title: PRC2 shields naïve human pluripotent cells from trophectoderm and mesoderm differentiation [scRNA-seq]

city: Solna

first: Simon

last: Elsässer

address: Tomtebodavägen 23A

department: Medical Biochemistry and Biophysics

institute: Karolinska Institutet

summary: Single-cell RNA-seq reveals that Polycomb repressive complex 2 (PRC2) maintains naïve pluripotency and restricts an intrinsic capacity of pre-implantation pluripotent stem cells to give rise to trophectoderm and mesoderm lineages. Inhibition of PRC2 forces naïve hESC into an ‘activated’ state through which differentiation into either trophectoderm or mesoderm lineages is triggered. This trajectory is distinct from embryonic lineage specification out of the post-implantation pluripotent state, hence PRC2-mediated repression provides a highly adaptive mechanism to restrict lineage potential during early human development.

overall_design: Primed and naïve cells were grown in respective media, supplemented with 10µM EZSolution™ EPZ-6438 (BioVision; 2428-5) for seven days, with daily media changes. Samples EZH2i d2, d4 and untreated control naïve as well as EZH2i d7 and untreated control primed cells were multiplexed following the manufacturer’s protocol. In brief, cells were dissociated, treated with mouse feeder removal beads (naïve cells), counted and resuspended in Cell Multiplexing Oligo. After oligo incubation, cells were washed for a total of three times using DPBS-/- containing 1% BSA (Sigma Aldrich; A7284) and counted using the NucleoCounter® NC-200™ automated cell counter. At this point, cells were pooled accordingly: EZH2i d2, d4 and WT naïve cells, and EZH2i d7 and WT primed cells, respectively, and each pool was counted once more.

first: Banushree

last: Kumar

first: Carmen

last: Navarro

first: Nerges

last: Winblad

first: John

middle: P

last: Schell

first: Cheng

last: Zhao

first: Jere

last: Weltner

first: Laura

last: Baqué-Vidal

first: Angelo

last: Mantero

first: Sophie

last: Petropoulos

first: Fredrik

last: Lanner


title: Neoantigen specific CD4+ T cells in human melanoma have diverse differentiation states and correlate with CD8+ T cell, macrophage, and B cell function

status: Public on Mar 12 2022

city: Seattle

first: Joshua

last: Veatch

state: Washington

address: 1100 Fairview Ave N

country: USA

institute: Fred Hutchinson Cancer Research Center

type: Expression profiling by high throughput sequencing

summary: Tumor antigen-specific CD4+ T cells are required for the efficacy of immune checkpoint inhibitors in murine models but their contributions in human cancer are less understood. We used targeted single cell RNA sequencing and matching of T cell receptor sequences to identify signatures and functional correlates of tumor antigen-specific CD4+ T cells infiltrating human melanoma tumors. CD4+ T cells that recognize tumor-specific neoantigens express CXCL13 and are subdivided into clusters expressing memory and T follicular helper markers, and those expressing cytolytic markers, exhaustion markers and IFN-. In a cohort of melanoma patients, the frequency of CXCL13+ CD4+ T cells in the tumor correlated with the transcriptional states of CD8+ T cells and macrophages, maturation of B cells, and patient survival. Similar correlations were observed in a breast cancer cohort. These results identify distinct phenotypes and functional correlates of tumor antigen-specific CD4+ T cells in melanoma and suggest the possibility of using such cells to modify the tumor microenvironment.

overall_design: Cryopreserved single cell suspensions from melanoma tumors from 20 patients were stained with oligonucleotide labelled antibodies and sorted for either lymphocytes or non-lymphocyte CD45+ hematopoietic cells and subjected to single cell sequencing targeted to TCR and VDJ recombination events and a panel of immune response genes using the BD Rhapsody platform.

middle: R

first: Sylvia

middle: M

last: Lee

first: Carolyn

last: Shasha

first: Naina

last: Singhi

first: Julia

middle: L

last: Szeto

first: Ata

middle: S

last: Moshiri

first: Teresa

last: Kim

first: Kimberly

last: Smythe

first: Paul

last: Kong

first: Matthew

last: Fitzgibbon

first: Brenda

last: Jesernig

first: Shailender

last: Bhatia

first: Scott

last: Tykodi

first: Evan

middle: T

last: Hall

first: David

last: Byrd

middle: A

last: Thompson

first: Venu

middle: G

last: Pillarisetty

first: Thomas

last: Duhen

first: A

last: Houghton

last: Newell

first: Raphael

last: Gottardo

first: Stanley

last: Riddell


title: Expression of immune-related genes in the normal colorectal mucosa and colorectal carcinoma samples from Lynch syndrome individuals

status: Public on Mar 14 2022

city: Heidelberg

first: Jan

last: Budczies

address: Im Neuenheimer Feld 224

department: Institute Of Pathology

country: Germany

institute: Heidelberg University Hospital

summary: Colorectal carcinomas arising in the context of Lynch syndrome, the most common inherited cancer syndrome, typically show deficiency of the DNA MMR (mismatch repair) system. Lack of functional MMR leads to accumulation of frameshift mutations at micosatellites (microsatellite instability, MSI). High load of highly immunogenic tumor-specific frameshift neoantigens results in strong immune response against Lynch syndrome MSI cancers. Previous studies have shown systemic immune responses against frameshift neoantigens in Lynch syndrome carriers long before tumor manifestation. In the present study, we analyzed the immune profile of normal colorectal mucosa in Lynch syndrome carriers without current or previous cancer history and in Lynch syndrome colorectal cancer patients, as well as of Lynch syndrome colorectal carcinomas. The unsupervised cluster analysis of gene expression data revealed a sharp differentiation between normal mucosa from Lynch syndrome individuals with and without manifest cancer as well as between normal mucosa in general and Lynch syndrome cancer tissue. Deconvolution analysis for predicting the prevalence of immune cell population among the three groups revealed 10 out of 14 investigated populations to be significantly different between the three tissue types (FDR=10%). In contrast to normal mucosa samples, tumor tissue showed overrepresentation of immune-suppressive cell populations, such as regulatory T cells and neutrophils. Taken together with the quantitative T cell density analysis on the basis of immunohistochemical T cell stainings, our data show strong immune infiltration of the normal colorectal mucosa in Lynch syndrome individuals even in the absence of a manifest cancer.

overall_design: Tissue specimens of normal colorectal mucosa from healthy, cancer-free Lynch syndrome carriers (n=10), Lynch syndrome CRC patients (n=10) and specimens of Lynch syndrome colorectal carcinoma samples (n=10) were analyzed for the expression of 770 immune-relevant genes using NanoString nCounter platform (PanCancer Human IO 360TM Panel). Deconvolution analysis was performed to predict the prevalence of 14 immune cell types in LS mucosal and tumor samples.

first: Lena

last: Bohaumilitzky

first: Klaus

last: Kluck

first: Robert

last: Hüneburg

first: Richard

last: Gallon

first: Jacob

last: Nattermann

first: Martina

last: Kirchner

first: Glen

last: Kristiansen

first: Oliver

last: Hommerding

first: Pauline

last: Pfuderer

first: Lelia

last: Wagner

first: Fabian

last: Echterdiek

first: Svenja

last: Kösegi

first: Nico

last: Müller

first: Konstantin

last: Fischer

first: Nina

last: Nelius

first: Ben

last: Hartog

first: Gillian

last: Borthwick

first: Elena

last: Busch

first: Georg

last: Haag

first: Hendrik

last: Bläker

first: Gabriela

last: Möslein

first: Magnus

last: von Knebel Doeberitz

first: Toni

last: Seppälä

first: Maarit

last: Ahtiainen

first: Jukka-Pekka

last: Mecklin

first: D

last: Bishop

first: John

last: Burn

first: Albrecht

last: Stenzinger

first: Matthias

last: Kloor

first: Aysel

last: Ahadova


title: Expansion, persistence and efficacy of donor memory-like NK cells for the treatment of post-transplant relapse

status: Public on Mar 11 2022

city: Cambridge

first: Guangan

last: Hu

state: MA

address: 500 Main Street

department: Koch Institute

institute: MIT

summary: Relapse of AML occurs in up to 50% of patients after allogeneic hematopoietic stem cell transplant (HCT) and heralds an extremely poor prognosis. Cytokine induced memory-like (CIML) human NK cells have enhanced ability to recognize and kill leukemia targets. We initiated a phase I trial of CIML NK cells to treat patients with relapsed myeloid malignancies after haplo-HCT (NCT04024761). Transcriptional profiling analysis by scRNAseq were used to characterize the immunophenotypes of infused NK cells post cell adoptive transfer and as well as their interactions with tumor cells.

overall_design: Patients with relapsed myeloid disease, including AML, MDS, myeloproliferative neoplasm (MPN), or BPDCN following haplo-HCT were eligible to be treated on an ongoing single center phase I dose de-escalation clinical trial of CIML NK cell therapy. CIML NK cells were derived from the same donor who provided stem cells for the original transplant. PBMC were collected and cryopreserved at day 0, day +7, day +21–28, day +42, day +60, day +100, and relapse time points after infusion.

first: Jianzhu

last: Chen

first: Rizwan

last: Romee


title: ZRF1 knockdown with shRNA in SK-N-BE(2) cells

city: Barcelona

first: Carlos

last: Jimenez

state: Barcelona

address: Pg. Vall dHebron 119-129, Edificio Collserola, lab 207

department: Pediatric Oncology

country: Spain

institute: Vall d'Hebron Institut de Recerca

summary: ZRF1 was silenced using two different shRNA in SK-N-BE(2) neuroblastoma cell line

overall_design: pLKO-NSC (Non Silencing Control) and pLKO-shZRF1 #1 and #3 lentiviral vectors were produced in HEK-293T, and viral supernatant was used to transduce SK-N-BE(2) cells. RNA extraction was performed 72 hours after transduction.

first: Miguel

middle: F

last: Segura

last: Jiménez


title: Transcriptome analysis of vascular smooth muscle cells from mice deficient for chaperone-mediated autophagy

city: Bronx

first: Julio

last: Madrigal-Matute

state: New York

address: 1300 Morris Park Ave, C504

department: Developmental and Molecular Biology

institute: Albert Einstein College of Medicine

summary: Chaperone-mediated autophagy (CMA) contributes to regulation of energy homeostasis by timely degradation of enzymes involved in glucose and lipid metabolism. Here, we report reduced CMA activity in vascular smooth muscle cells and macrophages in murine and human arteries in response to atherosclerotic challenges. We show that in vivo genetic blockage of CMA worsens atherosclerotic pathology through both systemic and cell-autonomous changes in vascular smooth muscle cells and macrophages, the two main cell types involved in atherogenesis. CMA deficiency promotes dedifferentiation of vascular smooth muscle cells and a pro-inflammatory state in macrophages. Conversely, a genetic mouse model with upregulated CMA shows lower vulnerability to the pro-atherosclerotic challenge. We propose that CMA could be an attractive therapeutic target against cardiovascular diseases.

overall_design: VSMC mRNA profiles were generated from 12-16 pasage wild type (WT) and LAMP-2AKO (L2A) cells incubated for 24h in DMEM media plus 5% LPDS with or without with or without LDL (150 μg/ml LDL-cholesterol) plus 0.1 U/ml of bovine lipoprotein lipase  by microarray, pooled in triplicate, using an Agilent Mouse 8x60K platform.

