title: Transcriptome -Wide m6A Methylome and Potential Drug Targets Prediction Analysis of Rheumatoid Arthritis

city: Beijign

first: Danping

last: Fan

address: Yinghua Donglu

institute: Institute of Clinical Medical Science, China-Japan Friendship Hospital

summary: This research aims at assessing the m6A methylome in the RA peripheral blood mononuclear cells (PBMCs) and perform potential drug targets prediction analysis. Five RA samples and ten control samples were obtained from China-Japan Friendship Hospital. The various expression of m6A methylation and genes in RA and control group were compared with methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing (RNA-seq). Bioinformatics explorations were also made to explore the enriched biological roles and paths of the differentially expressed m6A methylation and genes. Potential drug targets prediction and validation were also constructed to provide potential therapeutic targets for RA. In total, 583 dysregulated m6A peaks, of which 295 were greatly upregulated and 288 were greatly downregulated, were identified. Similarly, 1570 differentially genes were acquired by RNA-seq, including 539 upregulated and 1031 downregulated. According to deeper joint exploration, the m6A methylation and mRNA expression degrees of 35 genes varied greatly, of which including 13 hyper-methylated as well as upregulated genes (hyper-up), 12 hyper-methylated and downregulated genes (hyper-down), 5 hypo-methylated as well as upregulated genes (hypo-up), and 5 hypo-methylated and downregulated genes (hypo-down) GO and KEGG explorations showed the enrichment of these distinct genes in “protein transport” “protein binding” and “lysosome”. In addition, the mRNA levels of TUBB2A, IGF2BP3, DYNC1I1 and FOSL1 were increased and consistent with the trend of our sequencing results. While after the treatment of TP and MTX, their mRNA levels were decreased. This research set up the transcriptional map of m6A in RA PBMCs and displayed the hidden association between RNA methylation alternation and associated genes in RA. The outcomes highlight the importance of m6A modification as a gene regulatory system in RA. Furthermore, TUBB2A, IGF2BP3, DYNC1I1 and FOSL1 might be potential therapeutic targets of TP and MTX during RA treatment.

overall_design: Five patients suffering from RA were obtained. There were 10 subjects without RA or other autoimmune diseases enrolled, in which 5 control samples were used for MeRIP-seq, the other 5 control samples were used for experiments. The various expression of m6A methylation and genes in RA and control group were compared with methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing (RNA-seq). Bioinformatics explorations were also made to explore the enriched biological roles and paths of the differentially expressed m6A methylation and genes. Potential drug targets prediction and validation were also constructed to provide potential therapeutic targets for RA.

first: Cheng

last: Xiao

first: Yuan

last: Xu

first: Bailiang

last: Wang

first: Bin

last: Liu

first: Xiaofeng

last: Gu

first: Qian

last: Zhang

first: Qinbin

last: Ye

first: Xiaoyu

last: Xi

first: Ya

last: Xia

first: Qiong

first: Zheng


title: ZEB1 transcription factor promotes immune escape in melanoma

status: Public on Mar 23 2022

city: LYON

first: Julie

last: CARAMEL

address: 28 Rue Laennec

country: France

institute: Cancer Research Center of Lyon

type: Expression profiling by high throughput sequencing

summary: The efficacy of immunotherapies in metastatic melanoma depends on a robust T cell infiltration. Consistently, defining cancer cell intrinsic mechanisms mediating T cell exclusion and immune resistance is crucial. The EMT inducing transcription factor ZEB1 is a major regulator of melanoma cell plasticity, driving resistance to MAPK targeted therapies. Here, by analyzing the immune infiltrates of a cohort of melanoma patients, we demonstrate that high ZEB1 expression in tumor cells is associated with a decrease in CD8+ T lymphocyte infiltration, independently of beta-catenin pathway activation. Moreover, gain- or loss-of-function experiments in melanoma mouse models show that ZEB1 regulates tumor growth by controlling CD8+ T cell recruitment, via its negative action on the production of T cell attracting chemokines, and that its targeting improves the efficacy of anti-PD1 immunotherapy. Overall, the major role of ZEB1 in preventing T cell infiltration suggests it may constitute a new target in metastatic melanoma.

overall_design: RNA was extracted from cutaneous biopsies of melanoma samples. Bulk RNA-sequencing was performed on the FFPE samples.

last: Caramel


title: Next Generation Sequencing analysis at single-cell level of normal and degenerated nucleus pulposus cells transcriptomes

city: Wuhan

first: gaocai

last: li

state: HuBei

address: JIefang Avenve 1277, Jianghan District

department: Department of Orthopeadics

country: China

institute: wuhan union hospital

type: Non-coding RNA profiling by high throughput sequencing

summary: Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived retinal transcriptome profiling (RNA-seq) to microarray and quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods and to evaluate protocols for optimal high-throughput data analysis

overall_design: nucleus pulposus cells mRNA profiles of five degenerated and four normal human

first: Gaocai

last: Li

first: Cao

last: Yang

first: Kun


title: Response to cisplatin upon depletion of SNRNP200

status: Public on Mar 24 2022

city: Montréal

first: Alexandre

last: Orthwein

state: QC

address: 3755 Chemin de la Côte-Sainte-Catherine

department: Lady Davis Institute

country: Canada

institute: McGill University

summary: The goal is to define the impact of depleting SNRNP200 on the transcriptional response to the DNA damaging agent cisplatin

overall_design: This experiment contains duplicate for each condition, two distinct siRNAs targeting SNRNP200 compared to one scramble control and two time points for drug treatment (6 and 24hrs)


title: RRBS DNA methylation data of bovine embryonic fibroblast cells

city: Auburn

first: Wellison

middle: J. S.

last: Diniz

state: Alabama

address: CASIC Building 559 Devall Drive Auburn University, 205

department: Animal Sciences

country: USA

institute: Auburn University

type: Methylation profiling by high throughput sequencing

summary: We investigated the effects of one carbon metabolites supplementation on early embryonic development. To this end, the Bovine Embryonic Tracheal Fibroblast cell lines (EBTr; NBL-4; ATCC CCL-44) were cultured under different levels of glucose and OCM (folic acid, choline chloride, vitamin B12, and L-methionine).

overall_design:
The EBTr cells were cultured in Eagle’s Minimum Essential Medium, supplemented with 10% fetal bovine serum, 1% penicillin-streptomycin and 0.11 g/L Na pyruvate (Sigma). Treatments were arranged as a completely randomized design with a two glucose levels [1 g/L (low) or 4.5 g/L (high)] × 3 OCM levels [control, 2.5X, and 5X]. Control medium contained basal concentrations of folate (0.001g/L), choline (0.001 g/L), vitamin B12 (4 µg/L), and methionine (0.015 g/L). One-carbon metabolites (folic acid, choline chloride, vitamin B12, and L-methionine) were supplemented to the media to achieve 2.5 or 5 times.
For all treatments, cells were plated into T-75 culture flasks and allowed to become 90% confluent. Once target confluence was reached, cells were removed from the flask and cell pellets (1 × 10^6) were frozen in cryo vials (n = 3 tubes/trt) and shipped to the University of Minnesota Genomics Center. The DNA was extracted from the cell pellets with the DNeasy Blood and Tissue Kit (Qiagen, Hilden, Germany) and quantified using the PicoGreen DNA Quantification Kit (Invitrogen). Libraries (n = 18) were prepared from extracted DNA (20 ng/µL) using the NuGEN Ovation RRBS Methyl-Seq Kit (Tecan Genomics, Redwood City, CA). Libraries were bisulfite converted and PCR amplified prior to sequencing with NovaSeq S Prime in the 150 paired-end reads mode (Illumina, San Diego, CA).
After sequencing, raw data quality control was performed using the FastQC v0.11.8 and MultiQC v1.9 software. Adapter trimming and low-quality bases (Phred score < 20) were filtered out using Cutadapt v.2.10. NuGEN’s diversity trimming was carried out to remove diversity adaptors. Trimmed reads were mapped to the bovine reference genome (UCSC - bosTau8, Illumina IGenome) using the Nextflow methylseq pipeline v1.5. Differentially methylated cytosines (DMC) were identified by pair-wise comparison using edgeR v. 3.24.3, and considered significant for each of the contrasts when the P-value cutoff < 0.01.

middle: J

first: Matthew

middle: S

last: Crouse

first: Joel

last: Caton

first: Kate

last: Claycombe-Larson

first: Amanda

middle: K

last: Lindholm-Perry

first: Lawrence

middle: P

last: Reynolds

first: Carl

middle: R

last: Dahlen

first: Pawel

last: Borowicz

first: Alison

last: Ward


title: SB431452 affects extracellular matrix during induction of hiPSC-derived gastric organoid with muscularis mucosa

status: Public on Mar 22 2022

city: Kobe City

first: Michiyo

last: Koyanagi-Aoi

address: Chuo-ku, Kusunoki-cho 7-5-1

country: Japan

institute: Kobe University

summary: We report TGFβ signaling is involved in muscularis mucosa differentiation of hiPSC-derived gastric organoids.

overall_design: Comparison of RNA expression of hiPSC-derived gastric organoid treated with or without SB431542

first: Keiichiro

last: Uehara

first: Takashi

last: Aoi


title: Time course RNA−seq analysis during gastric organoids with muscularis mucosa induction derived from hiPSC

summary: We report time course analysys during gastric organoids with muscularis mucosa induction derived from hiPSC by RNA-seq.

overall_design: Comparison of RNA expression of hiPSC, hiPSC-derived gastric organoids (week1, week3, week6, week8) and stomach


title: Targeted paired transcriptomic analysis of head and neck carcinoma treated with chemoradiotherapy

city: Ulm

first: Johannes

last: Doescher

state: Deutschland

address: Frauensteige 12

department: Otorhinolaryngology, Head and Neck Surgery

country: Germany

institute: University hospital of Ulm

summary: We employed contemporary targeted autoimmune RNA sequencing (HTG molecular diagnostics, Precision Immuno-Oncology Panel) to head and neck tissue derived from 30 cancer patients patients before and after chemoradiotherapy. Differential gene expression was calculated and compared to therapy response.

overall_design:
Transcriptomic analysis of 60 samples from 30 head and neck cancer patients before and after CRT.
***Please note that raw data is not provided as raw files, basecall and fastq are not meaningful and can't be used by other researchers unless they are demultiplexed, parsed and aligned byt the manufacturer's software. With this technology, thousands of carefully designed probes (covering different isoforms) against thousands of genes were used and then after a Nuclease protection assay, all that is left are probes that are bound to their targets. Then a normal NGS library prep follows and in actuality the probes were sequenced and the number of reads per probe is used as an absolute quantification of mRNA transcripts per target. The produced fastq files are parsed in the manufacturer's software which aligns and counts reads by looking for the sequence of the probes (~100nt).

