title: RNA-sequencing of control (GFP) or NDRG3-depleted pancreatic cancer cell lines

status: Public on Aug 31 2022

city: Daejeon

first: Han

last: Koo

address: 125 Gwahak-ro, Yuseong-gu, Daejeon, Korea

institute: KRIBB

summary: We perfomed RNA-sequencing for searching upstream regulators regulated by NDRG3 gene. RNA profiles of each cell line were used to figure out Differentially Expressed Genes (DEG) by NDRG3 depletion, we found NDRG3-related upstream regulators by applicating DEG to Ingenuity Pathway Analysis (IPA).

overall_design: mRNA profiles of pancreatic cancer cell lines depleted control (GFP) or NDRG3 gene

first: Young-Il

last: Yeom

first: Dong-chul

last: Lee

first: Kyung-chan

last: Park


title: Multipartite super-enhancers function in an orientation-dependent manner [RNA-Seq]

status: Public on Aug 18 2022

city: Oxford

first: Mira

middle: Tony

last: Kassouf

state: England

address: John Radcliffe Hospital, Headly way

department: Weatherall Institute of Molecular Medicine

country: United Kingdom

institute: University of Oxford

type: Expression profiling by high throughput sequencing

summary: Transcriptional enhancers regulate gene expression in a developmental-stage and cell-specific manner. They were originally defined as individual regulatory elements that activate expression regardless of distance and orientation to their cognate genes. Genome-wide studies have shown that the mammalian enhancer landscape is much more complex, with different classes of individual enhancers and clusters of enhancer-like elements combining in additive, synergistic and redundant manners, possibly acting as single, integrated regulatory elements. These so-called super-enhancers are largely defined as clusters of enhancer-like elements which recruit particularly high levels of Mediator and often drive high levels of expression of key lineage-specific genes. Here, we analysed 78 erythroid-specific super-enhancers and showed that, as units, they preferentially interact in a directional manner, to drive expression of their cognate genes. Using the well characterised a-globin super-enhancer, we show that inverting this entire structure severely downregulates a-globin expression and activates flanking genes 5’ of the super-enhancer. Our detailed genetic dissection of the a-globin locus clearly attributes the cluster’s functional directionality to its sequence orientation, demonstrating that, unlike regular enhancers, super-enhancers act in an orientation-dependent manner. Together, these findings identify a novel emergent property of super-enhancers and revise current models by which enhancers are thought to contact and activate their cognate genes.

overall_design: An inversion spanning the alpha globin superenhancer and including both the Nprl3 and Mpg genes was genetically engineered in mouse ESCs and a mouse model was created by blastocyst injection of these mESCs. Primary erythroid cells derived from APH-treated spleens of these mice (INVEN) as well as WT mice were subjected to various analyses (ATAC-Seq, ChIP-Seq, RNA-Seq, Capture C) to examine the effect this inversion has on the expression of the erythroid -specific alpha globin genes. Embryonic blood (E10.5) was also analysed to differentiate the effect of the inversion on primitive (embryonic) versus dfinitive (adult) erythropoiesis. Moues ESCs were also generated; these harbour additional mutations that allow the effect of the inversion to be untangled from other confoudning factors (such as the boundary elements HS3839 or the repoistioned Mpg gene). These mESCs were subjected to an in vitro erythroid differentiation protocol (doi: 10.1371/journal.pone.0261950) and purified CD71+ erythroid cells derived from differentiated Wildtype (WT) or mutant mouse Embryonic Stem (mES) cells were analysed similarly to the primary spleen erythroid cells.


title: Multipartite super-enhancers function in an orientation-dependent manner [ChIP-Seq]

type: Genome binding/occupancy profiling by high throughput sequencing

first: Matthew

last: Gosden

first: Helena

last: Francis

first: Caroline

last: Harrold


title: RNA-seq analysis of MDA-MB-231_2b cells treated with control or TRIM37 specific antisense oligonucleotides

city: Davis

first: Sanchita

last: Bhatnagar

state: CA

address: 1275 Health Sciences, Tupper Hall 3134

department: Medical Microbiology and Immunology

country: USA

institute: University of California Davis

summary: TRIM37 is histon H2A ubiquitinase and important oncogene for breast cancer. To test what is the gene expression network regulated by TRIM37, we silenced its ixpression in MDA-MB-231_2b Triple Negative Breast Cancer (TNBC) cell line.

overall_design: RNA-seq on 6 samples: 3 control ASO, and 3 TRIM37 ASO

first: Piotr

last: Przanowski


title: Multipartite super-enhancers function in an orientation-dependent manner [Capture-C]


title: Multipartite super-enhancers function in an orientation-dependent manner

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

overall_design: Refer to individual Series


title: Paternal obesity alters the sperm epigenome and and is associated with changes in the placental transcriptome and its cellular composition [ChIP-seq]

status: Public on Sep 09 2022

city: Sainte-Anne-de-Bellevue

first: Anne-Sophie

last: Pépin

state: QC

address: 21111 Lakeshore road

department: Department of Pharmacology and Therapeutics

country: Canada

institute: McGill University

summary: Growing evidence point towards a strong contribution of paternal factors in placental health with implications in adult-onset complex disease risk. We have recently demonstrated that paternal diet-induced obesity alters sperm histone methylation and is associated with metabolic disturbances in the next generation. Diet-sensitive epigenetic regions in sperm were found at genes involved in trophectoderm and placental development, and corresponded to epigenetic and gene expression profiles in these tissues. We sought to investigate whether paternal diet-induced obesity before conception can alter the placental transcriptome, and whether differential gene expression corresponds with sperm obesity-associated epigenetic signatures. C57BL6/J males were fed either a control or high-fat diet for 10 weeks beginning at 6 weeks of age. They were then bred to control-fed C57BL6/J females to induce pregnancies and E14.5 placentas were collected. RNA-sequencing was performed (n=4 per group per sex) to detect sex-specific transcriptional changes associated with paternal diet. At necropsy, sperm was collected for chromatin immunoprecipitation followed by sequencing (ChIP-seq; n=3 per group) targeting histone H3 lysine 4 tri-methylation (H3K4me3) to detect obesity-induced changes in H3K4me3 enrichment. There were sex-specific differentially expressed genes in placentas with some overlapping with promoters showing obesity-associated sperm epimutations. A deconvolution analysis using single-cell RNA-seq data from mouse E14.5 placenta (Han et al., Cell, 2018) revealed significant differences in trophoblast subtype proportions in placentas derived from HFD sires. This study highlights a previously underappreciated role of the placenta at the origin of paternally-induced metabolic disturbances in offspring.