first: Jenny

last: De Bruijn

first: Kim

last: van Kuijk

first: Dario

last: Riasco-Bernal

first: Antonio

last: Diaz

first: Inmaculada

last: Tasset

first: Adrian

last: Martin-Segura

first: Marion JJ

last: Gijbels

first: Bianca

last: Sander

first: Susmita

last: Kaushik

first: Erik AL

last: Biessen

first: Simoni

last: Tiano

first: Mathieu

last: Bourdenx

first: Gregory

middle: J

last: Krause

first: Ian

last: McCracken

first: Andrew

last: Baker

first: Han

last: Jin

first: Nicholas

last: Sibinga

first: Jose Javier

last: Bravo-Cordero

first: Fernando

last: Macian

first: Rajat

last: Singh

first: Patrick

middle: C

last: Rensen

first: Jimmy FP

last: Berbee

first: Gerard

last: Pasterkamp

first: Judith

last: Sluimer

first: Ana Maria

last: Cuervo


title: The PAF1 complex promotes 3‘ processing of pervasive transcripts

status: Public on Mar 15 2022

city: Beijing

first: wei

middle: zi

last: guo

state: Beijing

address: ZhongGuanCun

country: China

institute: tsinghua university

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Persister state-directed transitioning and vulnerability in melanoma

city: Essen

first: Alexander

last: Roesch

address: Hufelandstrasse 55

department: Department of Dermatology

institute: University hospital Essen

summary: Phenotypic heterogeneity and plasticity represent major drivers of tumor relapse in melanoma. Expression of the H3K4 demethylase and pRB-binding protein JARID1B/KDM5B follows a dynamic continuum across melanoma cells. Highly JARID1B-expressing cells are enriched after short-term drug treatment. However, under persistent drug-exposure, JARID1B heterogeneity is reconstituted. Using a doxycycline-titratable system and a novel chemical enhancer of JARID1B-expression, we found that JARID1Bhigh cells are transiently 'pseudo-differentiated'. Melanoma cells use this JARID1Bhigh growth-arrested state to survive drugs, but must decrease JARID1B expression again to re-enter cell cycling for long-term tumor repopulation. Overcoming the reversion to phenotypic heterogeneity by exogenous homogenization of the JARID1Bhigh phenotype causes tumor growth arrest, also in MAPKi-resistant melanoma. Mechanistically, JARID1B represents an integral checkpoint for coordinating cell differentiation via transcriptional control, stabilization of hypophosphorylated pRB, and attenuation of cytokinetic abscission. Thus, the plasticity among tumor differentiation states per se represents a novel therapeutic target, which can be chemically overcome.

overall_design: RNAseq of melanoma cells (WM3734 and CSM152) treated with Cpd1 or control for 12h, 24h and 48h were analysed in triplicate using Illumina sequencer NextSeq 500.

first: H

last: Chauvistré

first: D

last: Picard

first: M

last: Remke

first: A


title: single-cell RNA transcriptome sequencing revealed that androgen/AR signaling promotes neutrophil precursors proliferation in bone marrow

city: Shanghai

first: Le-Xing

last: Yu

state: Shanghai

address: 225 Changhai Road

institute: Eastern Hepatobiliary Surgery Hospital

summary: The liver is one of the most-favored distant metastatic sites for solid tumors, and interactions between cancer cells and components of the hepatic microenvironment are essential for liver metastasis (LM). Although sex is one of the determinants for primary liver cancer, sexual disparity/dimorphism in LM (SDLM) and the underlying mechanisms remain unclear. We herein demonstrated a significant male-biased SDLM, which was attributed to host androgen/androgen receptor (AR) signaling that promoted hepatic seeding of tumor cells and subsequent outgrowth in a neutrophil-dependent manner. Mechanistically, androgen/AR signaling promoted hepatic accumulation of neutrophils by promoting proliferation and development of neutrophil precursors in the bone marrow, as well as modulating hepatic recruitment of neutrophils and their functions. Antagonizing the androgen/AR/neutrophil axis significantly mitigated LM in males. Our data thus reveal an important role of androgen in LM and suggest that androgen/AR modulation represents a promising target for LM therapy in men.

overall_design: Single cell RNA-sequencing of the CD45+Gr1+ cells sorted from bone marrow from a sham-operated male mouse (male), an orchiectomized-male mouse (ORX) or a female mouse (female)

first: Hong-Yang

last: Wang

first: Jiao-Jiao

last: Tang

first: Xiu-Liang

last: Cui


title: Dysfunction of S100A4+ effector memory CD8+ T cells aggravates asthma

first: Huilei

last: Zhang

address: 15, Datun road, Chaoyang District

institute: Institute of Biophysics

summary: Progressive loss of effector functions, especially IFN-γ secreting capability, in effector memory CD8+ T (CD8+ TEM) cells plays a causal role in asthma worsening. However, the mechanisms of CD8+ TEM cell dysfunction remain elusive. Here, we report that S100A4 drives CD8+ TEM cell dysfunction, impairing their protective memory response and promoting asthma worsening in OVA-induced asthma model. We find that allergic CD8+ TEM cells contain two subsets based on S100A4 expression. S100A4+ subsets exhibit dysfunctional effector phenotypes with increased proliferative capability, whereas S100A4- subsets retain effector function but are more inclined to apoptosis, giving rise a dysfunctional CD8+ TEM cell pool. Mechanistically, S100A4 upregulation of mitochondrial metabolism results in a decrease of acetyl-CoA levels, which impair the transcription of effector genes, especially ifn-γ, facilitating cell survival, tolerance and memory potential. Our findings thus reveal general insights into how S100A4 reprograms CD8+ TEM cells into dysfunctional and less protective phenotypes to promote asthma worsening.

overall_design: Protein-coding mRNA sequencing (RNA-seq) analysis of S100A4+ and S100A4- CD8+ TEM cells. There were three samples for each cell population.

first: Shuangqing

last: Liu

first: Yanan

last: Li

first: Jianru

first: Chen

last: Ni

first: Ming

last: Yang

first: Jun

last: Dong

first: Zhaoqing

first: Zhihai

last: Qin


title: RNA-seq (3-prime): Intestinal stem cell aging signature reveals a reprogramming strategy to enhance regenerative potential

status: Public on Mar 10 2022

city: Clayton

first: Monika

last: Mohenska

state: VIC

address: 15 Innovation Walk

country: Australia

institute: Monash University

summary: The impact of aging on intestinal stem cells (ISCs) has not been fully elucidated. In this study, we identified widespread epigenetic and transcriptional alterations in old ISCs. Using a reprogramming algorithm, we identified a set of key transcription factors (Egr1, Irf1, FosB) that drives molecular and functional differences between old and young states. Overall, by dissecting the molecular signature of aged ISCs, our study identified transcription factors that enhance the regenerative capacity of ISCs.

overall_design: Transcriptional (3-prime) profiles of intestinal stem cells of young (2 month) and old (22 month) mice.

first: Jose

last: Polo

first: Christian

last: Nefzger

first: Helen

middle: E

last: Abud

first: Thierry

last: Jarde


title: RNA-seq expression profiling of cerebral cortices from Taf8 deleted and Taf8 intact mouse embryos

city: Parkville

first: Gordon

middle: K

last: Smyth

state: Vic

address: 1G Royal Pde

department: Bioinformatics

institute: Walter and Eliza Hall Institute of Medical Research

summary:
Mutations in genes encoding general transcription factors cause neurological disorders. Despite clinical prominence, the consequences of defects in the basal transcription machinery during brain development are unclear. We found that loss of the TATA-box binding protein associated factor TAF8, a component of the general transcription factor TFIID, in the developing central nervous system affected the expression of many, but notably not all genes. Taf8 deletion caused apoptosis, unexpectedly restricted to forebrain regions. Nuclear levels of the transcription factor p53 were elevated in the absence of TAF8, as were the mRNAs of the pro-apoptotic p53 target genes Noxa, Puma and Bax. The cell death in Taf8–/– forebrain regions was completely rescued by additional loss of p53, but Taf8–/–p53–/– brains failed to initiate a neuronal expression program. Taf8 deletion caused aberrant transcription of promoter regions and splicing anomalies. We propose that TAF8 supports the directionality of transcription and co-transcriptional splicing, and that failure of these processes causes p53- induced apoptosis of neuronal cells in the developing mouse embryo.
Taf8 was deleted in the developing mouse central nervous system (CNS) using our Taf8 conditional allele. In the absence of global effects of loss of TAF8 on RNA levels, we determined effects on individual gene loci by RNA sequencing of cortices from Taf8 deleted (E14.5 Taf8lox/–NesCreT/+p53–/–)  and control (E14.5 Taf8+/+NesCreT/+p53–/–)  cortices. The p53-deficient background was used to avoid complications due to apoptosis, but prevented us from examining changes in p53-induced gene transcription.

overall_design: Four Taf8 deleted and four control corticle samples were prepared, each from a separate mouse. Stranded and unstranded RNA-seq libraries were prepared from each RNA sample.

first: Yunshun

last: Chen

first: Farrah

last: El-Saafin

first: Tim

last: Thomas

first: Anne

last: Voss


title: Stranded RNA-seq profiling of cerebral cortices from Taf8-deleted and Taf8-intact mouse embryos

summary: Taf8 was deleted in the mouse central nervous system using our Taf8 conditional allele. To determine transcriptional changes due to Ta8 deletion in the developing CNS, stranded RNA sequencing was performed on cortices from Taf8 deleted embryos (E14.5 Taf8lox/–NesCreT/+p53–/–)  and control embryos (E14.5 Taf8+/+NesCreT/+p53–/–).

overall_design: Four Taf8-deleted and four control samples were prepared, each from a separate mouse embryo. Stranded RNA sequencing was undertaken on an Illumina NextSeq 500 to produce 81bp paired-end reads.


title: DNA Methylation Analysis of Turner Syndrome BAV

city: Portland

first: Cheryl

last: Maslen

state: OR

address: 3181 SW Sam Jackson Park Rd

department: Division of Cardiovascular Medicine

institute: Oregon Health & Science University

type: Methylation profiling by high throughput sequencing

summary: Turner Syndrome (TS) is a rare cytogenetic disorder caused by the complete loss or structural variation of the second sex chromosome. The most common cause of early mortality in TS results from a high incidence of left-sided congenital heart defects, including bicuspid aortic valve (BAV), which occurs in about 30% of individuals with TS. BAV is also the most common congenital heart defect in the general population with a prevalence of 0.5-2%, with males being three-times more likely to have a BAV than females. TS is associated with genome wide hypomethylation when compared to karyotypically normal males and females. Alterations in DNA methylation in primary aortic tissue are associated with BAV in euploid individuals. Here we show significant differences in DNA methylation patterns associated with BAV in TS found in peripheral blood. When comparing TS with BAV to TS with no heart defects we identified a differentially methylated region encompassing the BAV-associated gene MYRF, and enrichment for binding sites of two known transcription factor contributors to BAV. When comparing TS with BAV to euploid women with BAV, we found significant overlapping enrichment for ChIP-seq transcription factor targets including genes in the NOTCH1 pathway, known for involvement in the etiology of non-syndromic BAV, and other genes that are essential regulators of heart valve development. Overall, these findings suggest that altered DNA methylation affecting key aortic valve development genes contributes to the greatly increased risk for BAV in TS.

overall_design: Cohort of patients from three clinically defined groups (TS BAV, TS TAV, NS BAV) underwent bisulfite sequencing using DNA derived from whole blood followed by differential methylation analysis to identify regions of biological interest.

first: Jacob

last: Gutierrez

first: Lucia

last: Carbone


title: The PAF1 complex promotes 3‘ processing of pervasive transcripts [RNA-seq]

first: wei

last: guo

summary: The PAF1 complex (PAF1C) functions in multiple transcriptional and co-transcriptional processes. Although many aspects of the requirement for PAF1 in coding gene transcription have been heavily studied, the function of PAF1C at sites of non-coding transcription is poorly understood. eRNAs and PROMPTs are pervasive transcripts transcribed by RNA Polymerase II and rapidly degraded after 3’ endonucleolytic cleavage by the Integrator complex (Integrator). Here we show that PAF1C depletion results in near universal 3’ end processing defects in pervasive transcripts, leading to transcription termination dysregulation. Mechanistically, PAF1C recruits Integrator to chromatin through direct interactions promoting timely cleavage and transcription termination of pervasive transcripts. We also show that PAF1C recruits Integrator to the promoters of coding genes, where PAF1C then dissociates from the Integrator complex upon entry into processive elongation. Our results demonstrate an unexpected function for PAF1C in limiting the length and accumulation of pervasive transcripts, while highlighting a general requirement for PAF1C in Integrator recruitment.