first: Adrian

last: von Witzleben

first: Konstantinos

last: Boukas

first: Stephanie

middle: E

last: Weissinger

first: Gareth

last: Thomas

first: Simon

last: Laban

first: Jaya

first: Thomas

last: Hoffmann

first: Christian

middle: H

last: Ottensmeier


title: Transcriptional CDK inhibitors as potential treatment option for testicular germ cell tumors

status: Public on Mar 26 2022

city: Bonn

first: Kai

last: Funke

state: NRW

address: Venusberg-Campus 1

department: Pathology, Developmental Pathology

institute: University Hospital Bonn

summary: Type II testicular germ cell tumors (TGCT) are the most frequently diagnosed solid malignancy in young men. Up to 15 % of patients with metastatic non-seminomas show cisplatin resistance having a very poor survival rate due to lacking treatment options. Transcriptional cyclin-dependent kinases (CDK) have been shown to be effective targets in treatment of different types of cancer. Here, we investigated the effects of the CDK inhibitors dinaciclib, flavopiridol, YKL-5-124, THZ1, NVP2, SY0351 and THZ531. XTT viability assay revealed strong cytotoxic impact of CDK7/ 12/ 13 inhibitor SY0351 and CDK9 inhibitor NVP2 on the TGCT wild type cell lines (2102EP, NCCIT, TCam2) and the cisplatin resistant cell lines (2102EP-R, NCCIT-R). The CDK7 inhibitor YKL-5-124 showed strong impact on 2102EP, 2102EP-R, NCCIT and NCCIT-R cell lines leaving the MPAF control cell line mostly unaffected. FACS based analysis revealed mild effects on the cell cycle of 2102EP and TCam2 cells after SY0351, YKL-5-124 or NVP2 treatment. Molecular analysis showed a cell line specific response for SY0351 and NVP2 inhibition while YKL-5-124 induced similar molecular changes in 2102EP, TCam2 and MPAF cells. Thus, after TGCT subtype determination CDK inhibitors might be a potential alternative for optimized and individualized therapy independent of chemotherapy sensitivity.

overall_design: Total RNA obtained from cell lines treated for 1 or 24 hours with CDK inhibitors compared to DMSO treated controls. Total 108 samples with n=3 for each condition.

first: Robert

last: Düster

first: Prince

last: De-Graft Wilson

first: Lena

last: Arévalo

first: Matthias

last: Geyer

first: Hubert

last: Schorle


title: Gene expression analysis during differentiation into gastric organoids with muscularis mucosa from hiPSC

first: Michiyo

last: Koyanagi-Aoi

summary: We induced gastric organoids with muscularis mucosa from hiPSC.

overall_design: Comparison of the hiPSC, hiPSC-derived gastric organoids (Day7, Day20, Day43 and Day56) and stomach

first: Keiichiro


title: The CHARGE syndrome ortholog CHD-7 regulates TGF-b pathways in C. elegans

city: Bethesda

first: Sijung

last: Yun

state: Maryland

address: 8908 Ewing Drive

institute: NIH/JHU

summary: CHARGE syndrome is a complex developmental disorder caused by mutations in the chromodomain helicase DNA-binding protein7 (CHD7) and characterized by retarded growth and malformations in the heart and nervous system. Despite the public health relevance of this disorder, relevant cellular pathways, and targets of CHD7 that relate to disease pathology are still poorly understood. Here we report that chd-7, the nematode ortholog of Chd7, is required for dauer morphogenesis, lifespan determination, stress response and body size determination. Consistent with our discoveries, we found chd-7 to be allelic to scd-3, a previously identified dauer suppressor from the DAF-7/TGF-E pathway. Epistatic analysis places CHD-7 at the level of the DAF-3/DAF-5 complex, but we found that CHD-7 also directly impacts the expression of multiple components of this pathway. Transcriptomic analysis revealed that chd-7 mutants fail to repress daf 9 for execution of the dauer program. In addition, CHD-7 regulates the DBL-1/BMP pathway components and shares roles in male tail development and cuticle synthesis. To explore a potential conserved function for chd-7 in vertebrates, we used Xenopus laevis embryos, an established model to study craniofacial development. Morpholino mediated knockdown of Chd7 led to a reduction in col2a1 mRNA levels, a collagen whose expression depends on TGF-E signaling. Both embryonic lethality and craniofacial defects in Chd7- depleted tadpoles were partially rescued by over-expression of col2a1 mRNA. We suggest that Chd7 has conserved roles in regulation of the TGF-E signaling pathway and pathogenic Chd7 could lead to a defective extracellular matrix deposition.

overall_design: RNA-seq on chd-7;daf-2 vs. daf-2

first: Judith

middle: L

last: Yanowitz

first: Daniel

last: Hochbaum


title: Chromatin Immunoprecipation Sequencing in 3T3-L1 preadipocytes using Par-4 antibody

status: Public on Mar 25 2022

city: LEXINGTON

first: Vivek

middle: M.

last: Rangnekar

state: KY

address: 789 s limestone

institute: University of Kentucky

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Prostate apoptosis response-4 (Par-4) is a tumor suppressor protein that is extensively investigated in cancer. The objective of this study is to determine the physiological function of Par-4 in normal cells. Our findings indicated that genetic loss of Par-4 in mice primarily results in adipocyte hypertrophy and obesity, and secondary leads to hepatic steatosis and insulin resistance. Moreover, we noted that Par-4 is downregulated in human subjects who are likely to become obese in the future, thereby serving as a predictor of obesity. Importantly, ChIP-Seq indicated that MDM2 is a target of Par-4 protein, which is known to function as a transcriptional coregulator.  We show that Par-4 loss upregulates MDM2 target protein p53, and that p53 further induces complement factor C3 and its proteolytic fragment acylation stimulating protein (ASP), an adipokine that is known to be causally associated with obesity. These studies led to the identification of the Par-4-MDM2-p53-C3/ASP axis in regulation of obesity by Par-4.

overall_design:
We report potential gene targets of Par-4 upon sequencing of ChIP in 3T3-L1 using Par-4 antibody.
Sequencing of pulled-down Par-4 antibody-bound chromatin.

first: James

last: Sledziona

first: Nathalia

last: Araujo

middle: M


title: Alteration of N6-Methyladenosine mRNAMethylation in an hiPSC-CMs Model of TKI-induced cardiotoxicity

status: Public on Mar 21 2022

city: Beijing

first: Dong

last: Han

address: No. 28, Fuxing Road, Haidian District

institute: 301 PLA hospital

summary: We performed a genome-wide analysis of m6A mRNA methylation in SUN-treated human pluripotent stem cells induced myocardial cells (hiPSC-CMs) by methylated RNA immunoprecipitation sequencing (MeRIP-seq).

overall_design: In this study, we used methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA sequencing (RNA-seq) to study the transcriptome and m6A modification sites of human pluripotent stem cells induced myocardial cells (hiPSC-CMs) treated with sunitinib (SUN). Through MeRIP-Seq, we identified a total of 2615 peaks with significant changes, of which 1694 were significantly up-regulated and 921 were significantly down-regulated.

first: Han

last: Dong

first: Ma

last: Yan


title: Whole transcriptome sequencing of major depressive disorder (MDD) patients and healthy controls

city: Zhanjiang

first: Yujie

last: Cai

state: Guangdong

address: Renmindadaonan 57th

institute: Affiliated Hospital of Guangdong Medical University

summary: We do this study to identify and validate the differentially expressed circRNAs in MDD patients and to evaluate their potential as diagnostic biomarkers and as novel therapeutic targets for MDD, to explore the potential clinical value and possible mechanism of circRNAs in MDD.

overall_design: Whole transcriptome sequencing of 4 MDD patients and 4 healthy controls

first: Dandan

last: Zhang

first: Yao

last: Ji


title: The effect of miR-322/-503 on myoblast differentiation in DM1 murine cell model

city: Wuhu

first: Xiaopeng

last: Shen

address: 1 East Beijing Rd.

institute: Anhui Normal University

summary: To study the gene expression profile difference caused by miR-322/-503 overexpression in the myoblast differentiation of DM1 group, we performed RNA-seq on the total RNA samples collected from the in vitro myoblast differentiation day 4 of control and miR-322/-503 overexpressing DM1 C2C12 cell models. The DM1 cell model was bulit by stably transfecting C2C12 cells with  GFP-CUG200 plasmid. Each group contained three biological replicates. The expression matrix was obtained by Hisat2 followed by Stringtie.

overall_design: RNA-seq of control and miR-322/-503 overexpressing DM1 C2C12 cell differentiated for 4 days


title: Interaction of modified oligonucleotides with nuclear proteins, formation of novel nuclear structures and sequence-independent effects on RNA processing

city: Parkville

first: Jack

middle: Aatto Laasonen

last: Cooper

state: Victoria

address: 49 College Crescent

country: Australia

institute: University of Melbourne

summary:
Oligonucleotides and nucleic acid analogues that alter gene expression are showing therapeutic promise for selected human diseases. The modification of synthetic nucleic acids to protect against nuclease degradation and to influence drug function is common practice, however, such modifications may also confer unexpected physicochemical and biological properties. Here we report backbone-specific effects of modified oligonucleotides on subnuclear organelles, altered distribution of nuclear proteins, the appearance of novel structured nuclear inclusions, and modification of RNA processing in cultured cells transfected with antisense oligonucleotides on a phosphorothioate backbone. Phosphodiester and phosphorodiamidate morpholino oligomers elicited no such consequences. Disruption of subnuclear structures and proteins elicit severe phenotypic disturbances, revealed by transcriptomic analysis of fibroblasts exhibiting such disruption. These data suggest that the toxic effects and adverse events reported after clinical evaluation of phosphorothioate nucleic acid drugs may be mediated, at least in part, by non-specific interaction of nuclear components with the phosphorothioate backbone.
The effects of antisense oligonucleotide transfection on morphology of subnuclear organelles, localisation of nuclear proteins and impact on RNA processing were examined in primary human cells.

overall_design: The effects of antisense oligonucleotide transfection on RNA processing were examined in primary human cells.

first: Loren

last: Flynn

first: Ruohan

last: Li

first: May

middle: T

last: Aung-Htut

first: Ianthe

last: Pitout

middle: A

first: Alysia

last: Hubbard

first: Lisa

last: Griffiths

first: Charlie

last: Bond

first: Steve

middle: D

last: Wilton

first: Archa

last: Fox

first: Sue

last: Fletcher


title: Unique molecular features and cellular responses differentiate two populations of motor cortical Layer 5b neurons in a preclinical model of ALS.

city: New York

first: Eric

middle: F

last: Schmidt

state: NY

address: 1230 York Avenue

institute: The Rockefeller University

summary: A major paradox of neurodegenerative disorders such as amyotrophic lateral sclerosis (ALS) is that many genes linked to disease are ubiquitously expressed, yet only discrete subtypes of cells degenerate. Therapeutic advancement depends on understanding unique molecular responses of vulnerable cells in the context of disease-causing mutations. Here, we describe two bacTRAP lines, Gprin3 and Colgalt2, that label distinct populations of projection neurons in layer 5b (L5b) of motor cortex. We demonstrate that these cells can be distinguished not only by their anatomical features, but by their underlying molecular properties. In the SOD1*G93A preclinical ALS model, both populations showed molecular responses in common, but Gprin3 corticospinal population showed additional, unique responses in disease that may reflect their disease vulnerability since Gprin3 cells were lost during disease progression while Colgalt2 cells were spared. These results tease apart molecular pathways that may contribute to ALS pathology and provide a framework for elucidating selective vulnerability.

overall_design: The translating ribosome affinity purification (TRAP) method was used to collect polysome-bound mRNAs were collected from two subpopulations of layer 5b pyramidal neurons using Gprin3-bacTRAP mice or  Colgalt2-bacTRAP mice. TRAP IPs were compared to whole tissue mRNAs (input) or TRAP IP mRNAs were compared between cell types (Gprin3 and Colgalt2), or between transgenic ALS mice (SOD1*G93A) and wildtype littermates within each cell type (Gprin3 and Colgalt2).  We collected 2-5 replicates for each experimental group.

first: Nathaniel

last: Heintz


title: Genome-wide maps of chromatin state in  WT and Whsc1 ko or Nlrc5  ko CT26 cells.

city: shanghai

first: jiale

middle: jia

last: ren

state: shanghai

address: yueyang road 320

department: shanghai institute of nutrition and health

institute: CAS

summary: Histone dimethyl transferase WHSC1 drives the transcription of MHC-I machinery in mouse and human colorectal cancer cells (CRCs); thus, WHSC1 downregulation potentiates CRCs to escape from cytotoxic CD8+ T cell responses. WHSC1 directly interacts with MHC-I transactivator, NLRC5 to selectively stimulate MHC-I gene expression. Thus, silencing Whsc1 diminished MHC-I level, impaired anti-tumor immunity and blunted immunotherapy efficacy.