overall_design: Examination of sperm H3K4me3 in control or high-fat fed sires (n=3 per group), examination of E14.5 placenta total RNA derived from control or high-fat fed sires (n=4 per sex per group)

last: Pepin

first: Sarah

last: Kimmins


title: Transcriptional study of the temporal evolution after severe traumatic spinal cord injury (SCI) and neural progenitor cells (NPCs)-transplantation intervention.

status: Public on Aug 16 2022

city: Valencia

first: Victoria

last: Moreno Manzano

state: Valencia

address: No. 3 Eduardo Primo Yúfera street

country: Spain

institute: Centro de Investigación Príncipe Felipe

summary:
The present study is intended to disclose the transcriptional profile of the temporal evolution through the different stages after SCI as well as the molecular processes underlying the NPC based-therapy in order to  transcriptionally characterize its therapeutical mechanism.
This transcriptional profile analysis of total RNA samples from  spinal cord homogenates of adult rats (Sprague Dawley) provides tissular (tought not cell-type specific) information of the critical time points after the injury from 1 to 8 weeks (acute, sub-acute and early-chronic and late-chronic stages) which conferred a wide temporal coverage making it ideal for studying the temporal dynamics of SCI. Furthermore, we have evaluated the impact of intramedullary acute or subacute transplantation of NPCs over the transcriptional regulation of the spinal cord tissue in order to define the functional outcomes of the NPCs therapy.

overall_design: For a better understanding of the molecular mechanisms involved in temporal evolution ocurring after severe traumatic spinal cord injury (SCI), a differential transcriptional profile was evaluated from acute to chronic stages. Severe traumatic contusion was induced by applying 250 kdyn at the T8 thoracic level in adult Sprague Dawley female rats . Spinal cord tissue, including the epicentre of the injury was collected at different time points representing the acute (1 week; T1), subacute (2 weeks; T2), early chronic (4 weeks; T4) and late chronic (8 weeks; T8) phases to perform the transcriptional profiling analysis using Rat Gene Expression microarray platform (Agilent-014879 Whole Rat Genome Microarray 4x44K G4131F, Agilent). Furthermore to evaluate the transcriptional impact of neural progenitor cells (NPC) transplantation in the injured spinal cord, we performed acute and subacute transplantation (right after or 1 week after injury, respectively) to injured rats . Each experimental group consisted in 4 biological replicates, sham uninjured animals (t0) served as a control for injured rats (t1, t2, t4, t8) and 1-week (t1) and 2-weeks (t2) injured animals served as time-matching control of acutely (u1t1) and subacutely (u2t2) transplanted animals, respectively.

first: Beatriz

last: Martínez-Rojas

first: Esther

last: Giraldo-Reboloso

first: Rubén

last: Grillo-Risco

first: Marta

last: Hidalgo-García

first: Francisco

last: García-García

last: Moreno-Manzano


title: Different characteristics of different scar fibroblasts

status: Public on Aug 06 2022

first: Jennifer

middle: Hyunjong

last: Shin

state: Yuseong-gu

address: 291 Daehak-ro

country: South Korea

institute: KAIST

summary: The goal of this study is to compare the different characteristics of fibroblasts from normal, hypertrophic and keloid scar through RNA-seq.

overall_design: Fibroblasts from normal, hypertrophic and keloid scars (n=3) were isolated and total RNA extracted from each cells were used for RNA-seq.

first: Minwoo

last: Kang

first: Ung

middle: H

last: Ko

first: Eun

middle: J

last: Oh

first: Hyun

middle: M

last: Kim

first: Ho

middle: Y

last: Chung


title: Landscape of single-cell transcriptomics of human placent in pathophysiological pregnancies

status: Public on Oct 12 2022

city: New York

first: Nanber

last: Zhong

address: 1050 Forest Hill Road

institute: New York State Institute for Basic Research in Developmental Disabilities

summary: Pathophysiological pregnancies amy result from gene-environmental interactions. In this study, investigation of the placental cell development in both chorionic villi and decidua of variant forms of pathophysiological pregnancies was pursued with a single cell RNA sequencing. Transcriptomic gene differential expression in variant placental cell clusters from chorionic villi and decidua of pregnancies, unique marker genes in trophoblasts that may lead to identification of biomarker for placental cell development states and cell-specific pathway activities were identified. In addition, gene expression profiles of ACE2 and TMPRSS2, the two key proteins involved in SARS-CoV-2 infection, in variant placental cell clusters was documented, which may facilitate exploration of vertical transmission of COVID-19 from mother to fetus.

overall_design:
SAMPLES A,B,C,D for the study of EPL.
SAMPLES N1-5, E1-5 for study of endometriosis.
10x Genomics Chromium Single Cell sequencing is applied to study the cell-specific transcriptomics for EPL.