overall_design: Examination of transcriptome of Set1d knockout and Paf1, Ctr9, Dis3 and Ints11 knockdown in mouse pancreatic ductal adenocarcinoma cells

first: Xinhong

last: Liu

first: Ziwei

last: Guo

first: Han

last: Jing

first: Bin

last: Zhang

first: Xiaoyu

last: Hu

first: Charles

last: David

first: Mo


title: Epigenetic activation of plasmacytoid DC drives IFNAR-dependent therapeutic differentiation of AML [ChIPseq DC]

city: Melbourne

first: Izabela

last: Todorovski

address: 305 Grattan Street

department: Cancer Research Division

institute: Peter MacCallum Centre

summary: Pharmacological inhibition of epigenetic enzymes can have therapeutic benefit against hematological malignancies.  Apart from affecting tumor cell growth and proliferation, these epigenetic agents may mediate anti-tumor immunity. Here we discovered a novel immuno-regulatory mechanism through inhibition of histone deacetylases (HDACs). In models of AML, leukemia cell differentiation and therapeutic benefit mediated by the HDAC inhibitor panobinostat required activation of the type I interferon (IFN) pathway. Plasmacytoid dendritic cells (pDCs) produced type I IFN after panobinostat treatment, through transcriptional activation of IFN genes concomitant with increased H3K27 acetylation at these loci. Depletion of pDCs abrogated panobinostat-mediated activation of type I IFN signaling in leukemia cells and impaired therapeutic efficacy, while combined treatment of panobinostat and IFN improved outcomes in mouse and human models. These discoveries offer a new therapeutic approach for AML and demonstrate that epigenetic rewiring of pDCs enhances anti-tumor immunity, opening the possibility of exploiting this population for immunotherapies. profiling by high throughput sequencing.

overall_design: Following two days of treatment with panobinostat or vehicle, plasmacytoid dendritic cells (pDCs) and conventional DCs (cDCs) were sorted from leukemia-bearing mice.

first: Pilar

last: Dominguez

first: Stephin

last: Vervoort


title: Transcriptomes of Enterococcus faecalis OG1RF and OG1RF Δbph in planktonic culture

city: Minneapolis

first: Julia

last: Willett

state: Minnesota

address: 689 23rd Avenue SE, 4-149 Micro Research Facility

institute: University of Minnesota

summary: Liquid cultures of Enterococcus faecalis OG1RF and OG1RF Δbph were grown in tryptic soy broth without added dextrose (TSB-D) for 2 and 4 hr.  At each time point, the transcriptomes were compared to identify differentially expressed genes in the Δbph mutant.

overall_design: 2 biological replicates per strain were analyzed at each time point.  Parental OG1RF (reference genome NC_017316.1) was used as a control.

first: Gary

last: Dunny


title: β-Ecdysterone Enhanced Bone Regeneration Through The BMP-2/SMADs/RUNX2/Osterix Signaling Pathway

status: Public on Mar 09 2022

city: 南充市

first: Cai-Ping

last: Yan

state: sichuan

address: No. 1 The South of Maoyuan Road

department: orthopedics

institute: North sichuan medical college

summary: Bone defects are a global public health problem. However, the available methods for bone regeneration are limited. In recent years, the application of traditional Chinese herbs for bone regeneration has attracted increasing attention. β-ecdysterone is a herbal extract similar to estrogen that promotes the metabolism of cell protein synthesis; however, its function on bone regeneration is not yet clear. In this study, we investigated the function of β-ecdysterone on osteoblast differentiation and bone regeneration in vitro and in vivo. MC3T3-E1 cells were used to test the function of β-ecdysterone on osteoblast differentiation and bone regeneration in vitro. The results of the CCK-8 assay suggested that the proliferation of MC3T3-E1 cells was promoted by β-ecdysterone. Furthermore, β-ecdysterone influenced the expression of osteogenesis-related genes, and the bone regeneration capacity of MC3T3-E1 cells was detected by PCR, the alkaline phosphatase (ALP) test, and the Alizarin Red test. β-ecdysterone could up-regulate the expression of osteoblastic-related genes, and promoted ALP activity and the formation of calcium nodules. Moreover, we also determined that β-ecdysterone increased the mRNA and protein levels of components of the BMP-2/Smad/Runx2/Osterix pathway. DNA sequencing further confirmed these target effects. β-ecdysterone promoted bone formation by enhancing gene expression of the BMP-2/Smad/Runx2/Osterix signaling pathway and by the enrichment biological processes. For in vivo experiments, the femur condyle defection model was constructed by drilling a critical-sized (3×3 mm) defect in 12-week-old SD male rats to further assess the bone regeneration functions of β-ecdysterone. The results of micro-CT showed that β-ecdysterone could accelerate bone regeneration and exhibited higher bone volume, bone surface, bone mineral density at each observation time point. Immunohistochemistry confirmed that the β-ecdysterone also increased the expression of collagen, osteocalcin, and BMP2 in the experiment group at 8 weeks. In conclusion, discovered β-ecdysterone is a new bone regeneration regulator that can stimulate MC3T3-E1 cell proliferation and induce bone regeneration through the BMP-2/Smad/Runx2/Osterix pathway. This new function of β-ecdysterone has revealed a new direction of osteogenic differentiation and has provided novel therapeutic strategies for treating bone defection.

overall_design: To observe and compare gene expression in MC3T3-E1 cells treated with β-ecdysterone, we performed RNA sequencing of the samples.

first: Xing-Kuan

last: Wang

first: Ke

last: Jiang

first: Yu-Ling

last: Li

first: Chong

last: Yin

first: Chao

last: Xiang

first: Yong

first: Chaoyu

last: Pu

first: Lu

last: Chen


title: Epigenetic activation of plasmacytoid DC drives IFNAR-dependent therapeutic differentiation of AML [RNAseq IFNAR KO tumor cells]

overall_design: Following three days of treatment with panobinostat or vehicle, leukemia tumor cells (GFP+) were sorted, using propidium iodide (PI) as a viability marker (3 replicates per group).


title: Differentially expressed genes in Mettl3-deficient livers compared to control livers

city: Jinan

first: Detian

last: Yuan

address: Wenhuaxi 44

institute: Shandong University

summary: Liver-specific deficiency of Mettl3 causes liver injury. By performing RNA sequencing (RNA-seq) analysis on the Mettl3-deficient versus control livers, we identified the potential target genes that were closely associated with the liver phenotype in liver-specific Mettl3 knockout mice. RNA-seq analysis revealed extensive metabolic reprogramming in Mettl3-deficient livers. These results demonstrated that Mettl3 coordinates metabolic homeostasis and functional maturation during postnatal liver development.

overall_design: mRNA profiles of 3 wild type (WT) and 3 Mettl3-deficient livers

first: Shiguan


title: Lamin B1 deletion in myeloid neoplasms alters hematopoietic cell fate and causes Pelger-Huët nuclear anomaly

first: Sergei

last: Doulatov

state: WA

address: 1705 N.E. Pacific Street

department: Hematology

institute: University of Washington

summary: The most common cytogenetic abnormality found in complex karyotype high risk MDS/AML is del5q. Disease is often identified through visual inspection and recognition of abnormalities such as pseudo-Pelger-Huet anomaly (PPHA), which presents as deviations of normal neutrophil nuclear morphology. The link between PPHA and leukemia is unknown, but mutations in the lamin B1 receptor (LBR) have been identified as the cause of benign presence in otherwise normal individuals. Lamin B1 (LMNB1) is the ligand for LBR and is important for maintaining nuclear lamina integrity and influencing 3D gene structural changes that impact transcription; the LMNB1 gene locus is also contained with chromosome 5q, so we hypothesized that appearance of PPHA is potentially caused by del5q in MDS/AML containing the LMNB1. To help characterize the potential role of LMNB1 in normal hematopoietic stem and progenitor cells (HSPCs), we infected CD34+ cord blood HSPCs with lentiviruses expressing shRNAs targeting LMNB1 (LB1) and luciferase (C11) as a control and assessed changes in transcription due to LMNB1 disruption via RNSeq and 3D genome structural changes through HiC. Neutrophils were also differentiated from LMNB1 shRNA and control treated CD34+ cord blood HSPCs and transcriptional/structural changes identified via RNAseq and HiC. Additionally, to understand long-term in vivo effects of LMNB1 loss on self-renewal and lineage bias, we engrafted control and LMNB1 shRNA infected CD34+ HSPCs into NSG mice and harvest hCD34+ cells 12 weeks later for scRNAseq.

overall_design: RNAseq, HiC, and 10X genomics scRNAseq

first: Serge

first: J.

last: Creamer

first: Andreea

last: Reilly

first: Jian

last: Ma

first: Zhijun

last: Duan

first: Yuchuan

first: Rachel

last: Wellington


title: Epigenetic activation of plasmacytoid DC drives IFNAR-dependent therapeutic differentiation of AML [RNAseq DC]

first: Izabela

last: Todorovski

overall_design: Following two days of treatment with panobinostat or vehicle, plasmacytoid dendritic cells (pDCs) and conventional DCs (cDCs) were sorted from leukemia-bearing mice (2 biological replicates per group; pool of 3-6 mice per sample).

first: Pilar

last: Dominguez

first: Stephin

last: Vervoort


title: Epigenetic activation of plasmacytoid DC drives IFNAR-dependent therapeutic differentiation of AML [ATACseq DC]


title: Epigenetic activation of plasmacytoid DC drives IFNAR-dependent therapeutic differentiation of AML


title: Intestinal stem cell aging signature reveals a reprogramming strategy to enhance regenerative potential

first: Monika

last: Mohenska


title: Epigenetic activation of plasmacytoid DC drives IFNAR-dependent therapeutic differentiation of AML [RNAseq WT tumor cells]


title: Pathogenic mutation of TDP-43 impairs RNA processing in a cell type-specific manner

city: Stanford

first: Kent

last: Imaizumi

state: CA

address: 318 Campus Drive

institute: Stanford University

summary: Gene expression profiling of human iPSC, iPSC-derived neural progenitors, and  iPSC-derived neurons with or without TARDBP K263E mutation

overall_design: 3 samples for each condition.

first: Hideyki

last: Okano


title: Mettl3-dependent m6A modification in postnatal liver

overall_design: mRNA was isolated and subject to m6A RNA immunoprecipitation, fractioning each mRNA pool into input, anti-m6A-antibody-tagged samples. Libraries were prepared and sequenced from each fraction, with 3 biological replicates per fraction per wild-type or Mettl3-deficient mice.

last: Wang


title: Next generation sequencing provides valuable insights into the global view of the transcriptome dynamics in Leishmania cells with depleted levels of profilin as compared to control cells.