overall_design: ChIP-seq:Examination of H3k36me2 histone modifications in WT and Whsc1 KO or Nlrc5 KO cells.

first: Jiale

last: Ren

first: Ni

first: Siyu

last: Pei

first: Yannan

last: Jiacheng Guo


title: Local euchromatin enrichment in lamina-associated domains anticipates their re-positioning in the adipogenic lineage

city: Oslo

first: Philippe

last: Collas

address: PO Box 1112 Blindern

department: Institute of Basic Medical Sciences

country: Norway

institute: University of Oslo

summary: Interactions of chromatin with the nuclear lamina via lamina-associated domains (LADs) confers structural stability to the genome. The dynamics of positioning of LADs during differentiation, and how LADs impinge on developmental gene expression, remains elusive, however. We examined changes in the association of lamin B1 with the genome in the first 72 hours of differentiation of adipose stem cells into adipocytes. We demonstrate a repositioning of entire stand-alone LADs and of LAD edges as a prominent nuclear structural feature of early adipogenesis. Whereas adipogenic genes are released from LADs, LADs sequester downregulated or repressed genes irrelevant for the adipose lineage. However, LAD repositioning only partly concurs with gene expression changes. Differentially expressed genes in LADs, including LADs conserved throughout differentiation, reside in local euchromatic and lamin-depleted sub-domains. In these sub-domains, pre-differentiation histone modification profiles correlate with the LAD versus inter-LAD outcome of these genes during adipogenic commitment. In addition, differentially expressed genes in LADs are linked to short-range enhancers which overall co-partition with these genes in LADs versus inter-LADs during differentiation. We conclude that LADs are predictable structural features of adipose nuclear architecture that restrain non-adipogenic genes in a repressive environment.

overall_design: We have analyzed nuclear lamin-chromatin contacts throughout in vitro adipose induction of human primary adipose stem cells (ASCs).  We have mapped lamina-associated domains (LADs) by ChIP-seq of lamin B1 on days 0 (here, T00), 1 (T24) and 3 (T72) of differentiation. LADs were also mapped from proliferating (Pro) ASCs in culture, before induction of cell cycle arrest at T00. Gene expression profiles were analyzed by RNA-sequencing at 0, 4, 8, 12, 16, 20, 24 and 72 h after differentiation induction. Histone H3K4me1, H3K4me3 and H3K27ac profiles were determined by ChIP-seq at the same timepoints between 0 and 24 h (T00 to T24). We set out to profile and characterize the fate (outcome) of LADs from T00 to T72, relate LAD changes to geen expression changes in concerved (constitutive) cLADs and in variable vLADs, and relate these changes to histone modification profiles.

first: Mohamed

last: Abdelhalim

first: Julia

last: Madsen-Østerbye

first: Marie-Odile

last: Baudement


title: Second generation molecules targeting the oncogenic signal mediator, RHOA in gastric cancer

city: Incheon

first: Seungyoon

last: Nam

state: Incheon

address: Namdong-gu, Namdongdae-ro 774-21

country: South Korea

institute: Gachon University Colleage of Medicine

summary: Previously, we designed pharmacologically optimized inhibitor RHOA, a major signaling hub in gastric cancer (GC), the fourth most common cause of death in the world. Here, based on that previous work, we perform the lead optimization and design new RHOA inhibitors for greater anti-GC potency. Two of these compounds, JK-206 and -312 inhibit cell viability and migration of GC cell lines. Furthermore, through our transcriptomics analysis of JK-206 treatment in GC cells, we revealed that pharmacologic inhibition of RHOA was associated with inhibition of mitogenic pathway, including BIRC5. Thus, RHOA→BIRC5 can be regarded as a new therapeutic strategy in GC, possibly regulating proliferation and migration in oncogenic mechanisms.

overall_design: The Affymetrix Whole transcript Expression array process was executed according to the manufacturer's protocol (GeneChip Whole Transcript PLUS reagent Kit). cDNA was synthesized using the GeneChip WT (Whole Transcript) Amplification kit as described by the manufacturer. The sense cDNA was then fragmented and biotin-labeled with TdT (terminal deoxynucleotidyl transferase) using the GeneChip WT Terminal labeling kit. Approximately 5.5 μg of labeled DNA target was hybridized to the Affymetrix GeneChip Human 2.0 ST Array at 45°C for 16hour. Hybridized arrays were washed and stained on a GeneChip Fluidics Station 450 and scanned on a GCS3000 Scanner (Affymetrix). Signal values were computed using the Affymetrix® GeneChip™ Command Console software.

first: Sungjin

last: Park

first: Hyo

last: Eom

first: Yon

last: Kim


title: TOX4 Facilitates Promoter-Proximal Pausing and C-Terminal Domain Dephosphorylation of RNA Polymerase II in Human Cells

status: Public on Mar 29 2022

city: Shanghai

first: Aiwei

last: Wu

state: Shanghai

address: 800 Dongchuan Road

department: School of Life Sciences and Biotechnology

institute: Shanghai Jiao Tong University

summary: TOX4 is one of the regulatory proteins of PP1 phosphatases with poorly understood functions. Here we show that chromatin occupancy pattern of TOX4 resembles that of RNA polymerase II (Pol II), and its loss increases cellular level of C-terminal domain (CTD) phosphorylated Pol II but mainly decreases Pol II occupancy on promoters. In addition, elongation rate analyses by 4sUDRB-seq suggest that TOX4 restricts pause release and early elongation but promotes late elongation. Moreover, TT-seq analyses indicate that TOX4 loss mainly decreases transcriptional output. Mechanistically, TOX4 may restrict pause release through facilitating CTD serine 2 and DSIF dephosphorylation, and promote Pol II recycling and reinitiation through facilitating CTD serines 2 and 5 dephosphorylation. Furthermore, among the PP1 phosphatases, TOX4 preferentially binds PP1α and is capable of facilitating Pol II CTD dephosphorylation in vitro. These results lay the foundation for a better understanding of the role of TOX4 in transcriptional regulation.

overall_design: Study the roles of TOX4 in transcriptional regulation

first: Ming

last: Yu

first: Ziling

last: Liu

first: Talang

last: Wang


title: Collateral Responses to Classical Cytotoxic Chemotherapies are Heterogeneous and Sensitivities are Sparse

city: Cambridge

first: Simona

last: Dalin

state: MA

address: 415 Main Street, 4017b

institute: Broad Institute

summary: Chemotherapy resistance is a major obstacle to curing cancer patients. Combination drug regimens have shown promise as a method to overcome resistance; however, to date only some cancers have been cured with this method. Collateral sensitivity – the phenomenon whereby resistance to one drug is co-occurrent with sensitivity to a second drug – has been gaining traction as a promising new concept to guide rational design of combination regimens. Here we evolved over 100 subclones of the Eµ-Myc; p19ARF -/- cell line to be resistant to one of four classical chemotherapy agents: doxorubicin, vincristine, paclitaxel, and cisplatin. We then surveyed collateral responses to acquisition of resistance to these agents. Although numerous collateral sensitivities have been documented for antibiotics and targeted cancer therapies, we observed only one collateral sensitivity: half of cell lines that acquired resistance to paclitaxel also acquired a collateral sensitivity to verapamil. However, we found that the mechanism of this collateral sensitivity was unrelated to the mechanism of paclitaxel resistance. Interestingly, we observed heterogeneity in the phenotypic response to acquisition of resistance to most of the drugs we tested, most notably for paclitaxel, suggesting the existence of multiple different states of resistance. Surprisingly, this phenotypic heterogeneity in paclitaxel resistant cell lines was unrelated to transcriptomic heterogeneity among those cell lines. These features of phenotypic and transcriptomic heterogeneity must be taken into account in future studies of treated tumor subclones and in design of chemotherapy combinations.

overall_design: Gene expression in 17 paclitaxel resistant cell lines, 12 DMSO control lines, and one parental line.

first: Beatrice

last: Grauman-Boss

first: Douglas

last: Lauffenburger

first: Michael

last: Hemann


title: An association study between somatic alterations and eQTL in immunity, DNA repair and inflammation pathways in esophageal squamous cell carcinoma (ESCC)

first: Maxwell

last: Lee

state: MD

address: Building 37, Room 5120

department: CCR

institute: National Cancer Institute, NIH

summary: Since ESCC is a complex and heterogeneous disease, it is increasingly important that analyses combine the evaluation of alterations in DNA, including somatic copy number alterations (SCNA), which distinguished from germ line copy number variants, allelic imbalance and LOH, and RNA, including those that occur in both mRNA and miRNA on the same specimen. Thus, in order to better understand their potential interactions in the development of cancers, in the present study, we studied alterations at DNA level as well as expression of mRNA and target miRNA in tumors and their matched normal tissues from 56 ESCC cases on a candidate tumor suppressor gene CSMD1 to characterize there alterations in this gene for import insights into the cellular detect that cause the complex disease, ESCC.

overall_design: Affymetrix SNP arrays were performed according to the manufacturer's directions on DNA extracted from esophageal squamous cell carcinoma diagnosed or peripheral blood samples.

first: Howard

last: Yang

first: Huaitian

last: Hu

first: Hua

last: Su

first: Chaoyu

first: Carol

last: Giffen

first: Philip

last: Taylor

first: Alisa

last: Goldstein

first: Max


title: Gill and liver transcript expression changes associated with gill damage in Atlantic salmon (Salmo salar)

city: St. John's

first: Mohamed

last: Emam

state: NL

address: 0 Marine Lab Rd, St. John's, NL

institute: Memorial University

summary: Gills of teleost fish represent a vital multifunctional organ; however, they are subjected to environmental stressors, causing gill damage. Gill damage is associated with significant losses in the Atlantic salmon aquaculture industry. Gill disorders due to environmental stressors are exacerbated by global environmental changes, especially with open-net pen aquaculture (as farmed fish lack the ability to escape those events). The local and systemic response to gill damage, concurrent with several environmental insults, are not well investigated. We performed field sampling to collect gill and liver tissue after several environmental insults. Using a 44K salmonid microarray platform, we aimed to compare the transcriptomes of pristine and moderately damaged gill tissue. The gill damage-associated biomarker genes and associated qPCR assays arising from this study will be valuable in future research aimed at developing therapeutic diets to improve farmed salmon gill health.

overall_design:
Using a 44K salmonid microarray platform, we aimed to compare the transcriptomes of pristine and moderately damaged gill tissue.
The fish were sampled on the 13th and 14th of November 2017. The average weights for all of the collected fish was 1348 grams. The gill samples were collected from the net pen-grown Atlantic salmon located in BC. Fish were randomly netted then euthanized using 400 mg L−1 tricaine-methane sulfonate bath (TMS; Syndel Laboratories, Nanaimo, BC, Canada) after 24 h of fasting. The sampled fish were weighed, fork length-measured, and their gills were scored following the farm’s protocol. Gills with no lesions visible and with single necrotic filament or spot were scored as G0 (i.e., pristine). Gills showing affected area up to 10% of the total gill filament were scored as G1, while G2 was defined as gills having damage affecting ~10% to 25% of the total gill filament. Gills with affected area > 25% of the total fill filament surface are scored as G3.  Fish with more than 50% of the total gill filament surface area covered with necrotic tissue were routinely observed but generally in mortalities or very moribund fish and not in healthy fish (B. Milligan, personal communication). Gills were collected for gene expression analyses. Samples around 50-100 mg of each tissue were preserved in 1 mL of RNAlater® solution (Ambion, Inc., Austin, TX, USA) and kept at 4°C overnight, then moved to -20°C until the shipping date to the Ocean Sciences Centre, Memorial University. All the collected samples were subjected for preliminary qPCR study, for sample choice and we aimed to target gills with moderate gill damage.  Using a 44K salmonid microarray platform, we aimed to compare the transcriptomes of pristine and moderately damaged gill tissue.