first: Nanbert

first: Xiaoling

last: Guo

first: Leslie

last: Myatt

first: James

last: Roberts

first: Yuhua

last: Zheng

first: Jing

last: Pan

first: Chenglai

last: Xia

first: Huadong

last: Zhou

first: Haiying

last: Chen

first: Weina

last: Ju


title: Multipartite super-enhancers function in an orientation-dependent manner [ATAC-Seq]

first: Mira

last: Kassouf

first: Matthew

last: Gosden

first: Helena

last: Francis


title: Methylation data from stored peripheral blood leukocytes from African American participants in the GENOA study with Infinium HumanMethylation450 BeadChip

status: Public on Aug 04 2022

city: Ann Arbor

first: Lulu

last: Shang

state: Michigan

address: 1415 Washington Heights

department: Biostatistics

institute: University of Michigan

type: Methylation profiling by genome tiling array

summary: African-American individuals of the GENOA study

overall_design: The Genetic Epidemiology Network of Arteriopathy (GENOA) study is a community-based study of hypertensive sibships that was designed to investigate the genetics of hypertension and target organ damage. The study includes African Americans (AA) from Jackson. In the initial phase of GENOA (Phase I: 1996-2001), all members of sibships containing at least 2 individuals with essential hypertension clinically diagnosed before age 60 were invited to participate, including both hypertensive and normotensive siblings. The exclusion criteria for GENOA included secondary hypertension, alcoholism or drug abuse, pregnancy, insulin-dependent diabetes mellitus, or active malignancy. Eighty percent of AA (N=1,482) from the initial study population returned for the second examination (Phase II: 2001-2005). Demographic information, medical history, clinical characteristics, lifestyle factors, and blood samples were collected in each phase. Written informed consent was obtained from all subjects and approval was granted by participating institutional review boards (University of Michigan, University of Mississippi Medical Center, and Mayo Clinic). Genomic DNA was extracted from stored peripheral blood leukocytes that was collected at Phase-1 (N=422) using AutoGen FlexStar (AutoGen, Holliston, MA). Bisulfite conversion was performed with the EZ DNA Methylation Kit (Zymo Research, Irvine, CA), and methylation was assessed using the Illumina Infinium HumanMethylation450 BeadChip. In total 418 samples remained after quality control and filtering.

first: Sharon

middle: L

last: Kardia

first: Jennifer

middle: A

last: Smith


title: The developmental origins of sex-biased expression in cardiac development

status: Public on Jul 29 2022

city: Camden

first: Jozef

last: Madzo

state: NJ

address: 403 Haddon Avenue

department: Bioinformatics

institute: Coriell Institute

summary: The sexually dimorphic gene expression profiles are established in early embryogenesis and epigenetically perpetuated at every stage of cardiac development.

overall_design: Examination of chromatin modifications in male and female mouse ES cells.

first: Daniel

middle: F

last: Deegan

first: Reza

last: Karbalaei

first: Rob

last: Kulathinal

first: Nora

last: Engel


title: Il-1-dependent enteric gliosis guides intestinal inflammation and dysmotility and modulates macrophage function

status: Public on Jul 11 2022

city: Bonn

first: Reiner

last: Schneider

address: Venusberg-Campus 1

department: Surgery Department

country: Germany

institute: University Clinics Bonn

summary: IL-1 signaling in enteric glia initiates acute intestinal inflamation and modulates postoperative motility disturbances by triggering a reactive glial phenotype named enteric gliosis. Enteric glia modulate macrophage function and activity in this reactive state by releasing a unique panel of chemokines and cytokines. An enteric glia-selective knockout of this pathway ameliorates acute postoperative inflammation and prevents postoperative ileus.

overall_design: Examination of enteric glial cells after IL-1b treatment in vitro and the effect of conditioned medium on BMDMs from IL1R1-KO animals

first: Patrick

last: Leven

first: Sven

last: Wehner


title: Effect of hypertonicity on Gene expression and contribution of Nfat5

status: Public on Aug 17 2022

city: Halle

first: Bayram

last: Edemir

address: Ernst-Grube-Str. 20

institute: University Hospital Halle (Saale)

summary: We analyzed the  gene expression in WT and Nfat5 deficient kidneys

overall_design: Renal cortex and renal inner medulla from WT and NFAT5-KO mice were used for gene expression analysis.


title: Methylation data from stored peripheral blood leukocytes from African American participants in the GENOA study with Infinium HumanMethylationEPIC BeadChip

overall_design: The Genetic Epidemiology Network of Arteriopathy (GENOA) study is a community-based study of hypertensive sibships that was designed to investigate the genetics of hypertension and target organ damage. The study includes African Americans (AA) from Jackson. In the initial phase of GENOA (Phase I: 1996-2001), all members of sibships containing at least 2 individuals with essential hypertension clinically diagnosed before age 60 were invited to participate, including both hypertensive and normotensive siblings. The exclusion criteria for GENOA included secondary hypertension, alcoholism or drug abuse, pregnancy, insulin-dependent diabetes mellitus, or active malignancy. Eighty percent of AA (N=1,482) from the initial study population returned for the second examination (Phase II: 2001-2005). Demographic information, medical history, clinical characteristics, lifestyle factors, and blood samples were collected in each phase. Written informed consent was obtained from all subjects and approval was granted by participating institutional review boards (University of Michigan, University of Mississippi Medical Center, and Mayo Clinic). Genomic DNA was extracted from stored peripheral blood leukocytes that was collected at Phase 1 (N=1106) or Phase 2 (N=304) using AutoGen FlexStar (AutoGen, Holliston, MA). Bisulfite conversion was performed with the EZ DNA Methylation Kit (Zymo Research, Irvine, CA), and methylation was assessed using the Illumina Infinium HumanMethylationEPIC BeadChip. In total 1,394 samples remained after quality control and filtering.