city: Bangalore

first: Purna Sudha

middle: Bindu

last: Ambaru

state: Karnataka

address: Biotech park, Electronic city, phase 1,, hosur main road, Bangalore

country: India

institute: Institute of Bioinformatics and Applied Biotechnology

summary:
Transcriptome analysis via high-throughput sequencing (RNA-Seq) captures significant changes in gene expression. The purpose of this study is to elucidate the transcriptomic changes in profilin deficient Leishmania promastigotes (LdPfn+/-) as compared to wild type control (LdPfn+/+). This research will enable us to gain insight into the role of  profilin in Leishmania cells. The data has been validated by qRT-PCR. Data analysis results are further supported by flow cytometry, immunofluorescence experiments and by western blot demonstrating a depletion in the protein expression of a eukaryotic translation initiation factor in the cells with depleted levels of profilin protein.
Methods: Total RNA from three independent biological replicates from each of wild-type (LdPfn+/+) and profilin knockout (LdPfn+/-) Leishmania promastigotes was extracted and after library preparation, RNA sequencing was carried out on Illumina HiSeq 2500.  The sequence reads that passed quality filters were aligned to the Leishmania donovani (BPK282A1) genomic data obtained from TriTrypDB version 51 using  Bowtie2 (-x option). All analyses were carried out using the Tophat pipeline with the following versions: Tophat v2.1.1, Bowtie2 v2.3.5.1. The HTSeq version 0.12.4 (htseq-count -f option) was used to count the number of reads aligned to protein-coding genes. DeSeq tool was used for differential gene expression analysis between samples in protein-coding genes. qRT-PCR validation was performed using SYBR Green assay.
Results: RNA-Seq datasets generated in this study with each sample control (LdPfn+/+) and profilin knockout (LdPfn+/-) have at least 30 million read pairs. RNA-seq data were aligned to the L.donovani genome (Leishmania donovani BPK282A1, NCBI taxon ID: 981087) and 8135 transcripts were identified in our data set. Data analysis revealed that a total of 254 genes were differentially expressed (DE) having a p-value <0.05. The data obtained from RNA-Seq have been validated by real-time PCR of 18 genes, 8 UP regulated and 10 down-regulated. Results of qRT-PCR analysis showed a strong correlation with the RNA-seq data. Inferences drawn from transcriptomic analysis were further validated by flow cytometry, immunofluorescence experiments and western blotting, revealing a critical role during the early phases of Leishmania cell division.
Conclusions: Present study represents a comprehensive report elucidating the transcriptomic changes in profilin deficient Leishmania promastigotes (LdPfn+/-) as compared to wild type (LdPfn+/+). This RNA-Seq data along with validation by qPCR, flow cytometry experiments, and immunofluorescent microscopy experiments indicate that profilin might play a critical role during the early phases of Leishmania cell division.

overall_design: mRNA profiles of wild type (LdPfn+/+) and profilin deficient (LdPfn+/-) Leishmania donovani promastigotes.

first: Bindu

first: Ganesh

last: Gangadharan

first: Hosahalli

last: Subramanya

first: Chhitar

last: Gupta


title: Bulk RNAseq anaysis of PSCs and differentiating cells under on-feeder and feeder-free cultures

summary: Gene expression profiling of human iPSCs [201B7] under on-feeder, feeder-free [StemFit], and FGF inhibitor [100 nM PD173074] -treated cultures, and differentiating cells.

first: Hirosato

last: Ideno


title: YAP mediates TNF effect on pro-fibrotic genes expression

city: Saint-Priest-en-Jarez

first: Robin

last: Caire

address: 10 rue de la Marandière

department: Biology

country: France

institute: Université Jean Monnnet Saint-Etienne

summary: YAP/TAZ are transcription co-factors recently described responsive to pro-inflammatory cytokines and involved in inflammatory-related disorders. However, the role of tumor necrosis factor (TNF), a major pro-inflammatory cytokine, on YAP signaling is not well understood and controversial. Here, we observe in vitro, using wild type and YAP knockout HEK293 cells, that TNF triggers YAP nuclear translocation and transcriptional activity, thus being dependent on Rho family of GTPases. In response to TNF, YAP transcriptional activity orientates cell fate toward survival. Transcriptional analysis with Nanostring technology reveals that YAP modulates TNF-induced increase in fibro-inflammatory pathways such as NF-κB, inflammasomes, cytokines or chemokines signaling as well as pro-fibrotic pathways involving TGF-β and extracellular matrix remodeling. Therefore, in response to TNF, YAP acts as a sustainer of the inflammatory response and as a molecular link between inflammation and fibrotic processes. This work identifies that YAP is critical to drive several biological effects of TNF which are involved in cancer and inflammatory disorders.

overall_design: HEK293 cells wild type (WT) or YAP knockout (YAP KO or YAP-/-), control (CTRL) or TNF treated were used. 12 samples were analyzed corresponding to 4 groups: WT CTRL, YAPKO CTRL, WT TNF, YAPKO TNF with n=3 samples per group. 4 comparisions were made between groups : WT CTRL vs WT TNF; WT CTRL vs YAPKO CTRL;  YAPKO CTRL vs YAPKO TNF; WT TNF vs YAPKO TNF

first: Hubert

last: Marotte


title: RNA-seq analysis of UFM1 deletion effect on gene expression during the RIG-I antiviral response

city: Durham

first: Stacy

last: Horner

state: North Carolina

address: 213 Research Drive, CARL 0049

department: MGM

institute: Duke University

summary: UFM1 is a post-translational modification that regulates protein function during the ER stress response, DNA damage response, and antiviral response. The goal of this experiment was to determine the role of UFM1 in regulating gene expression induced during RIG-I signaling. We found that deletion of UFM1 broadly decreases transcription of some interferon-stimulated genes during activation of antiviral innate immunity. In subsequent experiments, we found that this is due to UFM1/ufmylation regulation of RIG-I interaction with its trafficking protein 14-3-3epsilon.

overall_design: Wild-type or UFM1 CRISPR/CAS9 knock-out 293T cells were seeded in 6-well plates and infected with Sendai virus (SenV) (200HAU) or mock infection for 18 hours. RNA was extracted and RNA-seq was performed. 3 biological replicates were performed for each condition.

first: Daltry

last: Snider

first: Moonhee

last: Park

first: Kristen

last: Murphy

first: Dia

last: Beachboard


title: Epigenetic activation of plasmacytoid DC drives IFNAR-dependent therapeutic differentiation of AML [scRNAseq DC]

first: Izabela

last: Todorovski

overall_design: Following two days of treatment with panobinostat or vehicle, dendritic cells were isolated from leukemia-bearing mice through Optiprep gradient, followed by sorting of GFP- cells and magnetic negative selection using EasySep Mouse Pan-DC Enrichment Kit (Stem Cell Technologies, Cat# 19763). Cells then were incubated with Cell Hashing HTO-conjugated antibodies (Biolegend, TotalSeq™ anti-mouse Hashtag reagents Cat# A0301-A0306) and CITE-seq antibodies (Biolegend, TotalSeq™-A0106 anti-mouse CD11c and A0119 anti-mouse Siglec H). A total of 6 samples (vehicle and pano; leukemia-bearing mice and WT mice) were hashtagged and pooled for a single capture reaction.

first: Pilar

last: Dominguez

first: Stephin

last: Vervoort


title: A plant immune protein enables broad antitumor response by rescuing microRNA deficiency in cancers  [miRNA-seq]

status: Public on Mar 08 2022

first: Li

last: Ding

address: No.5, Yiheyuan Road

institute: Peking University

type: Non-coding RNA profiling by high throughput sequencing

summary: Cancer cells are featured with uncontrollable activation of cell cycle and proliferation,and microRNA deficiency drives tumorigenesis. The RNA-dependent RNA polymerase (RDR) is essential for small RNA-mediated immune response in plants, but absent in vertebrates with adaptive immunity. Here we show that ectopic expression of plant RDR1 can generally inhibit the proliferation of different cancer cells. Interestingly, we find that abnormal AGO2-free microRNA isoforms with 1-nucleotide shortening at the 3’ end are widely accumulated in many tumors. RDR1 with nucleotidyltransferase activity can recognize and modify these problematic microRNAs with mononucleotides to restore their AGO2 loading efficiency, which eventually rescues microRNA deficiency and elevates microRNA expression to target cell cycle in cancer specifically. The wide antitumor effects of RDR1 can be visualized in multiple xenograft tumor models in vivo, which can be realized by adeno-associated virus-mediated delivery. Altogether, we develop a broad microRNA-mediated antitumor stratagem using the plant immune protein RDR1.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Seven solid tumors (A549, HCT116, HepG2, HeLa, H1299, PC-3 and U-2 OS), three leukemia (Jurkat, K562 and NALM6), and three non-cancer somatic cell lines (NIH/3T3, RPE-1 and WPMY-1), as well as two embryonic stem cell lines (mouse V6.5 and human WIBR3) with or without RDR1 expression were used for RNA-seq at different time points. Small RNA-seq were performed at A549, HCT116, HepG2, HeLa, H1299, Jurkat, NIH/3T3 and RPE-1 cell lines with or without RDR1 expression for 5 or 6 days. siRNA-mediated AGO2 knockdown in cancer A549 cells with or without RDR1 expression were used for small RNA-seq to analyze the tailing percentage of miRNAs (siNC, Negative control). AGO2-IP followed by small RNA-seq analysis was performed from the A549 cell lines and HepG2 cell lines with inducible RDR1 expression (including miR-34c transfection assay). Tumor samples from the A549, H1299, PC-3, Jurkat and NALM-6 xenograft models were used for RNA-seq analysis (including AAV-mediated RDR1 delivery in A549 xenografts). EGFP+ cells in the peripheral blood of Jurkat xenotransplantation mice were sorted from vector group or AtRDR1 group at different time and used for single-cell RNA-seq. For nanoparticle-mediated RDR1 delivery, A549, HeLa, H1299, NIH/3T3 and RPE-1 cells were transfected with RDR1-containing nanoparticles and used for RNA-seq.

first: Ye

last: Qi

first: Peng

last: Du


title: MCM complexes are barriers that restrict cohesin-mediated loop extrusion [RNA-Seq]

city: Vienna

first: Bart

middle: Johan H

last: Dequeker

address: Dr. Bohr-Gasse 3

country: Austria

institute: IMBA - Institute of Molecular Biotechology

summary: Eukaryotic genomes are compacted into loops and topologically associating domains (TADs), which contribute to transcription, recombination and genomic stability. Cohesin extrudes DNA into loops that are thought to lengthen until CTCF boundaries are encountered. Little is known about whether loop extrusion is impeded by DNA-bound machines. Here we show that the minichromosome maintenance (MCM) complex is a barrier that restricts loop extrusion in G1 phase. Single-nucleus Hi-C of mouse zygotes revealed that MCM loading reduces CTCF-anchored loops and decreases TAD boundary insulation, suggesting loop extrusion is impeded before reaching CTCF. This effect extends to HCT116 cells, where MCMs affect the number of CTCF-anchored loops and gene expression. Simulations suggest that MCMs are abundant, randomly positioned, partially permeable barriers. Single-molecule imaging shows that MCMs are physical barriers that frequently constrain cohesin translocation in vitro. Remarkably, chimaeric yeast MCMs harbouring a cohesin-interaction motif from human MCM3 induce cohesin pausing, suggesting that MCMs are “active” barriers with binding sites. These findings raise the possibility that cohesin can arrive by loop extrusion at MCMs, which determine the genomic sites at which sister chromatid cohesion is established. Based on in vivo, in silico and in vitro data, we conclude that distinct loop extrusion barriers shape the 3D genome.

overall_design: 6 RNA-Seq libraries were generated from HCT116 MCM2-mAID cells

first: Johanna

last: Gassler

first: Sean

last: Powell

first: Imre

last: Gaspar

first: Ilya

last: Flyamer

first: Kikuë

last: Tachibana


title: A plant immune protein enables broad antitumor response by rescuing microRNA deficiency in cancers

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

overall_design: Refer to individual Series


title: RNA-seq analysis of metabolic tissues in high salt diet fed mice

city: shanghai

first: Fei

last: Mao

state: State...