first: Albert

last: Caballero-Solares

first: Xi

last: Xue

first: Navaneethaiyer

last: Umasuthan

first: Barry

last: Milligan

first: Richard

middle: G

first: Rachel

last: Balder

first: Matthew

last: Rise


title: Nucleoporin107 mediates female sexual differentiation via Dsx

city: Jerusalem

first: Tikva

last: Shore

address: Ein Karem Campus

country: Israel

institute: Hebrew University of Jerusalem

summary: We recently identified a missense mutation in Nucleoporin107 (Nup107; D447N) underlying XX-ovarian-dysgenesis (XX-OD), a disorder characterized by underdeveloped and dysfunctional ovaries. Specific knockdown of Nup107 in the somatic gonadal cells and moreover, modelling of the human mutation in Drosophila  result in ovarian-dysgenesis-like phenotypes in female flies. The aberrant phenotypes in larval and adult ovaries compromised for Nup107 are associated with hyperactivation of BMP signalling. Transcriptomic analysis identified the somatic sex-determination gene Doublesex (dsx) and the extracellular metalloprotease AdamTS-A as targets of Nup107.  Either loss or gain of Dsx in the gonadal soma is sufficient to respectively mimic or rescue the phenotypes induced by Nup107 loss. Furthermore, adamTS-A is transcriptionally regulated by Dsx, and its knockdown in the somatic gonad hyperactivates BMP signaling and to a large extent recapitulates loss of Nup107 phenotypes. Thus, Dsx acts downstream of Nup107 to impact female germline stem cells via sex-specific modulation of the BMP pathway.

overall_design: Transcriptome analysis of both wild-type and mutant-rescued Nup107-null Drosophila larval gonad samples, performed in triplicate, together with a single sample of yw control gonad analysis.

first: Tgst

last: Levi

first: Ariella

last: Weinberg-Shukron

first: Yuval

last: Nevo

last: Kalifa

first: Offer

last: Gerlitz

first: David

last: Zangen


title: BET protein inhibition regulates macrophage chromatin accessibility and microbiota-dependent colitis [RNA-seq]

status: Public on Mar 20 2022

city: Chapel Hill

first: Shehzad

middle: Z

last: Sheikh

state: North Carolina

address: 111 Mason Farm Road

department: Center of Gastrointestinal Biology and Disease

institute: University of North Carolina at Chapel Hill

summary: In colitis, macrophage functionality is altered compared to normal homeostatic conditions. Loss of IL-10 signaling results in an inappropriate chronic inflammatory response to bacterial stimulation. It remains unknown if inhibition of bromodomain and extra-terminal domain (BET) proteins alters usage of DNA regulatory elements responsible for driving inflammatory gene expression. We determined if the BET inhibitor, (+)-JQ1, could suppress inflammatory activation of macrophages in Il10-/- mice. We performed ATAC-seq and RNA-seq on Il10-/- bone marrow-derived macrophages (BMDMs) cultured in the presence and absence of lipopolysaccharide (LPS) with and without treatment with (+)-JQ1 and evaluated changes in chromatin accessibility and gene expression. Treatment with (+)-JQ1 suppressed LPS-induced changes in chromatin at distal regulatory elements associated with inflammatory genes, particularly in regions that contain motifs for AP-1 and IRF transcription factors.

overall_design: Mature bone marrow derived macrophages were cultured and biologically matched samples (n=3 biological replicates/condition) were generated for the following conditions: 1) unstimulated and no treatment, 2) unstimulated and (+)-JQ1 treatment (500nM), 3) unstimulated and IL-10 treatment (10ng/mL), 4) lipopolysaccharide (LPS) stimulation (50ng/mL) and no treatment, 5) LPS stimulation and (+)-JQ1 treatment, and 6) LPS stimulation and IL-10 treatment. Samples were treated with (+)-JQ1 or IL-10 or remained untreated in culture for 12 hours followed by LPS stimulation for 4 hours in for appropriate conditions (with PBS serving as a control).

first: Michelle

last: O'Connor

first: Ana

last: Berglind

first: Meaghan

last: Kennedy

first: Benjamin

last: Keith

first: Zachary

middle: J

last: Lynch

last: Schaner

first: Erin

middle: C

last: Steinbach

first: Jeremy

last: Herzog

first: Omar

last: Trad

first: William

last: Jeck

first: Janelle

last: Arthur

first: R

middle: B

last: Sartor

first: Terrence

last: Furey


title: BET protein inhibition regulates macrophage chromatin accessibility and microbiota-dependent colitis [ATAC-seq]

middle: P

first: Matthew

middle: R

middle: K

middle: S


title: Neutrophils direct preexisting matrix in to initiate repair of damaged organs

city: München

first: Safouane

last: Kadri

address: 1, Ingolstädter Landstraße

department: Institute of Regenerative Biology and Medicine

institute: Helmholtz Zentrum München

summary: Internal organs heal injuries with new connective tissue. However the cellular and molecular events, and the sources of this tissue, remain obscure. Here we tagged extracellular matrix around the mesothelium lining various mouse tissues: peritoneum, liver, and cecum, and applied various injury models. We discovered that preexisting matrix is transferred across organs into wounds. Using proteomics, genetic lineage-tracing and by selectively injuring juxtaposed organs, we demonstrate that the matrix tissue of origin likely dictates the final healing outcome: whether scarring or regeneration. Through single-cell RNA sequencing and genetic and chemical screens, we demonstrate that preexisting matrix is transferred by neutrophils in a heat shock factor, integrin AM/B2, kindlin3, cascade and pharmacologic inhibition of this axis prevented matrix transfer and peritoneal adhesions. Matrix transfer is thus an early event of wound repair that creates a therapeutic space to dampen scaring across a range of human conditions.

overall_design: Transcriptomic examination of liver cells exposed to irreversible electroporation in adult C57BL/6 mice.

first: Tim

last: Koopmans

first: Adrian

last: Fischer

first: Safwen

first: Yuval

last: Rinkevich


title: The Nuclear Lamina Binds the EBV Genome During Latency and Regulates Viral Gene Expression

status: Public on Mar 18 2022

city: Camden

first: Jozef

last: Madzo

state: NJ

address: 403 Haddon Avenue

department: Bioinformatics

institute: Coriell Institute

summary: Several cellular factors, including the nuclear lamina, regulate chromatin composition and architecture. While the interaction of the viral genome with the nuclear lamina has been studied in the context of EBV lytic reactivation, the role of the nuclear lamina in controlling EBV latency has not been investigated. Here, we report that the nuclear lamina is an essential epigenetic regulator of the EBV episome.

overall_design: We used two EBV-positive B cell lines: LCL and Mutu I cell lines. Chromatin immunoprecipitation (ChIP) was performed using Lamin B1 or Lamin A/C antibody or sonicate chromatin (input) were used to make ChIP-seq libraries. For peak calling we compare the Lamin B1 0r Lamin A/C enrichment to the input signal

first: Lisa

last: Caruso

first: Italo

last: Tempera


title: BET protein inhibition regulates macrophage chromatin accessibility and microbiota-dependent colitis.

first: Shehzad

last: Sheikh

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

overall_design: Refer to individual Series


title: H3K9 dimethylation safeguards cancer cells against activation of the Interferon pathway [SUV39H1_ChIP-seq]

city: Copenhagen

first: Bo

last: Porse

address: Ole Maaløes Vej 5, 4

department: Finsen Laboratory and BRIC

country: Denmark

institute: University of Copenhagen

summary: Activation of the interferon genes constitutes an important anticancer pathway, able to restrict proliferation of cancer cells. Here we demonstrate that the H3K9me3 histone methyl transferase (HMT) Suppressor Of Variegation 3-9 Homolog 1 (SUV39H1) is required for the proliferation of Acute Myeloid Leukemia (AML) and find that its loss leads to activation of the interferon pathway. Mechanistically, we show that this occur via destabilization of a complex composed of SUV39H1 and the two H3K9me2 HMTs G9a and GLP. Indeed, loss of H3K9me2 correlated with the activation of key interferon pathway genes and interference with the activities of G9a/GLP largely phenocopied loss of SUV39H1. Finally, we demonstrated that inhibition of G9a/GLP synergized with DNA demethylating agents and that SUV39H1 constitutes a potential biomarker the response to hypomethylation treatment. Collectively, we uncovered a clinical relevant role for H3K9me2 in safeguarding cancer cells against activation of the interferon pathway.

overall_design: Examination of H3K9me2 and H3K9me3 histone modifications after disruption of the SUV39H1/G9a/GLP protein complex

first: Anne

last: Hansen

first: Ying

last: Ge

first: Mikkel

last: Schuster

first: Sachin

last: Pundhir

first: Adrija

last: Kalvisa


title: H3K9 dimethylation safeguards cancer cells against activation of the Interferon pathway [RNA-seq]

overall_design: RNA-seq data after disruption of the SUV39H1/G9a/GLP protein complex


title: H3K9 dimethylation safeguards cancer cells against activation of the Interferon pathway [ChIP-seq]


title: H3K9 dimethylation safeguards cancer cells against activation of the Interferon pathway


title: Chemical Pretreatment Activated a Plastic State Amenable to Direct Lineage Reprogramming

status: Public on Mar 17 2022

first: Xiaochan

last: Xu

address: Blegdamsvej 17

summary: Somatic cells can be chemically reprogrammed into a pluripotent stem cell (CiPSC) state, mediated by an extraembryonic endoderm- (XEN-) like state. We found that the chemical cocktail applied in CiPSC generation initially activated a plastic state in mouse fibroblasts before transitioning into XEN-like cells. The plastic state was characterized by broadly activated expression of development-associated transcription factors (TFs), such as Sox17, Ascl1, Tbx3, and Nkx6-1, with a more accessible chromatin state indicating an enhanced capability of cell fate conversion. Intriguingly, introducing such a plastic state remarkably improved the efficiency of chemical reprogramming from fibroblasts to functional neuron-like cells with electrophysiological activity or beating skeletal muscles. Furthermore, the generation of chemically induced neuron-like cells or skeletal muscles from mouse fibroblasts was independent of the intermediate XEN-like state or the pluripotency state. In summary, our findings revealed a plastic chemically activated multi-lineage priming (CaMP) state at the onset of chemical reprogramming. This state enhanced the cells’ potential to adapt to other cell fates. It provides a general approach to empowering chemical reprogramming methods to obtain functional cell types bypassing inducing pluripotent stem cells.

overall_design: ChIP-seq and ATAC-seq data at day 4 of the reprogramming process

first: Zhenghao

last: Yang

first: Chan

last: Gu


title: Analysis of RNA-Seq with the deletion of components of the NSL complex

city: Changchun

first: hong

middle: sen

last: liu

address: qianjin

institute: jilin university

summary: The human males absent on the first (MOF)-containing non-specific lethal (NSL) histone acetyltransferase complex consists of 9 subunits can acetylate histone H4K16, H4K5 and H4K8. NSL complex is essential for the multiple transcriptional regulation mechanisms. In this work, mRNA profiles of WT and NSL3 knockout 293T cells were generated by deep sequencing, in triplicate. qRT–PCR validation was performed using SYBR Green assays. we identified more than 100 genes as NSL complex transcriptional targets, including several transcription factors (TFs), such as Yin Yang 1 (YY1) which are mainly involved in cell proliferation, biological adhesion, and metabolic processes. the NSL complex regulates gene expression by forming a gene regulatory network with multiple TFs. It is conceivable that the NSL complex participates in the regulation of a variety of biological functions in cells.