title: Methylation data from stored peripheral blood leukocytes from African American participants in the GENOA study


title: B11 cells created by replacing the NRAS-G12D mutation in THP1 cells with a dox-inducible NRAS-G12V mutation

status: Public on Jul 25 2022

city: Minneapolis

first: Zohar

last: Sachs

state: MN

address: 420 Delaware Street SE MMC 480

department: Department of Medicine, Division of Hematology, Oncology and Transplantation

institute: University of Minnesota Twin Cities

summary: To study the role of NRAS mutations in cell proliferation and self-renewal in acute myeloid leukemia (AML), the human AML cell line, THP1,  was modified to replace its naturally occurring heterozygous NRAS-G12D mutation with a doxycycline(dox)-inducible heterozygous NRAS-G12V mutation. The endogenous copies of the NRAS-G12D allele were deleted using CRISPR/Cas9 after a dox-inducible, CRISPR resistant, NRAS-G12V transgene was introduced into the THP1 cell line.  The resulting cell line was named B11.  RNA-seq data confirmed that endogenous NRAS G12D was successfully replaced by dox-inducible exogenous NRAS G12V in the B11 cell line.  As expected, depletion of dox induced G1 cell cycle arrest.  Interestingly, the B11 cells experienced ten-times higher expression of NRAS induced G2/S-phase cell cycle arrest.  Forty-nine genes were identified as signaling responsible genes associated with high expression of NRAS.

overall_design:
Using lenti-vector system, a CRISPR resistant dox-inducible NRAS was introduced to human THP1 cells, which also contained a G12V mutation and GFP.  Cells were treated with doxycycline and GFP positive were selected.  The selected cells were then transduced with a vector containing Cas9, Puromycin resistant gene, and dual gRNAs specific to upstrem and downstream of the start codon in NRAS locus.  Subclones were selected using puromycin.
One of the subclones, B11, was grown out and separated into 9 plates.  THP1 cells were also separated into 9 plates.  3 of the THP1 plates and 3 of the B11 plates were treated with dox at 1 ug/ml, 3 of the THP1 plates and 3 of the B11 plates were treated with dox at 10 ug/ml, and 3 of the THP1 plates and 3 of the B11 plates were left untreated.  After 5 days, RNA was collected and submitted for RNA-sequencing.

first: S

middle: K

last: Rathe

first: M

last: Kurata

first: D

last: Largaespada

first: Z


title: Pro-Inflammatory Cytokines Mediate the Epithelial-to-Mesenchymal-Like Transition of Pediatric Posterior Fossa Ependymoma (single-nucleus RNA-seq)

status: Public on Jun 24 2022

city: Philadelphia

first: Pablo

last: Camara

state: PA

address: 3700 Hamilton Walk

department: Genetics

institute: University of Pennsylvania

summary: Pediatric ependymoma is a devastating brain cancer marked by its relapsing pattern and lack of effective chemotherapies. This shortage of treatments is due to limited knowledge about ependymoma tumorigenic mechanisms. By means of single-nucleus chromatin accessibility and gene expression profiling of posterior fossa primary tumors and distal metastases, we reveal key transcription factors and enhancers associated with the differentiation of ependymoma tumor cells into tumor-derived cell lineages and their transition into a mesenchymal-like state. We identify NFkB, AP-1, and MYC as mediators of this transition, and show that the gene expression profiles of tumor cells and infiltrating microglia are consistent with abundant pro-inflammatory signaling between these populations. In line with these results, both TGF-b1 and TNF-a induce the expression of mesenchymal genes on a patient-derived cell model, and TGF-b1 leads to an invasive phenotype. Altogether, these data suggest that tumor gliosis induced by inflammatory cytokines and oxidative stress underlies the mesenchymal phenotype of posterior fossa ependymoma.

overall_design: Single-nucleus RNA-seq data was generated from 5 primary, 1 progressive, and 3 metastatic pediatric posterior fossa ependymal tumors using archived flash-frozen tissue.

first: Rachael

middle: G

last: Aubin

first: Emma

middle: C

last: Troisi

first: Javier

last: Montelongo

first: Adam

middle: N

last: Alghalith

first: MacLean

middle: P

last: Nasrallah

first: Mariarita

last: Santi


title: Pro-Inflammatory Cytokines Mediate the Epithelial-to-Mesenchymal-Like Transition of Pediatric Posterior Fossa Ependymoma

summary:
Pediatric ependymoma is a devastating brain cancer marked by its relapsing pattern and lack of effective chemotherapies. This shortage of treatments is due to limited knowledge about ependymoma tumorigenic mechanisms. By means of single-nucleus chromatin accessibility and gene expression profiling of posterior fossa primary tumors and distal metastases, we reveal key transcription factors and enhancers associated with the differentiation of ependymoma tumor cells into tumor-derived cell lineages and their transition into a mesenchymal-like state. We identify NFkB, AP-1, and MYC as mediators of this transition, and show that the gene expression profiles of tumor cells and infiltrating microglia are consistent with abundant pro-inflammatory signaling between these populations. In line with these results, both TGF-b1 and TNF-a induce the expression of mesenchymal genes on a patient-derived cell model, and TGF-b1 leads to an invasive phenotype. Altogether, these data suggest that tumor gliosis induced by inflammatory cytokines and oxidative stress underlies the mesenchymal phenotype of posterior fossa ependymoma.
This SuperSeries is composed of the SubSeries listed below.

overall_design:
Single-nucleus RNA- and ATAC-seq data was generated from primary and metastatic pediatric posterior fossa ependymal tumors using archived flash-frozen tissue.