address: Fudan university

institute: Fudan University

summary:
The goal of this study is to understand the effects of high salt diet on metabolic changes in different tissues including liver, WAT and muscle through RNA-seq.
Methods: Liver, WAT and muscle mRNA profiles of control WT and 12wks high salt diet fed mice were generated by RNA deep sequencing, using Illumina Hiseq 6000. Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm10 whole genome using Hisat2.
Results: We observed that several genes associated with de novo lipogenesis and cholesterol biosynthesis were significantly down-regulated in WAT and liver tissue of HSD mice group. Besides, combined with secretome datasets, our results further demonstrated that high salt diet could also down-regulate different organokines, and thus, possibly mediate crosstalk between different metabolic tissues
Conclusions: Our study investigated the mechanisms of changed gene expression pattern of metabolic organs induced by high salt in terms of whole transcriptome profiling.

overall_design: mRNA profiles of liver, WAT, muscle tissues of  high salt fed C57BL6J mice

first: Yao

last: Li


title: NDF Is a Transcription Factor that Stimulates Elongation by RNA Polymerase II

city: Piscataway

first: Jia

last: Fei

state: New Jersey

address: 604 Allison Road

department: Molecular Biology and Biochemistry

institute: Rutgers University - New Brunswick

summary: RNA polymerase II (Pol II) elongation is a critical step in gene expression. Here we find that NDF, which was identified as a bilaterian nucleosome-destabilizing factor, is also a Pol II transcription factor that stimulates elongation with plain DNA templates in the absence of nucleosomes. NDF binds directly to Pol II and enhances elongation by a different mechanism than does transcription factor TFIIS. Moreover, yeast Pdp3, which is related to NDF, binds to Pol II and stimulates elongation. Thus, NDF is a Pol II-binding transcription elongation factor that is localized over gene bodies and is conserved from yeast to humans.

overall_design:
Bru_seq analysis of nascent transcripts in HeLa S3 (WT) and NDF CRISPR-knockout (KO) cells.
Bru_seq analysis of nascent transcripts in SW480 (WT), NDF CRISPR-knockout (KO)  and KO rescue cells.

first: Jun

last: Xu

first: Ziwei

first: Kevin

first: Dong

last: Wang

first: George

last: Kassavetis

first: James

last: Kadonaga


title: RNA-seq (SPIA-seq): Intestinal stem cell aging signature reveals a reprogramming strategy to enhance regenerative potential

first: Monika

last: Mohenska

summary: The impact of aging on intestinal stem cells (ISCs) has not been fully elucidated. In this study, we identified widespread epigenetic and transcriptional alterations in old ISCs. Using a reprogramming algorithm, we identified a set of key transcription factors (Egr1, Irf1, FosB) that drives molecular and functional differences between old and young states. Overall, by dissecting the molecular signature of aged ISCs, our study identified transcription factors that enhance the regenerative capacity of ISCs.

overall_design: Full length transcriptional profiles of intestinal stem cells of young (2 month) and old (22 month) mice.

first: Jose

last: Polo

first: Christian

last: Nefzger

first: Helen

last: Abud

first: Thierry

last: Jarde


title: Human epigenetic and transcriptional T cell differentiation atlas for identifying functional T cell-specific enhancers [RNA melanoma]

city: Philadelphia

first: E

middle: JOHN

last: WHERRY

state: PA

address: 421 Curie Blvd,354 BRB II/III

department: Systems Pharmacology       and Translational Therapeutics,Institute for Immunology

institute: University of Pennsylvania

summary: RNA-seq was used to study the transcriptional landscape of CD8 T cells in locally advanced melanoma patients treated with neo-adjuvant/adjuvant Pembrolizumab.

overall_design: CD8 naive and Non naive cells were sorted from  PBMC and tumor samples from advanced melanoma patients treated with neo-adjuvant/adjuvant Pembrolizumab and transcriptional profiles were assayed using RNAseq for comparison analysis

first: Josephine

last: Giles

first: Sasi

last: Manne

last: Wherry


title: A plant immune protein enables broad antitumor response by rescuing microRNA deficiency in cancers [mRNA-seq]

city: Beijing

first: Li

last: Ding

first: Ye

last: Qi

first: Peng

last: Du


title: MCM complexes are barriers that restrict cohesin-mediated loop extrusion [snHi-C]

first: Bart

last: Dequeker

summary: Eukaryotic genomes are compacted into loops and topologically associating domains (TADs), which contribute to transcription, recombination and genomic stability. Cohesin extrudes DNA into loops that are thought to lengthen until CTCF boundaries are encountered. Little is known about whether loop extrusion is impeded by DNA-bound macromolecular machines. We demonstrate that the replicative helicase MCM is a barrier that restricts loop extrusion in G1 phase. Single-nucleus Hi-C of one-cell embryos revealed that MCM loading reduces CTCF-anchored loops and decreases TAD boundary insulation, suggesting loop extrusion is impeded before reaching CTCF. Single-molecule imaging shows that MCMs are physical barriers that constrain cohesin translocation in vitro. Simulations are consistent with MCMs as abundant, random barriers. We conclude that distinct loop extrusion barriers contribute to shaping 3D genomes.

overall_design: We generated 109 snHi-C libraries from G1 zygotes.

first: Johanna

last: Gassler

first: Sean

last: Powell

first: Imre

last: Gaspar

first: Ilya

last: Flyamer

first: Kikuë

last: Tachibana


title: Differential roles of BAF and PBAF subunits, Arid1b and Arid2, in MLL-AF9 leukemogenesis

city: Milwaukee

last: Zhu

state: WI

address: 8733 W Watertown Plank Rd

institute: Versiti Blood Research Institute Milwaukee


title: Bulk RNA-Seq of human primordial germ cells induced from human induced pluripotent cells

city: Minato-ku

first: Kei

last: Sato

state: Tokyo

address: 4-6-1 Shiroganedai

department: Department of Microbiology and Immunology

country: Japan

institute: The Institute of Medical Science, The University of Tokyo

summary: Mammalian germ cells first establish as primordial germ cells (PGCs). There is an increasing demand to investigate the characteristics of human germ cells including PGCs. Recent studies have established the methodologies to derive human germ cells such as PGCs from human induced pluripotent cells (iPSCs). The in vitro-derived PGCs are referred to as PGC-like cells (PGCLCs) and exhibit the characteristics of the PGCs immediately after specification. Here we performed bulk RNA-sequencing (RNA-Seq) of PGCLCs and their parental iPSCs to characterize the expression signatures of human genes and transposable elements in PGCLCs in detail.

overall_design: iPSCs (9A13 XY) line used in this study were established in the previous study (Hwang et al. [PMID: 33168808]). iPSCs were culture on a plate coated with Recombinant laminin-511 E8 (BG iMatrix-511 Silk, Peprotech) and were maintained under a feeder-free condition in the StemFit® Basic04 medium (Asinomoto) containing basic FGF (Peprotech) at 37 °C under an atmosphere of 5% CO2 in air. For the passage or the induction of differentiation, the cells were treated with a 1 to 1 mixture of TrypLE Select (Life Technologies) and 0.5 mM EDTA/PBS to enable their dissociation into single cells, and 10 mM ROCK inhibitor (Y-27632; Tocris) was added. PGCLCs were induced from iPSCs via incipient mesoderm-like cells (iMeLCs) as described previously (Sasaki et al. [PMID: 26189426]) and purified using the surface markers EpCAM and INTEGRINα6. Total RNA was extracted from iPSCs, and PGCLCs by using an RNeasy Micro Kit (QIAGEN) according to the manufacturer’s instructions. Synthesis of cDNAs using 1 ng of purified total RNA, and construction of cDNA libraries for RNA sequencing by using an SMART-Seq HT Kit (Takara) and Nextera XT DNA Library Preparation Kit (illumina) according to the manufacturer’s instructions. Libraries were sequenced using single-end sequencing protocol on an Illumina NextSeq 500 instrument.

first: Kotaro

last: Sasaki

first: Yasunari

last: Seita


title: Enhancer RNAs harness multivalent interactions to detach NELF from paused Pol II [GROSEQ, eCLIPSEQ, CHIPSEQ]

city: Bayreuth

first: Andreas

last: Pittroff

address: Universitätsstr. 30

department: BIOmac Research Center, NW1

institute: University of Bayreuth

summary: Enhancer RNAs (eRNAs) are a class of non-coding RNAs that originate from enhancers. Although eRNA transcription is a canonical feature of functionally activated enhancers in various organisms and cell types, its functionality in gene regulation has not been firmly established. This is in part due to a lack of knowledge regarding the molecular features required for eRNA function. Using the eRNA-dependent regulation of RNA polymerase II (Pol II) pause release as a model, we examined the requirement of sequence, structure and length for the ability of eRNAs to detach NELF from paused Pol II. We found no common structural or sequence motif that dictates eRNA function. Instead, efficient NELF release requires a single eRNA molecule to make multiple contacts with several NELF subunits as well as the presence of unpaired guanosine nucleosides. By revealing the molecular determinants for eRNA function, our study mechanistically links eRNAs to Pol II pause release and provides new insight into the regulation of metazoan transcription.

overall_design: GRO-, eCLIP-, or ChIP-seq profiles were generated from primary mouse cortical neurons at DIV7, comparing untreated neurons to neurons for different length of time.

first: Seung-Kyoon

last: Kim

first: Dahun

last: Um

first: Tae-Kyung


title: Differential cofactor dependencies define functionally distinct types of human enhancers (STAP-seq)

first: Vanja

last: Haberle

address: Campus-Vienna-Biocenter 1

institute: The Research Institute of Molecular Pathology (IMP)

summary: Development and homeostasis of all multicellular organisms rely on differential cell-type-specific gene expression that is regulated by non-coding genomic enhancer elements. Enhancers function through the transcription-factor-mediated recruitment of cofactors, a structurally and functionally diverse set of proteins that activate RNA polymerase II transcription at promoters. Despite the important roles of enhancers and cofactors in transcriptional regulation, it is still not clear if all cofactors are required for all enhancers or if different enhancers have distinct cofactor dependencies and whether differential cofactor dependencies could be used to functionally categorize enhancers.    Here, we quantified enhancer activities along the entire human genome using the massively parallel enhancer-activity assay STARR-seq in HCT116 cells, following the rapid auxin-inducible degradation of eight different cofactors. We identified groups of enhancers with distinct cofactor requirements, including enhancers whose activity is insensitive to the depletion of the core Mediator subunit MED14 or the bromodomain protein BRD4, respectively. In particular, Mediator seemed dispensable for P53-responsive enhancers and MED14-depleted cells in which transcription globally failed were still able to induce endogenous P53 target genes such as p21. Similarly, BRD4 was not required for genes with a CCAAT- and TATA-box proximal enhancer, including histone genes and LTR12 family retrotransposons, and for the induction of TATA-box containing heat-shock genes.     Taken together, we show that different types of enhancers with distinct cofactor dependencies exist, including enhancer types that function in the absence of well-known cofactors and are employed to regulate specific gene classes and transcriptional programs. This represents the first functional categorization of enhancers by their cofactor dependencies that improves our understanding of alternative ways to activate transcription, which can aid in developing more precise interventions to modulate gene expression.

overall_design: STAP-seq upon depletion of BRD4 in HCT116 cells with a library of candidate sequences consisting of wild-type BRD4-independent promoters, mutants for TATA- or/and CCAAT-box motifs and neutral genomic sequences with insertions of these motifs. All experiments were performed in 2 biological replicates.

first: Christoph

last: Neumayr

first: Cosmas

last: Arnold

first: Katharina

last: Bergauer

first: Martina

last: Rath

first: Michaela

last: Pagani

first: Alexander

last: Stark


summary: MOLM13 cells were used to perform Cleavage Under Targets and Tagmentation (CUT&Tag) analysis to identify genomic binding sites of ARID1B and ARID2 in MLL-AF9 cells.

overall_design: We performed CUT&Tag of ARID1B and ARID2 in MOLM13 cells.