overall_design: RNA-Seq analysis of differentially expressed genes in 293T  WT and NSL3KO cell line.

first: Yong

last: Cai

first: Jingji

last: Jin

first: Hongsen

last: Liu


title: Downregulation of m6A methyltransferase in the hippocampus of Tyrobp-/- mice and implications for learning and memory deficits

city: Jining

first: Zhanyun

last: Lv

state: Shandong

address: Guhuai Road

institute: the Affiliated Hospital of Jining Medical University

summary: Loss-of-function mutations in the gene that encodes TYRO protein kinase-binding protein (TYROBP) cause Nasu-Hakola disease, a heritable disease resembling Alzheimer's disease (AD). Methylation of N6 methyl-adenosine (m6A) in mRNA plays essential roles in learning and memory. Aberrant m6A methylation has been detected in AD patients and animal models. In the present study, Tyrobp−/− mice showed learning and memory deficits in the Morris water maze, which worsened with age. Tyrobp−/− mice also showed elevated levels of total tau, Ser202/Thr205-phosphorylated tau and amyloid β in the hippocampus and cerebrocortex, which worsened with aging. The m6A methyltransferase components METTL3, METTL14, and WTAP were downregulated in Tyrobp−/− mice, while expression of demethylases that remove the m6A modification (e.g., FTO and ALKBH5) were unaltered. Methylated RNA immunoprecipitation sequencing identified 498 m6A peaks that were upregulated in Tyrobp−/− mice, and 312 m6A peaks that were downregulated. Bioinformatic analysis suggested that most of these m6A peaks occur in sequences near stop codons and 3′-untranslated regions. These findings suggest an association between m6A RNA methylation and pathological TYROBP deficiency.

overall_design: m6A methylation in six months old  wild type and Tyrobp (-/-) mice

first: Lv

last: Zhanyun

first: Xu

last: Tongxiao

first: Li

last: Ran

first: Zheng

last: Dejie

last: Yanxin

last: Wei

first: Yang

last: Yan

first: Hao

last: Yanlei


title: Canonical Wnt Induces Focal Adhesion and Integrin Beta-1 Endocytosis

city: Parma

first: Marco

last: Morselli

state: Parma

address: Parco Area delle Scienze, 23/A

country: Italy

institute: University of Parma

summary: Activation of the Wnt pathway is at the core of many human cancers. During canonical Wnt signaling, the Lrp6 and Frizzled receptors bind to the Wnt growth factor, which leads to the complex being endocytosed. Glycogen Synthase Kinase 3 (GSK3), Dishevelled (Dvl), and Axin are sequestered inside the intraluminal vesicles of late endosomes, known as multivesicular bodies (MVBs). Here we present experiments showing that Wnt causes the endocytosis of focal adhesion (FA) proteins and depletion of Integrin β 1 (ITGβ1) from the cell surface. FAs and integrins provide link the cytoskeleton to the extracellular matrix. Wnt-induced macropinocytosis of the plasma membrane caused ITGβ1 depletion and was accompanied by striking changes in the actin cytoskeleton. In situ protease protection assays in cultured cells showed that ITGβ1 was sequestered within membrane-bounded organelles that corresponded to Wnt-induced MVBs containing GSK3 and focal adhesion-associated proteins. An in vivo model using Xenopus embryos dorsalized by Wnt8 mRNA showed that ITGβ1 depletion decreased Wnt signaling. The cross-talk between Wnt signaling, membrane trafficking, and focal adhesions should be relevant to human cancer and cell biology.

overall_design: RNA sequencing of Alexander1 ± Axin1 cells (Hepatocellular Carcinoma cell line). This HCC cell line has a deletion of the GSK3-binding region of the tumor suppressor Axin1 that results in constitutive activation of the Wnt pathway, and a stable cell line derivative that restores full-length Axin1 at physiological levels. See Albrecht et al., PNAS (2020): 115:E5317–25. DOI: 10.1073/pnas.1804091115, PMID: 29773710

first: Nydia

last: Tejeda-Muñoz

first: Yuki

last: Moriyama

first: Pooja

last: Sheladiya

first: Matteo

last: Pellegrini

first: Edward

middle: M

last: De Robertis


title: Transcriptional profiling of laser captured gonadal tissue from the planarian Schmidtea mediterranea

status: Public on Mar 15 2022

city: Madison

first: Phillip

last: Newmark

state: WI

address: 330 N Orchard St

department: Howard Hughes Medical Institute, Morgridge Institute for Research

institute: University of Wisconsin-Madison

summary: Female germ cells develop into oocytes, with the capacity for totipotency. In most animals, these remarkable cells are specified during development and cannot be regenerated. By contrast, planarians, known for their regenerative prowess, can regenerate germ cells. To uncover mechanisms required for female germ cell development and regeneration, we generated gonad-specific transcriptomes and identified genes whose expression defines progressive stages of female germ cell development. Strikingly, early female germ cells share molecular signatures with the pluripotent stem cells driving planarian regeneration. We uncovered spatial heterogeneity within somatic ovarian cells and found that a regionally enriched FoxL homolog is required for oocyte differentiation, but not specification, suggestive of functionally distinct somatic compartments. Unexpectedly, a neurotransmitter-biosynthetic enzyme, AADC, is also expressed in somatic gonadal cells, and plays opposing roles in female and male germ cell development. Thus, somatic gonadal cells deploy conserved factors to regulate germ cell development and regeneration in planarians.

overall_design: Acetone-fixed planarians were cryosectioned, then ovary, testis and non-gonadal tissue were collected using laser capture microdissection. Ovarian tissue samples were collected from 14 worms (10 worms used for cross- sections and 4 worms used for sagittal sections) for each replicate (3 replicates total). Testis and non-gonadal tissue samples were collected from 6 worms for each replicate (4 worms used for cross-sections and 2 worms used for sagittal sections). RNA was extracted using the Arcturus Picopure RNA isolation kit, followed by library generation and RNA-seq

first: Umair

middle: W

last: Khan


title: Cell type determination for cardiac differentiation occurs after seeding of human induced pluripotent stem cells

status: Public on Mar 19 2022

city: Philadelphia

first: Connie

middle: Lan

last: Jiang

state: Pennsylvania

address: 415 Curie Boulevard, Room number 415

institute: University of Pennsylvania

summary: Cardiac directed differentiation of human induced pluripotent stem cells consistently produces a mixed population of cardiomyocytes and non-cardiac cell types even when using very well-characterized protocols. We wondered whether differentiated cell types might result from intrinsic differences in hiPS cells prior to the onset of differentiation. By associating individual differentiated cells that share a common hiPS cell precursor, we were able to test whether expression variability in differentiated cells was pre-determined from the hiPS cell state. Although within a single experiment, differentiated cells that share an hiPS cell progenitor were more transcriptionally similar to each other than to other cells in the differentiated population, when the same hiPS cells were differentiated in parallel, we did not observe high transcriptional similarity across differentiations.

overall_design: Single cell RNA sequencing (10X Genomics single-cell RNA-seq kit v3) of barcoded hiPS cell-derived cardiomyocytes and hiPS cells. hiPS cells were lentivirally transduced with barcode libraries consisting of random 100-mers in the 3’ untranslated region of a green fluorescent protein (GFP) transgene after which they were permitted to divide approximately three times before being split and seeded for two parallel differentiations. On day 14 of differentiation (16 days after seeding), we used fluorescent activated cell sorting (FACS) gated for GFP positivity to enrich for barcode-containing cells on which we performed single cell RNA sequencing. As transcripts of the GFP transgene are captured by single cell RNA sequencing, we were also able to recover barcodes through a separate “side reaction” cDNA sequencing run and connect them to individual cells through analysis of GFP transcript 10x sequencing reads. Separately, we also transduced hiPS cells, allowed them to divide approximately six times before harvesting them for GFP sorting and sequencing. See Jiang et al. citation for additional details.

middle: L

first: Raj

last: Jain

first: Arjun

last: Raj


title: TGF-beta-independent function of SMAD4 preconditions naïve CD8 T-cells to prevent severe microbiota-driven chronic intestinal inflammation.

city: Bron

first: Hector

last: Hernandez-Vargas

state: Rhone

address: 36 bis Rue de la Batterie

institute: Genomics Consulting


title: Analysis of histone modification with the deletion of components of the NSL complex

first: hong

last: liu

summary: The human males absent on the first (MOF)-containing non-specific lethal (NSL) histone acetyltransferase complex consists of 9 subunits can acetylate histone H4K16, H4K5 and H4K8. It has been known that this complex shares WDR5 subunit with the MLL/SET histone methyltransferases, suggesting that NSL complex is essential for the multiple transcriptional regulation mechanisms. However, there are few reports on the function of NSL HAT in genome, It is necessary to clarify the details and specificity of histone H4 acetylation cooperates with H3K4 methylation to regulate gene transcription. In this work, Our results show that The NSL complex plays a critical role in regulating multiple histone modifications by using the CRISPR/Cas9 NSL3-KO 293T cell line and ChIP-Seq approach. The global levels of highly enriched H4K16ac, H4K5ac, H3K4me2 and H3K4me3 at TSSs (TSS ± 3 kb region) are inhibited by NSL3-KO. NSL3-KO seemed to have little effect on the enhancer mark H3K27ac and repressor mark H3K27me3, but H3K4me1 was greatly affected by NSL3-KO. Moreover, the NSL complex govern gene transcription and identified genes by a coordinative mode of H4K16ac and H3K4me2/me3. What’s more, De novo motif analysis of MOF and NSL3 targets indicated that the NSL complex forms a gene regulatory network with multiple transcription factors by regulating their genes expression or cooperating with some factors through binding to specific motif.

overall_design: ChIP analysis of different histone modifications, pol II and the NSL complex subunits in 293T WT and NSL3KO cell line.

first: Yong

last: Cai

first: Jingji

last: Jin

first: Hongsen

last: Liu


title: HiCuT: an efficient and low input method to identify protein-centric chromatin interactions

status: Public on Mar 16 2022

first: Satish

last: Sati

address: 421 Curie Blvd, BRB 1039

department: Dermatology

summary: Protein-DNA interactions regulate gene expression, and some interactions occur over large distances, such that they are nearby in 3-D space but are separated by many nucleotides in the linear genome. These long-range chromatin loops are essential for gene regulation but remain difficult to interrogate. Methods to capture these chromatin interactions mediated by a specific protein factor include Hi-C sequencing coupled with chromatin immunoprecipitation-sequencing (ChIP-seq), chromatin interaction analysis by paired-end tag sequencing (ChIA-PET), proximity ligation-assisted ChIP-seq (PLAC-seq), and HiChIP. These methods all require high amounts of starting material (0.5M – 100M cells) , which limits their general use. Here, we describe Hi-C Coupled chromatin cleavage and Tagmentation (HiCuT), an enzyme-based tagmentation strategy that provides efficient and high-resolution protein-centric chromatin mapping from as few as 5,000 cells. Thus, HiCuT will permit protein-centric 3-D genome binding assessment in rare cell populations that were previously not testable, including primary cells, human tissue samples, and personalized epigenomics.

