZ bacteriophage for 10 min, 10-40% glycerol gradient and fractionation into 21 fractions, RNA extraction, sequencing.

first: Milan

last: Gerovac


title: Diploid chromatin conformation capture (Dip-C) of single cells from the developing mouse cortex and hippocampus

first: Longzhi

last: Tan

country: USA

summary: To study the dynamics of 3D genome structure during postnatal development of the mouse forebrain, we performed Dip-C on 1,954 single cells from the mouse cortex and hippocampus at different ages throughout the first postnatal year.

overall_design: We performed Dip-C on 1,954 single cells and analyzed them with dip-c and hickit packages. We amplified 2544 samples (=26.5 96-well plates), and then filtered out 590 samples with fewer than 100K contacts per sample, which are considered as "empty wells" that were missed by the flow cytometer. The file GSE146397_metadata.excluded_samples.txt lists all excluded sample titles. The 1954 samples included in this study are listed in the file GSE146397_metadata.cells_contacts_100k.txt.gz.


title: llumina 450k array was used to identify DNA methylation data from formalin-fixed paraffine-embedded (FFPE) human uveal melanoma correlated to gene expression, metastasis and poor prognosis

summary:
Uveal melanoma (UM) is an aggressive malignancy, in which nearly 50% of the patients die from metastatic disease. Formalin-fixed paraffin-embedded (FFPE) samples represent a valuable source of tumor tissue. Our aim was to investigate differential DNA methylation in relation to histopathological classification and survival data. In addition, we sought to identify aberrant DNA methylation of genes that could be linked to metastatic disease and poor survival.
23 formalin-fixed paraffin-embedded human uveal melanoma tumours divided into 3 subgroups based on metastasis and survival; (1) 2-4 years “Early metastasis” (n=8), (2) 9-21 years “Late metastasis” (n=7), and (3) Alive or death of other cause ≥18 years  “No metastasis” (n=8). UM tumours were also divided into 3 groups based on tumour cell morphology; (1) Spindle (n=11), (2) Epitheloid (n=6), and (3) Mixed-both spindle and epitheloid (n=6).

overall_design: Methylation data of 23 FFPE human uveal melanoma samples were linked to histopathological classification, metastasis and survival data. Subset early metastasis (n=4) and Subset no metastasis (n=4) were used to investigate differential DNA methylation correlated to gene expression in relation to survival data of human uveal melanoma. We sought to identify aberrant DNA methylation of genes that could be linked to metastatic disease and poor survival.


title: DNA methylation profiling of normal, precursor lesion and cholangiocarcinoma samples

city: Heidelberg

first: Stephanie

last: Roessler

address: Im Neuenheimer Feld 224

institute: Heidelberg University Hospital

type: Methylation profiling by genome tiling array

summary: A detailed understanding of the molecular alterations in the course of cholangiocarcinogenesis is crucial for a better understanding of this disease and may pave the way to early diagnosis and better treatment options for cholangiocarcinoma (CCA) patients.  We analyzed a clinicopathologically well-characterized patient cohort with intraductal papillary (IPNB) or tubulopapillary biliary (ITPN) precursor lesions of the biliary tract. For a large subset of patients, the triplet sample set of non-neoplastic biliary epithelium, IPNB or ITPN, and invasive CCA was available.

overall_design: We conducted genome wide DNA methylation analyses of non-neoplastic bile duct, IPNB or ITPN and invasive CCA.

first: Reka

last: Toth

first: Damian

last: Stichel


title: Revealing nascent RNA processing dynamics with nano-COP

city: Sherbrooke

first: Karine

last: Choquet

state: Québec

address: 3201 rue Jean-Mignault

department: Biochemistry and Functional Genomics

institute: Université de Sherbrooke

summary: During maturation, eukaryotic precursor RNAs undergo processing events including intron splicing, 3’-end cleavage, and polyadenylation. Here, we describe nanopore analysis of CO-transcriptional Processing (nano-COP), a method for probing the timing and patterns of RNA processing. An extension of native elongating transcript sequencing (NET-seq), which quantifies transcription genome-wide through short-read sequencing of nascent RNA 3’ ends, nano-COP uses long-read nascent RNA sequencing to observe global patterns of RNA processing. First, nascent RNA is stringently purified through a combination of 4-thiouridine metabolic labeling and cellular fractionation. In contrast to cDNA or short-read–based approaches relying on reverse transcription or amplification, the sample is sequenced directly through nanopores to reveal the native context of nascent RNA. nano-COP identifies both active transcription sites and splice isoforms of single RNA molecules during synthesis, providing insight into patterns of intron removal and the physical coupling between transcription and splicing. The nano-COP protocol yields data within 3 days.

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct

overall_design: Stringent purification of nascent RNA and direct RNA sequencing on Oxford Nanopore Technologies platforms

first: Heather

middle: L

last: Drexler

first: Hope

middle: E

last: Merens

first: Paul

last: Tang

first: Jared

last: Simpson

first: L Stirling

last: Churchman


title: Predicting Changes in Renal Metabolism after Compound Exposure with a Genome-Scale Metabolic Model

city: Charlottesville

first: Jason

last: Papin

state: VA

address: 415 Lane Rd

department: Biomedical Engineering

institute: University of Virginia

summary: The kidneys are metabolically active organs that are important for several physiological tasks such as the secretion of soluble wastes into urine. Other functions of the kidneys include synthesizing glucose and oxidizing fatty acids for energy in fasting (non-fed) conditions. Once damaged, the metabolic capability of the kidneys becomes altered. Here, we define metabolic tasks in a computational modeling framework to capture kidney function in an update to the iRno network reconstruction of rat metabolism using literature-based evidence. To demonstrate the utility of iRno for predicting kidney function, we exposed primary rat renal proximal tubule epithelial cells to five compounds with varying levels of nephrotoxicity (acetaminophen, carbon tetrachloride, gentamicin, 2,3,7,8-tetrachlorodibenzodioxin, and trichloroethylene for six and twenty-four hours), and collected transcriptomics and metabolomics data to measure the metabolic effects of compound exposure. We observed changes in fatty acid metabolism and amino acid metabolism, consistent with changes in existing markers of kidney function such as Lcn2 (lipocalin), Clu (clusterin), and the carnitine transporter Octn2 (solute carrier family 22, member 5). The iRno metabolic network reconstruction was able to predict alterations in these same pathways after integrating transcriptomics data, and was able to distinguish between select compound-specific effects on the proximal tubule epithelial cells. Genome-scale metabolic network reconstructions with coupled omics data can be used to predict changes in metabolism to provide a step towards identifying novel biomarkers of kidney function and dysfunction.

overall_design: Examination of RNA from compound-treated primary rat renal proximal tubule epithelial cells for 6 conditions (including the control) with 3 replicates for each condition.

first: Kristopher

last: Rawls

first: Bonnie

middle: V

last: Dougherty

first: Glynis

last: Kolling

first: Kalyan

last: Vinnakota

first: Venkat

last: Pannala