first: Robert

last: Burns

first: Shikan

last: Zheng

first: Theresa

last: Bluemn


title: Bezafibrate reduces the elevated hepatic fumarate level in insulin-deficient streptozotocin mice [skeletal muscle tissue]

city: Neuherberg

first: Johannes

last: Beckers

address: Ingolstaedter Landstr. 1

department: Institute of Experimental Genetics

institute: Helmholtz Zentrum Muenchen

summary: In addition to high intracellular glucose level, the diabetic state is also characterized by altered metabolites, which are pivotal regulators of glucotoxic pathways. In this study we analyzed Bezafibrate (BEZ) treated Streptozotocin (STZ) mice, which showed improved glucose metabolism compared to untreated STZ controls. In order to identify the key molecules and pathways, which participate in the beneficial effects of BEZ, we analyzed the skeletal muscle, white adipose tissue (WAT) and liver samples of BEZ-treated mice using non-targeted metabolomics (NMR spectroscopy), targeted metabolomics (mass spectrometry), microarrays and enzyme activity measurements with a particular focus on the liver. The analysis of muscle and WAT demonstrated that BEZ improved the expressions of genes and levels of metabolites, which could partly regulate the beneficial effects of BEZ on the glucose metabolism. Furthermore, in the liver, BEZ treatment reduced the elevated hepatic fumarate level of STZ mice, which was accompanied by a decreased expression of urea cycle genes. Since the urea cycle is involved in the production of fumarate, which is shown to be participated in glucotoxic pathways, our data suggest that BEZ could attenuate the urea cycle, reduce fumarate level and in turn ameliorate glucotoxicity in the liver of STZ mice

overall_design: We performed gene expression microarray analysis on skeletal muscle tissue derived from mice treated with streptozotocin and bezafibrate

first: Andras

last: Franko

first: Martin

last: Irmler

first: Cornelia

last: Prehn

first: Silke

last: Heinzmann

first: Jerzy

last: Adamski

first: Jürgen-Christoph

last: von Kleist Retzow

first: Rudolf

last: Wiesner

first: Hans-Ulrich

last: Häring

last: Heni

last: Birkenfeld

last: Hrabĕ de Angelis


title: Human epigenetic and transcriptional T cell differentiation atlas for identifying functional T cell-specific enhancers [RNA HD]

first: E

last: WHERRY

summary: RNA-seq was used to study the transcriptional landscape of T cell subsets from PBMCs of healthy donors.

overall_design: CD8 and CD4 Immune cell subsets were sorted from young and older healthy donors PBMC samples and transcriptome was assayed using RNAseq for comparison across age and between various Immune cell subsets

first: Josephine

last: Giles

first: Sasi

last: Manne

last: Wherry


title: Enhancer RNAs make multivalent interactions with NELF to stimulate Pol II pause release

first: Andreas

last: Pittroff


title: Differential expression of miRNAs in trichloroethene-mediated inflammatory/ autoimmune response and its modulation by sulforaphane: Delineating the role of miRNA-21 and miRNA-69

city: Galveston

first: Nivedita

last: Banerjee

state: Texas

address: 301 University Blvd,

institute: University of Texas Medical Branch

summary: A contributory role of miR-21 and miR-690 in TCE- and its metabolite DCAC-mediated inflammation and autoimmune response

overall_design: Treated female MRL+/+ mice with TCE with/without antioxidant sulforaphane for 6 weeks

first: M

last: Khan


title: RNA-seq study on mouse bone marrow leukemia cells isolated from secondary transplant recipients receiving conditional knockout Arid1b or Arid2 MLL-AF9 cells

last: Zhu

summary: We isolated whole bone marrow leukemia cells from moribund secondary transplant recipients receiving VavCre+ (control), Arid1b fl/fl VavCre+ (knockout), Arid2 fl/fl VavCre+ (knockout), MxCre+ (control), Arid1b fl/fl MxCre+ (knockout), or Arid2 fl/fl MxCre+ (knockout) primary MLL-AF9 cells. Mice receiving MxCre+ (control), Arid1b fl/fl MxCre+ (knockout), or Arid2 fl/fl MxCre+ (knockout) MLL-AF9 cells were injected with pIpC 21 days post transplant. Total RNA was isolated followed by mRNA selection. cDNA libraries were generated and NextGen Sequencing was performed.

overall_design: We performed RNA-seq on mouse bone marrow leukemia cells from moribund secondary recipients transplanted with VavCre+ (control), Arid1b fl/fl VavCre+ (knockout), Arid2 fl/fl VavCre+ (knockout), MxCre+ (control), Arid1b fl/fl MxCre+ (knockout), or Arid2 fl/fl MxCre+ (knockout) MLL-AF9 cells.

first: Robert

last: Burns

first: Shikan

last: Zheng

last: Bluemn

first: Olivia

last: Arnold


title: Enhancer RNAs make multivalent interactions with NELF to stimulate Pol II pause release [Exo-seq]

summary: 5’ ExoSeq of total RNA (rRNA & signal recognition particle RNA depleted) from mouse cortical neurons before and after membrane depolarization by potassium chloride (KCl).

overall_design: Two libraries from KCl stimulated neurons (two biological replicates) and one library from unstimulated neurons (control) were analyzed.

first: Vladyslava

last: Gorbovytska

first: Claus-D

last: Kuhn


title: Bezafibrate reduces the elevated hepatic fumarate level in insulin-deficient streptozotocin mice [visceral adipose tissue]

first: Johannes

last: Beckers

overall_design: We performed gene expression microarray analysis on visceral adipose tissue derived from mice treated with streptozotocin and bezafibrate

first: Andras

last: Franko

first: Martin

last: Irmler

first: Cornelia

last: Prehn

last: Heinzmann

last: Adamski

last: von Kleist Retzow

last: Wiesner

last: Häring

last: Heni


title: JAGGED1 Promotes Tumor Progression through Recruitment of Macrophages and Functional Suppression of Tumor-killing T cells

first: Hanqiu

address: Qinghuayuan Sub-District, Haidian District

department: School of Medicine

institute: Tsinghua University


title: Human epigenetic and transcriptional T cell differentiation atlas for identifying functional T cell-specific enhancers

last: WHERRY


title: JAGGED1 Promotes Tumor Progression through Recruitment of Macrophages and Functional Suppression of Tumor-killing T cells [scRNA-seq]

summary: Breast cancer is one of the leading causes of cancer-related mortality in women. NOTCH signaling is a well conserved pathway which not only plays critical roles in normal development, but also in cancer progression. One of the Notch ligand, JAGGED1 is overexpressed in about 30% of breast cancer patients. However, the role of JAGGED1 in breast tumorigenesis has not been rigorously examined. By utilizing genetic engineered mouse models of mammary specific Jagged1 expression or knockout, we discover that Jagged1 promotes tumorigenesis in multiple spontaneous mammary tumor models. Jagged1 expression leads to increased infiltration of tumor associated macrophages and decreased presentation of T cells within tumor microenvironment. Depletion of macrophages or T cells by neutralizing antibodies diminishes the tumor-promoting effect caused by Jagged1. Mechanistically, Jagged1 activates Notch signaling in tumor cells, leads to increased expression and secretion of multiple cytokines, including IL-6 and WISP. These cytokines help recruit macrophages into the tumor microenvironment. Macrophages crosstalk with infiltrated T cells and inhibit their cytotoxic killing on tumor cells. In triple negative breast cancer patient samples, high expression level of JAGGED1 correlates with increased macrophage infiltration and decreased T cell infiltration within tumor tissues. JAGGED1 also promotes tumor progression in several other solid cancer types, including melanoma, and colon cancer in a T cell dependent manner. Co-administration of immune checkpoint inhibitor, PD-1 antibody with gamma-secretase inhibitor (GSI) significantly inhibits tumor growth. These findings identify a unique oncogenic crosstalk between tumor derived JAGGED1, Macrophages, and T cells to promote tumor progression.

overall_design: 10X Genomics to analyze immune subtype infiltrated in the WT/PyMT and Jagged1/PyMT mice.


title: JAGGED1 Promotes Tumor Progression through Recruitment of Macrophages and Functional Suppression of Tumor-killing T cells [RNA-seq]

overall_design: mRNA profiles of PyMT-A-Vector and PyMT-A-Jagged1 tumor cell lines.


title: Enhancer RNAs make multivalent interactions with NELF to stimulate Pol II pause release [SHAPE]

last: Pittroff

summary: A SHAPE-MaP structure probing experiment was performed on 40 eRNAs. The eRNA transcription start sites (TSSs) were identified by 5'-ExoSeq. The 5' end fragment (1-200 nucleotides) of each eRNA was cloned from mouse cortical neurons and the eRNAs were produced by in vitro transcription. Each eRNA was treated in one sample with DMSO (control) and in a second sample with the SHAPE reagent 1-Methyl-7-nitroisatoic anhydride (1M7). The 1M7 chemical  will react preferentially with the ribose 2'OH of the flexible nucleotides in single stranded regions and the produced adduct will lead to mutations during reverse transcription.

overall_design: Transcript specific SHAPE-MaP on 40 eRNAs (1-200 nt) for structural analysis


title: Differential cofactor dependencies define functionally distinct types of human enhancers

first: Vanja

last: Haberle

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

overall_design: Refer to individual Series


title: MCM complexes are barriers that restrict cohesin-mediated loop extrusion

status: Public on Mar 07 2022

first: Bart

last: Dequeker


title: Differential cofactor dependencies define functionally distinct types of human enhancers (ChIP-seq)

summary: Development and homeostasis of all multicellular organisms rely on differential cell-type-specific gene expression that is regulated by non-coding genomic enhancer elements. Enhancers function through the transcription-factor-mediated recruitment of cofactors, a structurally and functionally diverse set of proteins that activate RNA polymerase II transcription at promoters. Despite the important roles of enhancers and cofactors in transcriptional regulation, it is still not clear if all cofactors are required for all enhancers or if different enhancers have distinct cofactor dependencies and whether differential cofactor dependencies could be used to functionally categorize enhancers. Here, we quantified enhancer activities along the entire human genome using the massively parallel enhancer-activity assay STARR-seq in HCT116 cells, following the rapid auxin-inducible degradation of eight different cofactors. We identified groups of enhancers with distinct cofactor requirements, including enhancers whose activity is insensitive to the depletion of the core Mediator subunit MED14 or the bromodomain protein BRD4, respectively. In particular, Mediator seemed dispensable for P53-responsive enhancers and MED14-depleted cells in which transcription globally failed were still able to induce endogenous P53 target genes such as p21. Similarly, BRD4 was not required for genes with a CCAAT- and TATA-box proximal enhancer, including histone genes and LTR12 family retrotransposons, and for the induction of TATA-box containing heat-shock genes. Taken together, we show that different types of enhancers with distinct cofactor dependencies exist, including enhancer types that function in the absence of well-known cofactors and are employed to regulate specific gene classes and transcriptional programs. This represents the first functional categorization of enhancers by their cofactor dependencies that improves our understanding of alternative ways to activate transcription, which can aid in developing more precise interventions to modulate gene expression.

overall_design: MED1 ChIP-seq in WT and MED14-AID-tagged HCT116 cells, upon P53 induction by Nutlin-3a and/or MED14 depletion. MED14-AID-tagged cell line was treated with auxin (IAA) to deplete MED14 or water (mock) as a control, and with Nutlin-3a to induce P53 signalling on top of either auxin or mock treatment.