Refer to individual Series


title: Hypoxia-mediated stabilization of HIF1A in prostatic intraepithelial neoplasia promotes cell plasticity and malignant progression

status: Public on Jun 07 2022

first: Gilles

last: Laverny

address: 1 rue Laurent fries

country: France

institute: IGBMC

summary: Prostate cancer (PCa) is a leading cause of cancer-related deaths. The slow evolution of prostatic precancerous lesions to malignant tumors provides a broad time-frame for strategies targeting disease emergence. To characterize prostatic intraepithelial neoplasia (PIN) progression, we conducted longitudinal studies on prostates of genetically-engineered Pten(i)pe- /- mice. We discovered that early PINs are hypoxic and that hypoxia-inducible factor 1 alpha (HIF1A) signaling is activated in luminal cells during disease progression. Luminal HIF1A enhances glucose metabolism and promotes a PIN-derived secretome that increases the recruitment of myeloid-derived suppressor cells, thus dampening immune surveillance. Moreover, pharmacological inhibition of HIF1A induces apoptosis in early PIN lesions, and slows the proliferation of late ones. Therefore, our study identifies HIF1A as a target for PCa prevention. Importantly, we also demonstrate that HIF1A signaling correlates with the emergence of prostatic luminal cells expressing TGM2, the expression of which predicts early relapse after primary intervention in PCa patients.

overall_design: Pool of 3 dissociated prostates were analyzed by single cell analyis using the chromium 10X (10X Genomics)

first: Mohamed

last: Abu el Maaty

first: Daniel

last: Metzger

first: Celine

last: Keime


title: Telomerase-Dependent Ageing In The Zebrafish Brain

status: Public on Jun 09 2022

city: Sheffield

first: Catarina

middle: Martins

last: Henriques

address: Bateson Centre, Firth Court, The University of Sheffield, Western Bank

department: Oncology & Metabolism

institute: The University of Sheffield

summary:
Decreased telomerase expression, telomere shortening, senescence-associated markers and inflammation have all been independently observed in the ageing brain and associated with disease. However, causality between limited telomerase expression and brain senescence and neuro-inflammation in the natural ageing setting is yet to be established. Here, we address these questions using the zebrafish as an ageing model which, akin to humans, displays premature ageing and death in the absence of telomerase and where telomere shortening is a driver of cellular senescence.
Our work shows for the first time that telomerase deficiency (tert-/-) accelerates key hallmarks of ageing identified in the Wild Type (WT) zebrafish brain at transcriptional, cellular, tissue and functional levels. We show that Tert-dependent transcriptomic changes associated with dysregulation of gene expression, stress response and dysregulation of immune genes are accompanied by accelerated accumulation of senescence-associated markers and inflammation in the aged brain. Importantly, In vivo, these changes correlate with increased blood-brain barrier permeability and altered cognitive behaviour. Of note, telomerase-dependent accumulation of senescence-associated markers in the brain occurs not only in the expected proliferative areas but also in non-proliferative ones, where it is unlikely due to telomere-dependent replicative exhaustion, suggesting that non-canonical roles of telomerase may be involved.
Together, our work suggests that telomerase has a protective role in the zebrafish brain against the accumulation of senescence and neuro-inflammation and is required for blood brain barrier integrity.

overall_design:
RNA-Sequencing of whole brain tissues, throughout the lifespan of WT and telomerase-deficient (tert-/-) fish. The data 4 age-groups of WT (2, 9, 22 and >30 months), corresponding to young, adult, median lifespan and old; and 3 age-groups of telomerase-deficient fish (2, 9, and 22 months), which correspond to young, medium lifespan and old. Each group has a sample size of 3 animals.
To identify signatures of ageing, WT brain samples were subjected separately to DESEq2 analysis, comparing the time points 9, 22 and >30 months with the time-point of 2 months. Then, tert-/- samples at 2, 9 and 22 months were compared with WT at 2 months, to identify telomerase-dependent ageing processes.

first: Raquel

middle: R

last: Martins

first: Savandara

last: Besse

first: Pam

middle: S

last: Ellis

first: Naomi

last: Hartopp

first: Nadiyah

last: Mughal

first: Owain

last: Evans

first: Mina

last: Wahib

first: Zerkif

last: Yazigan

first: Heather

last: Mortiboys

first: Michael

last: Rera


title: Ribosome impairment regulates intestinal stem cell identity via ZAKɑ activation

status: Public on Jun 08 2022

city: Amsterdam

first: William

last: Faller

address: Plesmanlaan 121

department: Oncogenomics

country: Netherlands

institute: Netherlands Cancer Institute

summary: We show that ribosome collisions, induced by Rptor deletion or aminoacid starvation, causes a ZAK alpha mediated identity switch in intestinal stem cells by activating a more fetal-like stem cell signature. This switch also causes changes in metabolic profiles of the cells, which is heavily regulated by RNA translation efficiency of different group of metabolic genes. These observations originate from performing in vitro Riboseq and RNAseq in mouse intestinal organoids, and in vivo Riboseq in Lgr5 expressing intestinal stem cells from WT and Rptor.fl/fl mice.

overall_design: In vitro Ribo-seq and RNAseq in WT-stem cell enriched, WT-stem cell depleted and Rptor fl/fl mouse intestinal organoids. In vitro RNAseq in WT mouse intestinal organoids. In vivo Ribo-seq in Lgr5 expressing intestinal stem cells from WT and Rptor fl/fl mouse.

first: Ferhat

last: Alkan

first: Joana

last: Silva


title: Central Lung Gene Expression Associates with Myofibroblast Activity in Idiopathic Pulmonary Fibrosis

status: Public on May 31 2022

city: Charlottesville

first: Yong

last: Huang

state: VA

address: 1340 Jefferson Park Ave

department: Medicine

institute: University of Viginia

summary: Non-fibrotic central regions of IPF lungs demonstrate massive dysregulated gene expression involving enhanced T-cell differentiation, pulmonary fibrosis idiopathic, and wound healing pathways. Enhancement of myofibroblast features without apparent extra-cellular matrix deposition in the central non-fibrotic region of IPF lungs renders a window for cell-based molecular targeting therapy.

overall_design: IPF lungs were grossly examined within 6-12 hours from the time of explant, and multiple 1cm3 biopsies were obtained from macroscopically non-fibrotic central (IPF.C) and fibrotic peripheral (IPF.P) areas. Non-IPF donor lung biopsies of central (CTR.C) and peripheral (CTR.P) were similarly obtained. H&E-stained biopsies were reviewed and confirmed by two clinical pulmonary pathologists (Jefree Schulte, Aliya Husain). Representative central and peripheral samples from IPF patients and non-IPF donors were selected and underwent pathologic classification and scoring to quantify fibrosis, fibroblastic foci, honeycombing, inflammation, and pulmonary hypertension.