overall_design: HiCuT assay on GM12878 and primary Keratinocytes

first: Parker

last: Jones

first: Hali

last: Kim

first: Linda

middle: A

last: Zhou

first: Thomas

last: Leung


title: Large-scale H3.1/H3.2 - H3K9me3 deposition defines the epigenetic signature of extraembryonic lineage cells [HiC]

city: Tsukuba

first: Masashi

last: Hada

state: Ibaraki

address: Koyadai 3-1-1

department: Bioresource Research Center

institute: RIKEN

summary: The extraembryonic lineage in mammals is unique in its unipotent differentiation ability and gestation-limited development. Here we identified the epigenomic characters of mouse trophoblast stem cells (TSCs) focusing on the genome-wide enrichment of histone H3.1/H3.2 and H3K9me3. Comparative ChIP-sequencing (ChIP-seq) analysis of TSCs and embryonic stem cells (ESCs) revealed that the TSC genome uniquely contained large (over 1Mb) H3.1/H3.2-H3K9me3 domains in the intergenic regions. This feature was common to extraembryonic cells in mice and humans. Depletion of CAF1, a H3.1/H3.2 chaperone, led to downregulation of TSC marker genes such as Cdx2 and Elf5 and ectopic expression of Oct3/4, an ESC marker. Nuclear transfer cloning using TSCs resulted in extremely poor embryonic development, but removal of H3K9me3 from their genome resulted in birth of the first TSC-cloned mice. Thus, the signature of the extraembryonic epigenome is safeguarded by the H3.1/H3.2-H3K9me3 enrichment, which protects the genome from cell fate transition.

overall_design: ChIP-seq analysis of H3K9me3, H3.1/H3.2, and H3.3 in ESCs, TSCs, control TSCs, and P150 knockdown TSCs. ATAC-seq analysis of ESCs and TSCs.  Hi-C analysis and Repli-seq analysis of TSCs. RNA-seq analysis of control TSCs and P150 knockdown TSCs.

first: Hisashi

last: Miura

first: Akie

last: Tanigawa

first: Shogo

last: Matoba

first: Kimiko

last: Inoue

first: Narumi

last: Ogonuki

first: Michiko

last: Hirose

first: Naomi

last: Watanabe

first: Ryuichiro

last: Nakato

first: Katsunori

last: Fujiki

first: Ayumi

last: Hasegawa

first: Akihiko

last: Sakashita

first: Hiroaki

last: Okae

first: Kento

first: Daiki

last: Shikata

first: Takahiro

last: Arima

first: Katsuhiko

last: Shirahige

first: Ichiro

last: Hiratani

first: Atsuo

last: Ogura


title: Transcriptional regulation of insulin action and sensitivity via a GSK3β-FBXW7-ERRα axis

city: Montréal

first: Vincent

last: Giguère

state: QC

address: 1160 Pine Ave West, McIntyre Bldg Room 710

department: Goodman Cancer Centre

institute: McGill University

summary: Insulin resistance, a harbinger of the metabolic syndrome, is a state of compromised hormonal response for which transcriptional dysregulation is a major contributor. Genetic or pharmacological inhibition of the nuclear receptor ERRα preserves insulin sensitivity during diet-induced obesity. However, how ERRα integrates insulin signaling at the molecular level with whole body metabolism remains unknown. Herein, we reveal that insulin-mediated gene expression requires the nuclear stabilization of ERRα through a GSK3β/FBXW7 axis. Liver-specific deletion of GSK3β or FBXW7 or mice harboring mutations of ERRα phosphosites (ERRα3SA) co-targeted by GSK3β/FBXW7 result in accumulated ERRα proteins that no longer respond to insulin. ERRα3SA mice display reprogrammed liver and muscle transcriptomes, resulting in insulin resistance and compromised metabolic homeostasis, effects largely reversed by pharmacological inhibition of ERRα. These findings uncovered a previously unrecognised  ERRα-dependent insulin regulatory pathway that could be targeted to improve insulin resistance and associated metabolic diseases.

overall_design: Liver RNA-sequencing data of WT and ERRα KO mice treated with saline or insulin as well as liver/muscle RNA-sequencing data of WT and ERRα3SA mice.


title: RNA sequencing of secondary heart field- and cardiac neural crest-derived cells from murine aortas

status: Public on Mar 14 2022

city: Baltimore

first: Tyler

last: Creamer

state: Maryland

address: 733 N Broadway

department: Genetic Medicine

institute: Johns Hopkins University

summary: To better understand the role of lineage in smooth muscle cell heterogenity along the length of the aorta, we performed bulk RNA-sequencing transcriptomic analysis on secondary heart field (SHF)-derived and cardiac neural crest derived (CNC)-derived sorted cells from murine aortas.

overall_design: Comparison of the transcriptome of SHF-derived (n=3) and CNC-derived (n=3) cells in murine aortas.

first: Rustam

last: Bagirzadeh

first: Emily

middle: E

last: Bramel

first: Elena

last: Gallo MacFarlane


title: Nuclear and cytoplasmic whole transcriptome RNA-sequencing of mouse endothelial cell line and mmu-miR-466c knock-down cell line

city: Kuopio

first: Mikko

last: Turunen

address: Yliopistonranta 1 E

department: A. I. Virtanen Institute

country: Finland

institute: University of Eastern Finland

summary: To analyze the role of miR-466c in the regulation in hypoxic response, we used CRISPR-mediated gene editing to remove 97 bp region from the Sfmbt2 intron 10 which contains the miR-466c hairpin. We performed whole transcriptome RNA-sequencing of the parental mouse endothelial cell line C166 and the mmu-miR-466c knock-down cell line.

overall_design: Two cell lines (C166 wildtype and C1.1 mutant cell lines) were cultured in normoxia or hypoxia (2h and 24h timepoints) and split to nuclear and cytoplasmic fractions. All samples were done in triplicate.

first: Pia

last: Laitinen

first: Thomas

last: Roberts

first: Tiia


title: H3K9 dimethylation safeguards cancer cells against activation of the Interferon pathway [WGBS]

first: Bo

last: Porse

summary: Activation of interferon genes constitutes an important anticancer pathway able to restrict proliferation of cancer cells. Here we demonstrate that the H3K9me3 histone methyl transferase (HMT) Suppressor of Variegation 3-9 Homolog 1 (SUV39H1) is required for the proliferation of Acute Myeloid Leukemia (AML) and find that its loss leads to activation of the interferon pathway. Mechanistically, we show that this occurs via destabilization of a complex composed of SUV39H1 and the two H3K9me2 HMTs, G9A and GLP. Indeed, loss of H3K9me2 correlated with the activation of key interferon pathway genes, and interference with the activities of G9A/GLP largely phenocopied loss of SUV39H1. Finally, we demonstrate that inhibition of G9A/GLP synergized with DNA demethylating agents and that SUV39H1 constitutes a potential biomarker for the response to hypomethylation treatment. Collectively, we uncovered a clinically relevant role for H3K9me2 in safeguarding cancer cells against activation of the interferon pathway.

overall_design: Examination of the DNA methylation on retrotransposons in SUV39H1 depleted cells

first: Anne

last: Hansen

first: Ying

last: Ge

first: Mikkel

last: Schuster

first: Sachin

last: Pundhir

first: Janus

last: Jakobsen

first: Adrija

last: Kalvisa

first: Sandra

last: Gordon

first: Francesca

last: Ambri

first: Frederik

middle: O

last: Bagger

first: Deo

last: Pandey

first: Kristian

last: Helin

middle: T


title: RNAseq analysis of human macrophages differentiated from monocytes exposed to extracellular vesicles from tumor cells or recombinant CSF-1

city: Paris

first: Elodie

last: Segura

address: 156 rue de Vaugirard

department: INSERM U1151

institute: INSERM

summary: Tumor  associated  macrophages  (TAMs),  which  differentiate  from  circulating  monocytes,  are pervasive across human cancers and comprise heterogeneous populations. The contribution of tumor-derived signals to TAM heterogeneity is not well understood. In particular, tumors release both soluble factors and extracellular vesicles (EVs), whose respective impact on TAM precursors may be different. Here, we show that triple negative breast cancer cells (TNBC) release EVs and soluble  molecules  promoting  monocyte  differentiation  towards  distinct  macrophage  fates.

overall_design: Human CD14+ monocytes were cultured with extracellular vesicles from MDA-MB-231 tumor cells (0.02  ng/ml) or recombinant CSF-1 (100  ng/ml) for 5 days.

first: Mercedes

last: Tkach

first: Clotilde

last: Théry


title: Global analysis of expression upon RNA Pol I inhibition (iPol I) in ESCs

city: London

first: Michelle

last: Percharde

address: Du Cane Road

department: Imperial College London

country: United Kingdom

institute: MRC Laboratory of Medical Sciences

summary: Immediately following fertilisation, the genomes of mammalian embryos undergo extensive reprogramming to generate a totipotent zygote with the capacity to form the entire developing organism and associated extraembryonic tissues. The embryo must also switch from dependence on maternal transcripts to transcribing from its own genome, and is accompanied in mice by rapid and transient upregulation of MERVL transposons and MERVL-driven genes at the 2-cell stage. The mechanisms and requirement for MERVL and 2-cell (2C) gene upregulation are poorly understood. Moreover, this program must be rapidly shut off upon 2C exit and developmental progression. Here, we report that robust ribosomal RNA synthesis and nucleolar integrity are essential for exit from the 2C state. 2-cell embryos and 2C-like cells show immature nucleoli with altered structure and reduced function, and we reveal that nucleolar disruption induces conversion to the 2C-like state in ESCs. Mechanistically, the 2C/MERVL activator Dux is held in perinucleolar chromatin in ESCs and released into the nucleoplasm upon nucleolar disruption or direct inhibition of nucleolar phase separation, leading to its rapid upregulation. In embryos, nucleolar disruption prevents proper Dux silencing and leads to 2-4 cell arrest. Our findings reveal an intriguing link between rRNA synthesis, nucleolar maturation and gene repression during early development.

overall_design: 3 replicates each of mouse ESCs at 0/2/4/8h after treatment with 1uM BMH-21 or CX-5461 used for RNA extraction and library generation (21 samples total)

first: Sheila

last: Xie


title: DUX4 is a multifunctional factor priming human embryonic genome activation

city: Yokohama

first: Shruti

last: Bhagat

state: Kanagawa

address: W424, RIKEN Yokohama Campus, 1-7-22 Suehiro-cho, Tsurumi-ku

department: RIKEN Center for Integrative Medical Sciences

institute: RIKEN, Yokohama

summary: Double homeobox 4 (DUX4) is expressed in at the early preimplantation stage in human embryos. Here we show that induced human DUX4 expression substantially alters the non-coding chromatin accessibility of non-coding DNA and activates thousands of newly identified putative transcribed enhancer-like regions, preferentially located within ERVL-MaLR repeat elements. CRISPR activation of transcribed enhancers by C-terminal DUX4 motifs results in the increased expression of target embryonic genome activation (EGA) genes ZSCAN4 and KHDC1P1. We show that DUX4 is markedly enriched in human zygotes, followed by intense nuclear DUX4 localization preceding and coinciding with minor EGA. DUX4 knockdown in human zygotes led to changes in the EGA transcriptome but did not terminate the embryos. We also show that the DUX4 protein interacts with the Mediator complex via the C-terminal KIX binding motif. Our findings contribute to the understanding of DUX4 as a regulator of the non-coding genome.

overall_design: This study contains 40 samples (ATAC-seq: 8 samples; CAGE: 4 samples; NET-CAGE: 4 samples; bulk STRT: 24 samples) comprising of controls (dox minus) and Doxycycline treated (dox plus). All experiments were performed in DUX4 TetOn H1 and H9 human embryonic stem cell (hESC) lines. The DUX4 TetOn hESCs have been treated with 1μg/ml of Doxycycline for 4 hours. For ATAC-seq, 4 samples were controls and 4 samples were Doxycycline teated. For CAGE, 2 samples were controls and 2 samples were Doxycycline teated, and same was done also for the NET-CAGE samples. For bulk STRT, 12 samples were controls and 12 samples were Doxycycline teated.