first: Christoph

last: Neumayr

first: Ann

last: Boija

first: Charles

last: Li

first: Katharina

last: Bergauer

first: Martina

last: Rath

first: Michaela

last: Pagani

first: Richard

last: Young

first: Alexander

last: Stark


title: Robust regulatory architecture of pan-neuronal gene expression

city: New York

first: Eduardo

last: Leyva-Diaz

state: NY

address: 1212 Amsterdam Ave

department: Dept Biological Sciences

institute: Columbia University

summary: Panneuronally expressed genes, such as genes involved in the synaptic vesicle release cycle or in neuropeptide maturation, are critical for proper function of all neurons, but the transcriptional control mechanisms that direct such genes to all neurons of a nervous system remain poorly understood in any organism. We show here that six members of the CUT superfamily of homeobox genes, which are each panneuronally expressed, control panneuronal identity specification in C. elegans. Single mutants show barely any effects on panneuronal gene expression or global nervous system function, but such effects become apparent and progressively worsen upon removal of additional CUT family members. Phenotypes of compound mutants can be rescued non-discriminately by overexpression of individual family members, indicating that gene dosage is critical for CUT gene function. Genome-wide binding profiles as well as mutation of CUT binding sites by CRISPR/Cas9 genome engineering show that CUT genes directly control the expression of panneuronal features. Lastly, we found that CUT genes act in conjunction with neuron-type specific transcription factors to control panneuronal gene expression. In conclusion, our study provides a previously missing key insight into how neuronal gene expression programs are specified and reveals a highly buffered and robust mechanism that controls the most critical functional features of all neurons.

overall_design: Immunoprecipitation of neuronal nuclei at the L1 larval stage (C. elegans) in control animals as well as CUT sextuple mutant (ceh-38;ceh-44;ceh-48;ceh-41,ceh-21,ceh-39 null)

first: Leyva-Diaz

last: Eduardo

first: Hobert

last: Oliver


title: MKL-1 Merkel cell carcinoma cells treated with EPZ011989

city: Boston

first: Ashley

middle: K.

last: Gartin

state: MA

address: 450 Brookline Ave. Mayer 444

department: Medical Oncology

institute: Dana-Farber Cancer Institute


title: Bidirectional epithelial - macrophage crosstalk promotes tissue repair in the lung

city: Charlottesville

first: Eugene

last: Ke

state: Virginia

address: P.O. Box 800734, 1340 Jefferson Park Avenue

department: MIC

institute: University of Virginia

summary: Epithelial structures are frequently damaged and injured by different insults, with rapid repair being essential to restore function and homeostasis. Dysregulated epithelial tissue repair occurs in several human disease states, yet how individual cell types communicate and interact to coordinate tissue regeneration is incompletely understood. Using RNAseq, we show that Macrophage-dependent induction of Nras and Bcas2 expression within the regenerating epithelium positively promoted epithelial proliferation, with amphiregulin from macrophages enhancing Nras expression.

overall_design: RNAseq comparing flow sorted purified epithelium from Panx1+/+ and Panx1-/- mice at day four after napthalene-induced injury

first: Christopher

last: Lucas

last: Medina

first: David

last: Dorward

first: Michael

last: Raymond

first: Brady

last: Barron

first: Turan

last: Tufan

first: Magdalena

last: Oremek

first: J

middle: I

last: Etchegaray

first: Sanja

last: Arandjelovic

first: Emily

last: Farber

first: Suna

last: Onngut-Gumuscu

first: Moria

last: Whyte

first: Andriano

last: Rossi

first: Kodi

last: Ravichandran


title: Next-generation mRNA sequencing of MKL-1 Merkel cell carcinoma cells treated with EPZ011989

summary: The overarching goal of this study was to characterize mechanisms of sensitivity to EZH2 inhibitors in Merkel cell carcinoma cell lines. To that end, early and late transcriptional changes were profiled in MKL-1 cells treated with the EZH2 inhibitor EPZ011989 for 6 or 12 days.

overall_design: MKL-1 cells were treated in triplicate with 3 µM EPZ011989 or vehicle (1% ethanol) for 6 or 12 days with refreshment every 3 days. 12 samples are analyzed.


title: Gene expression changes in 48hr APF retinas and optic lobes in Blimp-1 loss of function and gain of function flies

first: Jessica

middle: E.

last: Treisman

address: 540 First Ave, Skirball Institute

department: Cell Biology

institute: NYU Grossman School of Medicine

summary: Blimp-1 encodes a zinc-finger transcription factor, which is involved in the terminal differentiation of the Drosophila eye. We found that Blimp-1 mutants display plano-covex corneal lenses  and rhabdomere extension defects. To identify genes that it regulates in the retina, we performed an RNA-seq experiemtn to compare retinas expressing Blimp-1 RNAi or UAS-Blimp-1 in all cell types in the eye with lGMR-GAL4 to control lGMR-GAL4/+. We examined RNA from dissected retinas and attached optic lobes from 48 hr APF pupae. Among the genes that were affected, we found that the transcription factor Slow border cells (Slbo) was significantly up-regulated in Blimp-1 knockdown samples, and Slbo overexpression could recapitulate Blimp-1 loss of function phenotypes. Many other affected genes may collectively contribute to the corneal lens and photoreceptor phenotypes caused by loss of Blimp-1..

overall_design: Three biological replicates were analyzed for each tissue (48hr APF retinas and optic lobes) in each condition (lGMR>Blimp-1 RNAi, lGMR>UAS Blimp-1, lGMR-GAL4/+..

first: Hongsu

last: Wang

first: Carolyn

last: Morrison

first: Neha

last: Ghosh

first: Gerald

last: Call


title: Next-generation sequencing of CUT&RUN of MKL-1 Merkel cell carcinoma cells treated with EPZ011989

first: Ashley

last: Gartin

summary: The overarching goal of this study was to characterize mechanisms of sensitivity to EZH2 inhibitors in Merkel cell carcinoma cell lines. To that end, CUT&RUN for H3K27me3 and H3K4me3 histone marks was performed in MKL-1 cells treated with the EZH2 inhibitor EPZ011989 for 6 days.

overall_design: MKL-1 cells were treated in duplicate with 3 µM EPZ011989 or vehicle (1% ethanol) for 6 days with refreshment on day 3. Each sample was split into thirds and CUT&RUN was performed with antibodies against H3K27me3 or H3K4me3 or control IgG. 12 samples are analyzed.


title: Bezafibrate reduces the elevated hepatic fumarate in insulin-deficient mice.

first: Johannes

last: Beckers


title: Differential cofactor dependencies define functionally distinct types of human enhancers (STARR-seq)

first: Vanja

last: Haberle

summary: Development and homeostasis of all multicellular organisms rely on differential cell-type-specific gene expression that is regulated by non-coding genomic enhancer elements. Enhancers function through the transcription-factor-mediated recruitment of cofactors, a structurally and functionally diverse set of proteins that activate RNA polymerase II transcription at promoters. Despite the important roles of enhancers and cofactors in transcriptional regulation, it is still not clear if all cofactors are required for all enhancers or if different enhancers have distinct cofactor dependencies and whether differential cofactor dependencies could be used to functionally categorize enhancers. Here, we quantified enhancer activities along the entire human genome using the massively parallel enhancer-activity assay STARR-seq in HCT116 cells, following the rapid auxin-inducible degradation of eight different cofactors. We identified groups of enhancers with distinct cofactor requirements, including enhancers whose activity is insensitive to the depletion of the core Mediator subunit MED14 or the bromodomain protein BRD4, respectively. In particular, Mediator seemed dispensable for P53-responsive enhancers and MED14-depleted cells in which transcription globally failed were still able to induce endogenous P53 target genes such as p21. Similarly, BRD4 was not required for genes with a CCAAT- and TATA-box proximal enhancer, including histone genes and LTR12 family retrotransposons, and for the induction of TATA-box containing heat-shock genes. Taken together, we show that different types of enhancers with distinct cofactor dependencies exist, including enhancer types that function in the absence of well-known cofactors and are employed to regulate specific gene classes and transcriptional programs. This represents the first functional categorization of enhancers by their cofactor dependencies that improves our understanding of alternative ways to activate transcription, which can aid in developing more precise interventions to modulate gene expression. "

overall_design: Enhancer activity measured by STARR-seq upon AID-mediated depletion of individual transcriptional cofactors (COFs) in HCT116 cells. In total 11 COF-AID-tagged cell lines and the Parental cell line (control) were assayed, either with a genome-wide (8 tagged cell lines and Parental) or with a focused BAC-derived STARR-seq library (3 tagged cell lines). Each cell line was treated with auxin (IAA) or water (mock) and enhancer activity was assessed by STARR-seq after 6h of treatment, with 2-4 biological replicates performed per COF and condition.

first: Christoph

last: Neumayr

first: Katharina

last: Karner

last: Bergauer

first: Martina

last: Rath

first: Michaela

last: Pagani

first: Alexander

last: Stark


title: Profiling selective packaging of host RNA and viral RNA modification in the SARS-CoV-2 virion

status: Public on Mar 06 2022

city: Chicago

first: Tao

last: Pan

state: Illinois

address: 929 E. 57th Street

department: Biochemistry and Molecular Biology

institute: University of Chicago

summary: Viruses package host RNAs in their virions which are associated with a range of functions in the viral life cycle. Previous transcriptomic profiling on host RNA packaging were mostly focused on retroviruses. Which host RNAs are packaged in other viruses at the transcriptome level has not been thoroughly examined. Here we apply both small RNA and large RNA sequencing of SARS-CoV-2 virions from six individual isolates in Vero cell cultures to profile packaging of host RNAs in a coronavirus and to explore SARS-CoV-2 genomic RNA modifications. We find selective packaging of specific tRNAs, tRNA fragments and signal recognition particle RNA into SARS-CoV-2 virions. Virions from all individual clones package the same set of host RNAs, suggesting a common mechanism of selective packaging. We estimate that a SARS-CoV-2 virion contains up to 1 SRP RNA and 4 tRNA molecules. We identify tRNA modification differences between the virion-packaged tRNAs and those in the uninfected host cells. Furthermore, we find subgenomic viral RNAs in the virions, and uncharacterized candidate modifications in the SARS-CoV-2 genomic RNA. Our results reveal an under-explored aspect of viral-host interaction that may be explored for viral therapeutics.

sample_organism: Severe acute respiratory syndrome coronavirus 2

overall_design: Small RNA seq of six SARS CoV 2 virion isolates and large RNA seq of three Vero E6 cells to identify RNA abundance and RNA modifications

first: Noah

last: Peña

first: Wen

last: Zhang

first: Christopher

last: Watkins

first: Hala

last: Alshammary

first: Viviana

last: Simon

last: Katanksi


title: Differential cofactor dependencies define functionally distinct types of human enhancers (PRO-seq)

overall_design: Nascent RNA transcription measured by PRO-seq in WT, MED14- or BRD4-AID-tagged HCT116 cells, upon P53 induction by Nutlin-3a and/or respective COF depletion. Each COF-AID-tagged cell line was treated with auxin (IAA) to deplete the respective COF or water (mock) as a control. Each cell line was in addition treated with Nutlin-3a to induce P53 signaling on top of either auxin or mock treatment. For MED14-AID cell line, two different treatment regimens were performed: 1) 3h auxin/mock treatment followed by 3h Nutlin-3a treatment, or 2) 12h auxin/mock treatment followed by 6h Nutlin-3a treatment. All experiments were performed in 2 biological replicates.

first: Oliver

last: Hendy

first: Leonid

last: Serebreni

first: Cosmas

last: Arnold


title: Frequent 4EBP1 amplification induces synthetic dependence on FGFR signaling in cancer

status: Public on Mar 05 2022

first: Prathibha

last: Mohan

address: 1275 York Avenue

department: Cancer Biology and Genetics

institute: Memorial Sloan Kettering Cancer Center

summary: The eIF4E translation initiation factor has oncogenic properties and concordantly, the inhibitory eIF4E-binding protein (4EBP1) is considered a tumor suppressor. The exact molecular effects of 4EBP1 activation in cancer are still unknown. Surprisingly, 4EBP1 is a target of genomic copy number gains (Chr. 8p11) in breast and lung cancer. We notice that 4EBP1 gains are genetically linked to gains in neighboring genes including WHSC1L1 and FGFR1. Our results show that FGFR1 gains act to attenuate the function of 4EBP1 via PI3K mediated phosphorylation at Thr37/46, Ser65, and Thr70 sites. This implies that not 4EBP1 but instead FGFR1 is the genetic target of Chr. 8p11 gains in breast and lung cancer. Accordingly, these tumors show increased sensitivity to FGFR1 and PI3K inhibition and this is a therapeutic vulnerability through restoring the tumor-suppressive function of 4EBP1. Ribosome profiling reveals genes involved in insulin signaling, glucose metabolism, and inositol pathway to be the relevant translational targets of 4EBP1. These mRNAs are among the top 200 translation targets and are highly enriched for structure and sequence motifs in their 5’UTR that depends on the 4EBP1-EIF4E activity. In summary, we identify the translational targets of 4EBP1-EIF4E that facilitate the tumor suppressor function of 4EBP1 in cancer.