first: Robert

last: Guzy

first: Shwu-Fan

last: Ma

first: Imre

last: Noth


title: Transcriptional dysregulation of BAHCC1 in melanoma

status: Public on Jun 03 2022

first: Pietro

last: Berico

state: NY

address: 550 1st Ave

department: Pathology

institute: NYU Langone Health

overall_design: Refer to individual Series


title: Genomic alterations drive brain metastases formation in colorectal cancer: The role of IRS2

status: Public on May 16 2022

city: Tel Aviv

first: Dor

last: Simkin

address: 6 Weizmann st.

country: Israel

institute: Department of Oncology, Tel Aviv Sourasky Medical Center, Tel Aviv, Israel

summary: Colorectal cancer (CRC) is currently the fourth leading etiology of brain metastasis (BM). Yet, mechanisms supporting the formation of CRC BM are mostly unknown. In order to identify drivers that lead to tropism and adaptation of CRC cells to the brain environment, we analyzed an extensive genomic database, consisting of over 36,000 human CRC primary and metastasis samples. Several genomic alterations specific for BM were noted, among them increased prevalence of insulin receptor substrate 2 (IRS2) amplification, observed in 7.6% of BM, compared to only 2.9% of primary tumors or other metastatic sites (p<7E-05). This observation was validated by Immunohistochemistry studies of human clinical samples, showing increased expression of IRS2 protein in BM. IRS2 is a cytoplasmic adaptor mediating effects of insulin and IGF-1 receptors and is involved in more aggressive behavior of different cancer types. In order to study the ability of IRS2 to promote growth of CRC cells under brain microenvironment conditions, we employed an in vitro system consisting of cultured human astrocytes or their conditioned media. Indeed, IRS2-overexpressed CRC cells survived better and formed larger 3D spheres when grown in brain-mimicking conditions, while IRS2-silenced CRC cells showed a mirror image. Similarly, In vivo studies, using intracranial CRC BM mouse model, demonstrated that IRS2-overexpressed cells generated larger brain lesions, while silencing IRS2 dramatically decreased tumor outgrowth and extended survival. Transcriptomic analysis revealed enrichment of oxidative phosphorylation (OXPHOS) and Wnt/β-catenin pathways by IRS2. Indeed, IRS2-expressing cells showed increased mitochondrial activity and glycolysis-independent viability. Furthermore, IRS2-expressing cells had increased β-catenin transcriptional activity, and either β-catenin inhibition or IRS2 inhibition in IRS2-expressing cells decreased their viability, β-catenin transcriptional activity, and mitochondrial activity. These data suggest involvement of IRS2 in modulating OXPHOS through β-catenin. Exploiting this mechanism as a potential vulnerability allowed us to develop novel treatment strategies against CRC BM. NT219 is a novel IRS1/2 inhibitor already being tested in clinical trials. Treatment of mice harboring CRC BM with NT219 and 5-flourouracil reduced tumor growth and prolonged mice survival. These data reveal, for the first time, the unique genomic profile of CRC BM and imply the IRS2 role in promoting CRC BM. These effects may be mediated, at least in part, by modulation of the β-catenin and OXPHOS pathway. These findings may pave the way for clinical trials evaluation this novel IRS2-based strategy for the treatment of CRC BM.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for SW403 colon adenocarcinoma cell line grown in DMEM in two conditions: normal IRS2 expression versus IRS2 knockdown.

first: Inbal

last: Greenberg

first: Anat

last: Klein-Goldberg

first: Ethan

last: Sokol

first: Moshe

last: Benhamou

first: Michal

last: Raz

first: Rachel

last: Grossman

first: Eilam

last: Yeini

first: Paula

last: Ofek

first: Ronit

last: Satchi-Fainaro

first: Bertrand

last: Liang

first: Hadas

last: Reuveni

first: Tami

last: Rubinek

first: Ido

last: Wolf


title: Determination of human DNA replication origin position and efficiency reveals principles of initiation zone organisation

status: Public on Jun 15 2022

city: Cambridge

first: Guillaume

last: Guilbaud

address: Francis Crick Avenue

department: PNAC

institute: Medical Research Council - LMB

summary: We describe a modification of initiation site sequencing (ini-seq, Langley et al., NAR 2016) in which density gradient centrifugation separates ‘heavy’ replicated DNA containing BrdU from DNA containing only ‘light’ dTTP before sequencing. This approach allows us to assign a replication initiation efficiency score to each of the 23905 origins identified. We also performed Short Nascent Strand sequencing (SNS-seq), an established method to map replication origins, in the same cell line as in Ini-seq2 for comparison.

overall_design: Massive sequecing of human replication origin isolated from EJ30 cell line by either Ini-seq2 or SNS-seq method