first: Masahito

last: Yoshihara

first: Anastassius

last: Damdimopoulos

first: Shintaro

last: Katayama

first: Sanna

last: Vuoristo


title: The NFκB pathway promotes endocrine therapy tolerance through activation of integrated stress response in ER positive breast cancer

city: Chicago

first: Leonid

last: Feferman

state: Illinois

address: 1737 West Polk St

department: Research Resources Center

institute: University of Illinois at Chicago

summary: While estrogen receptor positive breast tumors generally respond well to endocrine therapy (ET), up to 40% of patients will experience relapse, either while on endocrine therapy or after ET is completed. We have previously shown that the selective pressure of tamoxifen activates the pro-survival NFkB pathway in patient tumors, breast cancer cell lines, and breast cancer xenograft tumors, and that this activation allows for survival of a population of ET-tolerant cells, which can contribute to relapse after ET withdrawal. Here, we utilized single cell RNA-sequencing to characterize survival pathways in NFkB+, ET-tolerant cells and identified activation of the integrated stress response as a critical survival mechanism. Moreover, we found key players in this pathway are regulated by ET in patient derived organoids. In addition, we developed an ET-tolerant gene signature that can be found in metastatic cell populations and that predicts poor outcome to ET. Our findings suggest that co-targeting of ER and key players in the integrated stress response may be a viable therapeutic strategy to eliminate cells that survive the selective pressure of ET.

overall_design: scRNA-seq (OneCellBio)

first: Svetlana

last: Semina

first: Jonna

last: Frasor


title: Analysis of small ncRNAs in stool samples from ASD patients

status: Public on Mar 12 2022

city: Segrate

first: Ettore

last: Mosca

address: Via F.lli Cervi 93

institute: National Research Council - Institute of Biomedical Technologies

summary: Intestinal microorganisms impact on health maintaining gut homeostasis and shaping the host immunity, while gut dysbiosis associates with many conditions including autism, a complex neurodevelopmental disorder with multifactorial aetiology. In autism, gut dysbiosis correlates with symptom severity and is characterized by a reduced bacterial variability and a diminished beneficial commensal relationship. Microbiota can influence the expression of host microRNAs that, in turn, regulate the growth of intestinal bacteria by means of bidirectional host-gut micro-biota cross-talk. We investigated possible interactions among intestinal microbes and between them and host transcriptional modulators in autism. To this purpose, we analysed, by “omics” technologies, faecal microbiome, mycobiome and small non-coding-RNAs (particularly miRNAs and piRNAs) of children with autism and neurotypical development. Patients displayed gut dysbiosis, related to a reduction of healthy gut micro- and mycobiota, and up-regulated tran-scriptional modulators. The targets of dysregulated non-coding-RNAs are involved in intestinal permeability, inflammation and autism. Furthermore, microbial families, underrepresented in patients, participate to the production of human essential metabolites negatively influencing the health condition. Here, we propose a novel approach to analyse faeces as a whole and, for the first time, we detected miRNAs and piRNAs in faecal samples of patients with autism.

overall_design: 6 stool samples from individuals with Autism Spectrum Disorders and 6 age matched stool samples from neurotypical individuals. "ASD" samples are the case group, and "Ctrl" samples are the controls group.

first: Federica

last: Chiappori

first: Francesca

last: Cupaioli

first: Arianna

last: Consiglio

first: Noemi

last: Di Nanni

first: Vito

middle: F

last: Licciulli

first: Alessandra

last: Mezzelani


title: TGF-beta-independent function of SMAD4 preconditions naïve CD8 T-cells to prevent severe microbiota-driven chronic intestinal inflammation [RNAseq]

first: Hector

last: Hernandez-Vargas

summary: SMAD4, a key mediator of TGF-beta signaling, plays a crucial role in T cells to prevent chronic intestinal inflammation through unknown mechanisms. We reveal that SMAD4 in CD8 T cells prevents chronic intestinal inflammation primarily in a TGF-beta-independent manner. Mechanistically, SMAD4, in CD8 T cells, acts as a basal and tonic repressor of TGF-beta-target genes at the transcriptional and epigenetic level, prior to any TGF-beta signal.  SMAD4 deletion affects aberrantly a wide range of TGF-beta-target genes, thereby promoting accumulation and epithelial retention of CD8.alpha.beta T cells inversely to total TGF-beta signaling disruption. Moreover, SMAD4 deletion unleashes the expression of TGF-beta-signaling-repressors and hampers TGF-β-mediated CD8 T cell immunosuppression, eliciting their chronic activation. Hence, in a feedforward mechanism, SMAD4 both blocks the TGF-beta signature in CD8 T cells and pre-sensitizes them to TGF-beta.

overall_design: We used the CD4-CRE conditional deletion system to establish several mouse strains lacking one or several TGF-β signaling pathways. We established mice with a deletion of SMAD4 (SKO), TGF-beta RII (R2KO), and double deletion of TGF-beta RII and SMAD4 (R2SKO) in T cells. We next performed SMAD4 RNAseq of CD8 T cells from WT, SKO, R2KO, and R2SKO mice. In order to rule out any potential side effects of the inflammatory environment, we used F5 transgenic CD8 T cells in a RAG2KO background, because these mice do not develop any inflammation.

first: Ramdane

last: Igualouzene

first: Saidi

last: M’Homa Soudja


title: TGF-beta-independent function of SMAD4 preconditions naïve CD8 T-cells to prevent severe microbiota-driven chronic intestinal inflammation [ChIPseq]

overall_design: We used the CD4-CRE conditional deletion system to establish several mouse strains lacking one or several TGF-β signaling pathways. We established mice with a deletion of SMAD4 (SKO), TGF-beta RII (R2KO), and double deletion of TGF-beta RII and SMAD4 (R2SKO) in T cells. We next performed SMAD4 ChIPseq of CD8 T cells from WT, SKO and R2KO mice. In order to rule out any potential side effects of the inflammatory environment, we used F5 transgenic CD8 T cells in a RAG2KO background, because these mice do not develop any inflammation.


title: HSF2 cooperates with HSF1 to drive a transcriptional program critical for the malignant state

first: Marc

last: Mendillo

state: IL

address: 303 E Superior St

department: Biochemistry and Molecular Genetics

institute: Feinberg School of Medicine, Northwestern University

summary: Heat shock factor 1 (HSF1) is well known for its role in the heat shock response (HSR), where it drives a transcriptional program comprising heat shock protein (HSP) genes, and in tumorigenesis, where it drives a program comprising HSPs and many non-canonical target genes that support malignancy. Here, we find that HSF2, an HSF1 paralog with no significant role in the HSR, physically and functionally interacts with HSF1 across diverse types of cancer. HSF1 and HSF2 have strikingly similar chromatin occupancy and regulate a common set of genes which include both HSPs and non-canonical transcriptional targets with roles critical in supporting malignancy. Loss of either HSF1 or HSF2 results in a dysregulated response to nutrient stresses in vitro and reduced tumor progression in cancer cell line xenografts. Taken together, these findings establish HSF2 as a critical cofactor of HSF1 in driving a cancer cell transcriptional program to support the anabolic malignant state.

overall_design: Chromatin occupancy of HSF1 and HSF2 was measured in two cancer cell lines by ChIP-seq. Gene expression profiles were obtained from a panel of cancer cell lines with HSF1 or HSF2 depletion by siRNA or CRISPR/Cas9 gene editing. Gene expression profiles were also obtained from tumors resulting from implantation of HSF1 or HSF2 knockout cancer cell lines in immunocompromised mice.

first: Roger

last: Smith

first: Seesha

last: Takagishi

first: David

last: Amici

first: Kyle

last: Metz

first: Sitaram

last: Gayatri

first: Milad

last: Alasady

first: Yaqi

last: Wu

first: Sonia

last: Brockway

first: Stephanie

last: Taiberg

first: Natalia

last: Khalatyan

first: Mikko

last: Taipale

first: Sandro

last: Santagata

first: Luke

last: Whitesell

first: Susan

last: Lindquist

first: Jeffrey

middle: N

last: Savas


title: SAGA Complex Subunits in Candida albicans Differentially Regulate Filamentation, Invasiveness and Biofilm Formation

status: Public on Mar 09 2022

city: Montreal

first: Saima

last: Rashid

state: Quebec

address: 7141 Sherbrooke St W

department: Department of Arts and Science

institute: Concordia University

summary: In this study we were interested in studying the role of SAGA subunits in various cellular processes- morphogenetic changes, growth, invasiveness, biofilm formation and to check the role of these subunits under various cellular and genotoxic conditions. In this work, we investigated conditional and null mutants of components of the SAGA complex modules; Ngg1 of the HAT module, Ubp8 of the Dub module, Tra1 of the recruitment module, Spt7 of the architecture module, and Spt8 of the TBP interaction unit to assess their role in processes such as filamentation, invasiveness, and biofilm formation. Spt7 and Spt8 deletion in Candida albicans resulted in ifilamentatation, invasiveness  in YPD media at 30 degrees Celsius and  also showed sensitivity to antifungal drugs. Wild type SN148 do not make any filaments  in YPD at 30 degrees Celsius and didn't show antifungal sensitivity  . The aim was to look for transcription profiling of SAGA mutants against wild type to find genes up and down regulated in the mutant especially those ones critical for filamentation, invasiveness and antifungal drug resistance.

overall_design: Liquid cultures of wild-type (WT)  and SAGA subunits were adjusted to OD600 at 0.8 in 10 mL liquid YPD medium and incubated at room temperature for 8 hours with shaking at 220 rpm.

first: Malcolm

last: Whiteway


title: miRNA profiling in heart tissue of AGAT-/- mice

city: Hamburg

first: Märit

last: Jensen

address: Martinistraße 52

institute: University Medical Center Hamburg-Eppendorf (UKE)

summary: miRNA sequencing was performed in left ventricular heart tissue of wt (n = 5), AGAT-/- (n = 5), and AGAT-/- mice supplemented with homoarginine (n = 5). Small RNA libraries were prepared using TruSeq Small RNA Sample Preparation Kits (Illumina, San Diego, CA, USA) according to the manufacturer’s instructions. The CLC Genomics Workbench (clcbio.com/products/clc-genomics-workbench/) was used to map reads from miRNA sequencing against the murine set of all known miRNAs, which was retrieved from miRBase (www.mirbase.org/). The number of reads falling in mature miRNAs were extracted and further processed in R. Only miRNAs covered by more than ten reads were kept for further analyses. Differential expression of miRNAs between groups of mice was calculated by R/Bioconductor package DESeq2 and the False Discovery Rate (FDR) based Benjamini-Hochberg method was used to account for multiple tests. Differentially expressed miRNAs with an FDR ≤ 0.05 were considered significant.

overall_design: Heart tissue miRNA profiles in wt, AGAT-/- and AGAT-/- mice supplemented with homoarginine were generated by deep sequencing using Illumina HiSeq 2500.