overall_design: We performed ribosome footprinting and RNA sequencing on six samples of 293T 4A cells including three untreated samples (DMSO) and three samples that were collected 2 hrs after treatment with Doxycycline (2mg/ml). DMSO treated samples are indicated as CT and Dox treated samples are indicate as Dox.

first: Joyce

last: Pasion

first: Nathalie

last: Lailler

first: Elisa

last: de Stanchina

first: Anke

last: van den Berg

first: Arjan

last: Diepstra

first: Giovanni

last: Ciriello

first: Viraj

last: Sanghvi

first: Hans-Guido

last: Wendel

first: Kamini

last: Singh


title: RNA Pol IV induces antagonistic parent-of-origin effects on Arabidopsis endosperm [PareSeq]

status: Public on Mar 04 2022

city: Cambridge

first: Satyaki

last: PRV

address: 455 Main street, Room number 549

institute: Whitehead Institute for Biomedical Research

summary: Gene expression in endosperm – a seed tissue that mediates transfer of maternal resources to  offspring – is under complex epigenetic control. We show here that plant-specific RNA  Polymerase IV mediates parental control of endosperm gene expression. Pol IV is required for the production of small interfering RNAs that typically direct DNA methylation. We compared small RNAs, DNA methylation, and mRNAs in A. thaliana endosperm from reciprocal heterozygotes produced by crossing wild-type plants to Pol IV mutants. We find that maternally and paternally acting Pol IV have divergent effects on endosperm. Losses of maternal and paternal Pol IV impact sRNAs and DNA methylation at distinct genomic sites. Strikingly, maternally and paternally-acting Pol IV have antagonistic impacts on gene expression at some  loci, divergently promoting or repressing endosperm gene expression. Antagonistic parent-of origin effects have only rarely been described and are consistent with a gene regulatory system  evolving under parental conflict

overall_design: 5' end sequencing of endosperm mRNAs from two replicates of wild-type (Ler x Col-0 F1) and three replicates of nrpd1-/- (Ler nrpd1-/- x Col-0 nrpd1-/-).

first: Mary

last: Gehring

first: PRV

last: Satyaki


title: Single cell analysis identifies the key role of basophils orchestrating Th17 immunity and kidney fibrosis

city: philadelphia

first: tomohito

last: doke

state: Pennsylvania

address: smilow, 3400 Civic Center Blvd STRC 12-188

department: Medicine

summary: Inflammation is a key component of fibrosis, however, the immune processes that orchestrate kidney fibrosis are poorly understood. By performing single-cell sequencing of a mouse model of kidney fibrosis, we identified a subset of kidney tubule cells with a profibrotic-inflammatory phenotype, characterized by the expression of cytokines and chemokines associated with immune cell recruitment. Receptor-ligand interaction analysis and experimental validation indicated that CXCL1 secreted by profibrotic tubules recruits CXCR2+ basophils. In mice, these basophils were the key source of IL-6 and were responsible for the recruitment of Th17 cells. Genetic deletion or antibody-based depletion of basophils resulted in reduced renal fibrosis. Human kidney single-cell, bulk gene expression, and immunostaining results validated the role of basophils in patients with kidney fibrosis. Collectively, these studies identify basophils as key contributors to the development of renal fibrosis and suggest that targeting these cells may be a useful strategy for the management of chronic kidney disease.

overall_design: Single cell and bulk transcriptome analysis of unilateral ureter obstruction kidneys

first: Tomohito

last: Doke

first: Daniel

middle: L

last: Aldridge

first: Ya-Wen

last: Yang

first: Jihwan

last: Park

first: Christina

last: Hernandez

first: Amin

last: Abedini

first: Michael

last: Balzer

first: Rojesh

last: Shrestra

first: Gaia

last: Coppock

first: Juan

middle: M

last: Inclan Rico

first: Seung

middle: Y

last: Han

first: Junhyonk

last: Kim

first: Sheng

last: Xin

first: Adrian

last: Piliponsky

first: Veronique

last: Lefebvre

first: Mark

last: Siracusa

first: Christopher

last: Hunter

first: Katalin

last: Susztak


title: RNA Pol IV induces antagonistic parent-of-origin effects on Arabidopsis endosperm

first: Satyaki

last: PRV


title: Effects of a vaccine against the tumor vasculature

city: Amsterdam

first: Judy

last: van Beijnum

state: Noord Holland

address: CCA rm 2.52, De Boelelaan 1118

country: Netherlands

institute: VUMC

summary: Mice were vaccinated with iBoost technology against a tumor vascular target, prior to B16F10 tumor inoculation. Tumors from control and target vaccinated mice were excised and subject to RNAseq

overall_design:
mRNA profiles of target and control vaccinated mice' B16F10 tumor tissue
The TRX_CP are treatment samples, TRX is control

middle: R

first: Else

last: Huijbers

first: Arjan

middle: W

last: Griffioen


title: CREBBP/EP300 acetyltransferase inhibition targets lineage specific enhancers to inhibit the proliferation of ER+ breast cancer cells [scATAC-Seq]

first: David

last: Mayhew

address: 500 Technology Square, Suite 700

institute: Foghorn Therapeutics

summary: MCF7 cells were treated with a CREBBP/EP300 acetyltransferase inhibitor CPI-1612 and chromatin status was measured via single-cell ATAC-seq

overall_design: MCF7 cells were treated with DMSO or CPI-1612 (50 nM).  Libraries were prepared with the 10X single-cell ATAC-seq protocol.

first: Archana

last: Reddy

first: Sungmi

last: Park-Chouinard

first: Andy

last: Conery


title: RNA Pol IV induces antagonistic parent-of-origin effects on Arabidopsis endosperm [bisulfite_sequencing]

overall_design: Sequencing of two replicates of bisulfite treated DNA from WT (Ler x Col-0 F1), mat nrpd1+/- (Ler nrpd1-/- x  Col-0 F1), and pat nrpd1+/- (Ler x Col-0  nrpd1-/-  F1)

first: Mary

last: Gehring

first: PRV

last: Satyaki


title: Endomitosis controls tissue-specific gene expression during development

city: Utrecht

first: Matilde

last: Galli

address: Uppsalalaan 8

institute: Hubrecht Institute

summary: Polyploid cells contain more than two copies of the genome and are found in many plant and animal tissues. Different types of polyploidy exist, in which the genome is confined to either one nucleus (mononucleation) or two or more nuclei (multinucleation). Despite the widespread occurrence of polyploidy, the functional significance of different types of polyploidy are largely unknown. Here, we assess the function of multinucleation in C. elegans intestinal cells through specific inhibition of binucleation without altering genome ploidy. Through single worm RNA sequencing, we find that binucleation is important for tissue-specific gene expression, most prominently for genes that show a rapid upregulation at the transition from larval development to adulthood. Regulated genes include vitellogenins, which encode yolk proteins that facilitate nutrient transport to the germline. We find that reduced expression of vitellogenins in mononucleated intestinal cells leads to progeny with developmental delays and reduced fitness. Together, our results show that binucleation facilitates rapid upregulation of intestine-specific gene expression during development, independently of genome ploidy, underscoring the importance of spatial genome organization for polyploid cell function.

overall_design: 152 individual C. elegans animals expressing an intestinal specific TIR1 and containing an endogenous AID::knl-1 or AID::cdk-1 tag and grown in with auxin or under control conditions were collected as young adults (72 hours of development)

first: Lotte

last: van Rijnberk

first: Reinier

last: van der Palen

first: Erik

middle: S

last: Schild

first: Hendrik

last: Korswagen


title: Regulation of Chromatin Accessibility by hypoxia and HIF_3

city: Liverpool

first: Sonia

last: Rocha

address: Crown Street

country: United Kingdom

institute: University of Liverool

summary: Integrative analysis of RNA-seq with ATAC-seq to investigate changes in chromatin accessibility in repsonse to hypoxia.

overall_design: HeLa cells were grown at 21% oxygen, treated with control siRNA and exposed to 0 (control), 1 and 24 hs 1% oxygen (hypoxia). A549 cells were grown at 21% oxygen, treated with control siRNA and exposed to 0 (control) and 24 hs 1% oxygen (hypoxia). RNA-seq was performed to measure differential transcript expression.

first: Michael

last: Batie


title: A hotspot mutation targeting the R-RAS2 GTPase acts as a potent oncogenic driver in a wide spectrum of tumors

city: Salamanca

first: L. Francisco

last: Lorenzo-Martín

state: Castilla y León

address: Campus Miguel de Unamuno

institute: University of Salamanca (USAL) – Centro de Investigación del Cáncer (CIC)

summary:
A missense change in RRAS2 (Gln72-to-Leu), analogous to the Gln61-to-Leu mutation of RAS oncoproteins, has been identified as a hotspot mutation in cancer and Noonan syndrome. However, the relevance of this mutation for in vivo tumorigenesis remains unknown.
Here, we show that R-RAS2(Q72L) is required for optimal tumorigenic activity of human cancer cells.

overall_design: Genome-wide gene expression analyses of control and Rras2(Q72L) knockdown (sh#89) fibrosarcoma cells.

first: Isabel

last: Fernández-Pisonero

first: Laura

last: Clavaín

first: Javier

last: Robles-Valero

first: L

middle: F

first: Rubén

last: Caloto

first: Blanca

last: Nieto

first: Carmen

last: García-Macías

first: Clara

last: Oeste

first: Manuel

last: Sánchez-Martín

first: Antonio

last: Abad

first: Alejandro

last: Hortal

first: Dolores

last: Caballero

first: Marcos

last: González

first: Miguel

last: Vicente-Manzanares

first: Mercedes

last: Dosil

first: Balbino

last: Alarcón

first: Xosé

last: Bustelo


title: Deep learning approach identified a gene signature predictive of the severity of renal damage caused by chronic cadmium accumulation

city: SHANGHAI

first: XIAN

last: JIN

state: SHANGHAI

address: Cailun Road 720, 2-404

institute: ENNOVABIO

summary: Repetitive CdCl2 exposure resulted in cadmium accumulation and remarkable renal injuries in the animals. In the present study, we established an animal model to mimic chronic cadmium exposure-induced renal injury and performed whole transcriptome profiling studies. RNA-Seq data analysis showed that cadmium treatment induced dramatic gene expression changes in kidneys in a dose-dependent manner and higher severity of injury in ob/ob mice.

overall_design: Kidney mRNA profiles of CdCl2 treated ob/ob (ob) and normal C57BL/6 (b6j) mice

first: Feng

last: Xuefang

first: Jin

last: Xian


title: Next Generation Sequencing of TRAIL Resistant Gene Expression Profile of GBM Cells.

city: Istanbul

first: Tugba

last: Bagci-Onder

state: Sariyer

address: Koc University School of Medicine

country: Turkey

institute: Koc University

overall_design: Examination of TRAIL resistance on gene expression profile of GBM cell lines. 2 sets of each cell line were prepared for Resistant and Sensitive (parental/control) conditions.

first: Tuğba

first: Ahmet

last: Cingoz

first: Tunc

last: Morova