first: Pierre

last: Murat

first: Helen

last: Wilkins

first: Leticia

last: Lerner

first: Julian

middle: E

last: Sale

first: Torsten

last: Krude


title: Single-cell analysis of CD45+ cells from duodenal RCDII tissue

status: Public on Apr 30 2022

city: Leiden

first: Tessa

last: Dieckman

address: Albinusdreef 2

department: Immunology

institute: LUMC

summary: Refractory Celiac Disease type II (RCDII) is a rare lymphoma in the small intestine characterized by a clonally expanded intra-epithelial iCD3+sCD3-CD7+CD56- aberrant cell population. However, RCDII pathogenesis is ill-defined. Here, we aimed at single-cell characterization of the innate and adaptive immune system in RCDII. Paired small intestinal and blood samples from 12 RCDII patients and 6 healthy controls were assessed by single-cell mass cytometry with a 39 cell surface marker antibody panel, designed to capture heterogeneity of the innate and adaptive immune system. A second single-cell mass cytometry panel which included transcription factors and immune checkpoints was used for analysis of paired samples from 5 RCDII patients. Single-cell RNA sequencing analysis was performed on duodenal samples from 2 RCDII patients. Finally, we developed a 40-marker imaging mass cytometry antibody panel to evaluate cell-cell interactions in duodenal biopsies of RCDII patients. We provide evidence for inter- and intra-tumoral cell heterogeneity within the duodenal and peripheral aberrant cell population present in RCDII. We observed that part of the aberrant cell population proliferated and observed co-localization of aberrant cells with CD163+ APCs in situ. Additionally, we observed phenotypic discrepancy between peripheral and duodenal aberrant cells. Novel high-dimensional technologies reveal substantial inter- and intra-tumoral heterogeneity in the aberrant cell population in RCDII. This may underlie variability in refractory disease status between patients and responsiveness to therapy, pointing to the need for personalized therapy in RCDII based on patient-specific immune profiles.

overall_design: Isolation of live single intra-epithelial and lamina propria CD45+ lymphoyctes from duodenal tissue derived from 2 RCDII patients, which were processed on two different days on the Chromium 3' single cell platform (10X Genomics) and sequenced simultaneously on the NovaSeq 6000 system (Illumina).

first: Frits

last: Koning

first: Susan

last: Kloet


title: Transcriptional dysregulation of BAHCC1 in melanoma [CUT&Tag]

first: Pietro

last: Berico

summary: Transcriptional addiction of cancer cells for Super-Enhancer (SE) associated genes has been proven through an extensive use of transcriptional inhibitors against specific co-activators toward which SE are esponentially dependent compared to canonical enhancers. However, each cancer type displays different sensitivity against this new class of drugs whose efficacy may vary according to their transcriptomic profile including the expression of enzymatic proteins able to confer intrinsic tolerance or resistance. Moreover, SE has been observed as well in healthy cells which point a realistic concern toward the specificity of transcriptional inhibitors and their side effects on healthy tissues. Therefore, we looked at SE-associated genes specific for cancer using melanoma as a model. Here, we identified BAHCC1 whose activity is higher in melanoma compare to other cancer types and normal tissues. BAHCC1 is a protein involved both in gene expression and genomic stability. To explore BAHCC1 role in gene expression, we performed functional studies in melanoma cells in basal conditions or upon BAHCC1 silencing. Strikingly, BAHCC1 regulates an E2F-dependent subset of genes together with SWI/SNF chromatin remodelling complex.

overall_design: Cleavage Under Targets and Tagmentation (CUT&Tag) of BAHCC1 followed by deep sequencing

first: Frederic

last: Coin

first: Irwin

last: Davidson


title: Multi-omics analysis of the cervical epithelial integrity of women using depot medroxyprogesterone acetate

status: Public on Apr 29 2022

city: Stockholm

first: Kristina

last: Broliden

state: Solna

address: Visionsgatan 4, J7:20

department: Dept. of Medicine Solna

country: Sweden

institute: Karolinska Institutet

summary: Depot medroxyprogesterone acetate (DMPA) is an injectable hormonal contraceptive used by millions of women worldwide. However, experimental studies have associated DMPA use with genital epithelial barrier disruption and mucosal influx of human immunodeficiency virus (HIV) target cells. We explored the underlying molecular mechanisms of these findings. Ectocervical biopsies and cervicovaginal lavage (CVL) specimens were collected from HIV-seronegative Kenyan sex workers using DMPA (n=32) or regularly cycling controls (n=64). Tissue samples were assessed by RNA-sequencing and quantitative imaging analysis, whereas protein levels were measured in CVL samples. The results suggested a DMPA-associated upregulation of genes involved in immune regulation, including genes associated with cytokine-mediated signaling and neutrophil-mediated immunity. A transcription factor analysis further revealed DMPA-associated upregulation of RELA and NFKB1 which are involved in several immune activation pathways. Several genes significantly downregulated in the DMPA versus the control group were involved in epithelial structure and function, including genes encoding keratins, small proline-rich proteins, and cell-cell adhesion proteins. Pathway analyses indicated DMPA use was associated with immune activation and suppression of epithelium development, including keratinization and cornification processes. The cervicovaginal microbiome composition (Lactobacillus dominant and non-Lactobacillus dominant) had no overall interactional impact on the DMPA associated tissue gene expression. Imaging analysis verified that DMPA use was associated with an impaired epithelial layer as illustrated by staining for the selected epithelial junction proteins E-cadherin, desmoglein-1 and claudin-1. Additional staining for CD4+ cells revealed a more superficial location of these cells in the ectocervical epithelium of DMPA users versus controls. Altered protein levels of SERPINB1 and ITIH2 were further observed in the DMPA group. Identification of specific impaired epithelial barrier structures at the gene expression level, which were verified at the functional level by tissue imaging analysis, illustrates mechanisms by which DMPA adversely may affect the integrity of the genital mucosa.

overall_design:
In this study we compared the gene expression between a group of women using DMPA (n=32) and a group of women not using any type of hormonal contraceptives (n=64), acting as control group. RNA used for sequencing was extracted from the ectocervical biopsies.
NOTE FROM SUBMITTER: The raw sequencing data cannot be held in a public repository due to the sensitive nature of such personal data. Request for access to raw data can be made to the Data Access Committee, and access will be granted if the request meets the requirements of the data policy.


title: Transcriptional dysregulation of BAHCC1 in melanoma [RNA-seq]

overall_design: Total RNA sequencing


title: Patient-derived brain organoids uncover reversible cellular abnormalities in Pitt-Hopkins Syndrome IV

status: Public on Mar 02 2022

city: Campinas

first: Fabio

last: Papes

state: SP

address: Rua Carl Von Linaeus

country: Brazil

institute: University of Campinas

summary: We evaluate cellular phenotypic abnormalities and alterations in molecular pathways due to mutations in the gene TCF4 in cells derived from Pitt-Hopkins patients, in comparison with respective controls (parents).