first: Christian

last: Müller

first: Tanja

last: Zeller


title: Large-scale H3.1/H3.2 - H3K9me3 deposition defines the epigenetic signature of extraembryonic lineage cells [Repli-seq]

first: Masashi

last: Hada

first: Hisashi

last: Miura

first: Akie

last: Tanigawa

first: Shogo

last: Matoba

first: Kimiko

last: Inoue

first: Narumi

last: Ogonuki

first: Michiko

last: Hirose

first: Naomi

last: Watanabe

first: Ryuichiro

last: Nakato

first: Katsunori

last: Fujiki

first: Ayumi

last: Hasegawa

first: Akihiko

last: Sakashita

first: Hiroaki

last: Okae

first: Kento

first: Daiki

last: Shikata

first: Takahiro

last: Arima

first: Katsuhiko

last: Shirahige

first: Ichiro

last: Hiratani

first: Atsuo

last: Ogura


title: A case of genetic hypoglycemia in an adult: A Molecular Approach

city: Fitzroy

first: Balasubramanian

last: Krishnamurthy

state: VIC

address: 41 Victoria Parade

department: Department of Endocrinology and Diabetes

institute: St. Vincent’s Hospital Melbourne

summary:
Context: We present a case of a young adult with GCK mutation who had neonatal hypoglycaemia, re-emerging with hypoglycaemia later in life. Mechanistic insight is gained from detailed clinical, cellular and genomic analysis.
Conclusion: This case highlights the variable phenotype of GCK mutations. In depth molecular analyses in the islets has revealed possible mechanisms for nesidioblastosis and insulin hypersecretion.

overall_design: We investigated a 22-year-old man with asymptomatic hypoglycaemia with a family history of maternal hypoglycaemia. Genetic testing yielded a novel GCK mutation that was subsequently found in his mother, sister and nephew. After partial pancreatectomy, immunohistochemistry revealed diffuse nesidioblastosis. Functional analysis of his islets by assessing intracellular calcium, a surrogate marker for insulin secretion, revealed normal basal but increased glucose stimulated insulin secretion. Unbiased gene expression analysis using 10X single cell sequencing revealed upregulated INS gene, upregulated CKB gene and downregulated DLK1 and NPY genes in beta-cells, which may explain the metabolic and histological phenotype associated with glucokinase mutation. There were more larger islets in the patient’s pancreas than in control subjects but there was no difference in the proportion of beta cells in the islets. His hypoglycaemia was persistent after pancreatectomy and was refractory to diazoxide therapy. Octreotide partially improved hypoglycaemia.


title: Single-cell transcriptomic analysis of Smad3-deficent mice

first: Tyler

middle: J

last: Creamer

summary: To analyze both early and late events associated with Smad3-inactivation, and to identify how specific smooth muscle cell (SMC) phenotypes may confer increased susceptibility to disease in the aortic root, we performed single-cell transcriptomic analysis on aortic cells from control (Smad3+/+; Myh11-CreER; + or – EGFP-L10a mice are refered to as Smad3+/+) and Smad3-deficient (Smad3lox/lox; Myh11-CreER; + or - EGFP-L10a  are referred to as Smad3SmKO) mice at 10- and 18-weeks post-deletion, when aortic dilation is undetectable and moderate, respectively. Analysis of SMC-specific transcriptional changes show that, in addition to the expected decrease in TGF-β -induced expression of extracellular matrix components, Smad3-inactivation resulted in a broad reduction in the expression of components and modulators of focal adhesions, including that of several integrins, anchoring proteins, adaptors and modulators.

overall_design: Examination of the aortic transcriptome of mice with postnatal, smooth-muscle specific deletion of Smad3 at two different timeponts, 10- and 18-weeks post-deletion when dilation is undetectable and moderate, respectively.

first: Emily

last: Bramel

first: Elena

middle: G

last: MacFarlane


title: Exon Level Expression Profiling:HTA 2.0 array

city: Houston

first: Ruojia

last: Wu

address: 6500 Main St

institute: Rice University

summary:
Transcriptome analysis of  RNA samples extracted from human placenta FFPE sample
RNA expression level comparison

overall_design: We analyzed RNA from commercial human placenta FFPE tissue using the Affymetrix HTA2.0 array. Array data was processed by Affymetrix Exon Array Computational Tool. Techinical Duplicates were performed.

first: David

last: Zhang

first: Jinny

first: Peng

last: Dai


title: Derivation of stem cell-like cells from spherical culture of astrocytes for enhanced neural repair after middle cerebral artery occlusion

city: changsha

first: dan

last: zhu

address: hunan

institute: Hunan University

summary: Neural precursor cells (NPCs) tend to aggregate and develop into three-dimensional (3D) spheres, which in return help maintain the stemness of the cells. This close relationship between spherical environment and cell stemness direct us to assume that 3D spheres of astrocytes may facilitate the acquiring of stem cell-like features and generate sufficient seed cells for regeneration of neurons. Primary astrocytes (ASTs) were isolated from the cerebra of postnatal day 1 to day 2 C57BL/6 mouse. ASTs were loaded onto the agarose-coated dishes and cultured to induce the formation of AST spheres. The status of the ASTs over time in cell spheres was characterized by immunochemistry, flow-cytometry, western blotting, and RNA-sequencing. The potency of AST-derived stem cells (A-iSCs) in repairing neural injuries was evaluated in mouse models of middle cerebral artery occlusion (MCAO), spinal cord injury (SCI), and peripheral nerve injury (PNI). In vitro results confirmed that mouse astrocytes cultured on agarose surfaces spontaneously formed cell spheres and exhibited molecular features similar to stem cells, capable of further differentiating into neurons particularly and forming functional synaptic networks with synchronous burst activities. RNA-sequencing results revealed the similarity of A-iSCs to NPCs in global gene expression profiles. It was observed that the transplanted A-iSCs expressed a series of markers for neural differentiation, such as NeuN, Tuj1, and Map2, indicating the conversion of the transplanted A-iSCs into neurons in these scenarios. We also found that the injured mice injected with A-iSCs exhibited significant improvements in sensorimotor functions over 8 weeks compared with the sham and control mice. Taken together, mouse astrocytes form cell spheres on agarose surfaces and acquire stem cell-associated features, and the derived A-iSCs possess the capacity to differentiate into neurons and facilitate the regeneration of damaged nerves across the nervous system.

overall_design: To elucidate the mechanism involved in the induction of ASTs in 3D cell spheres, global gene expression profile of ASTs (0 day incubation, negative control) and A-iSCs (5 days incubation) were characterized by RNA-sequencing, with the profiles of brain-derived P56-NPCs and P0-NPCs as positive controls. Three replicates per sample.

first: Dan

last: Zhu

first: Zheming

last: Cao

first: Xiaoyang

last: Pang

first: Wei

last: Jiang


title: Predictive biomarkers for survival benefit with ramucirumab in urothelial cancer in the RANGE trial

city: New York

first: Erik

middle: Roy

last: Rasmussen

state: NY

address: 430 East 29th Street

institute: Eli Lilly

summary:
The RANGE study (NCT02426125) evaluated ramucirumab (an anti-VEGFR2 monoclonal antibody) in patients with platinum-refractory advanced urothelial carcinoma (UC). Here, we use programmed cell death-ligand 1 (PD-L1) immunohistochemistry (IHC) and transcriptome analysis to evaluate the association of immune and angiogenesis pathways, and molecular subtypes, with overall survival (OS) in UC. Higher PD-L1 IHC and immune pathway scores, but not angiogenesis scores, are associated with greater ramucirumab OS benefit. Additionally, Basal subtypes, which have higher PD-L1 IHC and immune/angiogenesis pathway scores, show greater ramucirumab OS benefit compared to Luminal subtypes, which have relatively lower scores. Multivariable analysis suggest patients from East Asia as having lower immune/angiogenesis signature scores, which correlates with decreased ramucirumab OS benefit. Our data highlight the utility of multiple biomarkers including PD-L1, molecular subtype, and immune phenotype in identifying patients with UC who might derive the greatest benefit from treatment with ramucirumab.
Please note that the RANGE clinical trial data generated in this study have been deposited at www.vivli.org. RANGE clinical trial data are available under restricted access to protect patient privacy, and access can be obtained by submitting a request. For details on submitting a request, see the instructions provided at www.vivli.org. For specific study details, see here: https://search.vivli.org/?search=NCT02426125.

overall_design:
RNA was isolated from tumor sample specimens from 394 patients with MIBC in the RANGE Phase 3 cliinical trial and whole transcriptome profiling was performed. Samples were classified according to four published molecular subtyping methods.
Please note that CEL files contain the 'Decipher' string for chp information. The array data (i.e. CEL files) were generated for Eli Lilly by Decipher Biosciences (as part of a service contract to perform gene expression profiling and apply their proprietary bladder cancer molecular subtype classifier to these n=394 patient samples) and the 'HuEx-1_0-st' array was positioned as their proprietary 'Decipher' array.

first: M

last: van der Heijden

first: T

last: Powles

first: D

last: Petrylak

first: R

last: de Wit

first: A

last: Necchi

first: C

last: Sternberg

first: N

last: Matsubara

first: H

last: Nishiyama

last: Castellano

first: S

last: Hussain

last: Bamias

first: G

last: Gakis

first: J-L

last: Lee

last: Tagawa

first: U

last: Vaishampayan

first: J

middle: B

last: Aragon-Ching

first: B

last: Eigl

last: Hozak

first: E

last: Xia

last: Rhodes

last: Wijayawardana

first: K

last: Bell-McGuinn

last: Aggarwal

last: Drakaki


title: Next Generation Sequencing Facilitates Quantitative Analysis of osteogenic differentiation treated with metformin

status: Public on Mar 11 2022

city: shenyang

first: Keda

last: Yang

state: --- Select One ---

address: North Nanjing Street 155

institute: The First Affiliated Hospital of China Medical University

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to determine the differentially expressed genes of osteogenic differentiation after treatment with metformin by using NGS-derived transcriptome profiling (RNA-seq)
Methods: Osteogenic mRNA profiles of MC3T3-E1 cells treated in osteogenic medium and in osteogenic medium with metformin, in triplicate, using Illumina instrument.
Results: Using an optimized data analysis workflow, the data reveals 1946 up-regulated and 1544 down-regulated genes after metformin treatment.
Conclusions: Our study represents the first detailed analysis of osteogenic transcriptomes after metformin treatment, with biologic replicates, generated by RNA-seq technology. We conclude that RNA-seq based transcriptome characterization would contribute the target prediction and drug discovery.

overall_design: Osteogenic mRNA profiles of MC3T3-E1 cells in osteogenic medium and in osteogenic medium with metformin.

first: Yue

last: Zhu

first: Lin

last: Tao

first: Fangming

last: Cao


title: Tenogenic induction from induced pluripotent stem cells unveils the trajectory towards tenocyte differentiation.

city: Fukuoka

first: Yasuyuki

last: Ohkawa

address: 3-1-1 Maidashi

institute: Medical Institute of Bioregulation

summary: The musculoskeletal system is integrated by tendons that are characterized by expression of a functionally important transcription factor Scleraxis (Scx). To date, there has been no solid culture system for tenogenic differentiation. Here we developed the tenocyte induction method using induced pluripotent stem cells established from ScxGFP transgenic mice by monitoring fluorescence that reflects a series of dynamic differentiation process. Among several developmentally relevant factors, TGF-2 was the most potent inducer for differentiation of tenocytes expressing Tenomodulin, a mature differentiation marker. Single-cell RNA sequencing (scRNA-seq) revealed 11 distinct clusters including mature tenocyte population and tenogenic differentiation trajectory, which well recapitulates in vivo developmental process. Dataset of scRNA-seq and transcriptome of Scx deficient tendon enabled identification of Scx-related genes in which tendon-related genes are enriched. Collectively, this study leads to a new opportunity for tendon research and provides further insight into mechanistic understanding of the differentiation pathway to a tenogenic fate.

overall_design: Single cell-based RNA-sequencing during tenogenic induction from iPSCs and profiling of Achilles tendon mRNA of 14-day old wild type (Wt) andScx -/- (KO) mice.

first: Yuki

last: Yoshimoto

first: Akiyoshi

last: Uezumi

first: Madoka

last: Ikemoto-Uezumi

first: Shinsei

last: Yambe

first: Yusuke

last: Sotomaru

first: Chisa

last: Shukunami

first: Kazumitsu

last: Maehara

first: Kaori

last: Tanaka