overall_design: Comparison between RNA-Seq libraries of neurons derived from Pitt-Hopkins Syndrome patients and respective parents of matching sex. Libraries for each individual include three biological replicates.

first: Antonio

last: Camargo


title: Patient-derived brain organoids uncover reversible cellular abnormalities in Pitt-Hopkins Syndrome

overall_design: Comparison between RNA-Seq libraries of neural progenitor cells and neurons derived from Pitt-Hopkins Syndrome patients and respective parents of matching sex. Libraries for each individual include three biological replicates.


title: KLF10 promotes nonalcoholic steatohepatitis progression through transcriptional activation of zDHHC7 [KFL10 overexpression]

status: Public on Feb 25 2022

city: Shenzhen

first: Shu

last: Yang

address: 1017 Dongmen North Road, Luohu District, Shenzhen

country: China

institute: shenzhen people's hospital

summary: Nonalcoholic steatohepatitis (NASH), characterized by hepatic steatosis, inflammation, and liver injury, has become a leading cause of end-stage liver diseases and liver transplantation. Krüppel-like factors 10 (KLF10) is a Cys2/His2 zinc finger transcription factor that regulates cell growth, apoptosis, and differentiation. However, whether it plays a role in the development and progression of chronic liver diseases like NASH remains poorly understood. In the present study, we found that KLF10 expression was selectively upregulated in the mouse models and human patients with NASH, compared with simple steatosis (NAFL). Gain- and loss-of function studies demonstrated that hepatocyte-specific overexpression of KLF10 aggravated, whereas its depletion alleviated diet-induced NASH pathogenesis in mice. Mechanistically, transcriptomic analysis and subsequent functional experiments showed that KLF10 promotes hepatic lipid accumulation and inflammation through the palmitoylation and plasma membrane localization of fatty acid translocase CD36 via transcriptionally activation of zDHHC7. Indeed, both expression of zDHHC7 and palmitoylation of CD36 are required for the pathogenic roles of KLF10 in NASH progression. Thus, our results identify an important role for KLF10 in NAFL-to-NASH development through zDHHC7-mediated CD36 palmitoylation.

overall_design: primary hepatocyte transfected with lentivirus expressing negative control or KLF10

first: Lijing

last: Jia

first: Jiaqing

last: Xiang

first: Guangyan

first: Shanhu

last: Qiu

first: Lin

last: Kang

first: Peilin

last: Zheng

first: Yan

last: Lu

first: Zhen

last: Liang


title: KLF10 promotes nonalcoholic steatohepatitis progression through transcriptional activation of zDHHC7 [NAFL vs NASH]

overall_design: liver mRNA profiles of HFD fed mice (NAFL) and WD+CCl4 treated mice (NASH)


title: Patient-derived brain organoids uncover reversible cellular abnormalities in Pitt-Hopkins Syndrome III

overall_design: Comparison between single cell RNA-Seq libraries of brain organoids derived from Pitt-Hopkins Syndrome patients and respective parents of matching sex.


title: Reducedfemalefertility duetosequestration ofRNA Pol II by pervasive transcription in exosome RNasedepleted oocytes

status: Public on Feb 11 2022

city: Ashburn

first: Di

last: Wu

state: Virginia

address: 19700 Helix Drive

institute: HHMI Janelia

summary: We performed mouse single oocyte RNA-seq and bulk oocyte CUT&Tag assays in the current project. In details, SMART-seq based single oocyte RNA-seq was performed at adult GV stage, GV3h stage and MII stage, using control and Dis3 oocyte-speicfic knockout (cKO) oocytes. RiboMinus-seq based single oocyte RNA-seq was performed at adult GV stage using control and Dis3 cKO oocytes, and at p20 GV stage using control, Dis3 cKO, Exosc10 cKO and Dis3/Exosc10 double cKO (dcKO) oocytes. Bulk CUT&Tag of anti-H3K27me3 was done in WT and Dis3 cKO oocytes at adult GV stage, and in WT and Dis3/Exosc10  dcKO oocytes at p20 stage. In addition, CUT&Tag of anti-RNA polymerase II (Ser2+Ser5) was performed in WT and Dis3 cKO oocytes at GV stage. All CUT&Tag experiments share the same rabbit-Igg negative control. All p20 stage oocytes were specified as p20. The non-specified GV oocytes were all adult GV oocytes.

overall_design: In SMART-seq based single oocyte RNA-seq, totally 72 samples were performed, including 24 samples at GV stage, 24 samples at GV3h stage and 24 samples at MII stage. At each stage, 8 samples were from control group and 16 samples were from Dis3 cKO group. In RiboMinus RNA-seq, all groups (adult-GV-ctr, adult-GV-Dis3cKO, p20-ctr, p20-Dis3cKO, p20-Exosc10, p20-dcKO) have 8 samples each. In CUT&Tag assays, each group has 3 replicates in totall. The CUT&Tag sequencing results were all firstly randomly sampled for 3M reads for downstream analysis (without deduplication).

first: Jurrien

last: Dean


title: Cellular translational enhancer elements that recruit eukaryotic initiation factor 3

status: Public on Jul 06 2022

first: Rachel

middle: Orion

last: Niederer

state: MI

address: 1150 W. Medical Center Dr

department: Biological Chemistry

institute: University of Michigan

summary: Translation initiation is a highly regulated process which broadly affects eukaryotic gene expression. Eukaryotic initiation factor 3 (eIF3) is a central player in canonical and alternative pathways for ribosome recruitment. Here we have investigated how direct binding of eIF3 contributes to the large and regulated differences in protein output conferred by different 5′-untranslated regions (5′-UTRs) of cellular mRNAs. Using an unbiased high-throughput approach to determine the affinity of budding yeast eIF3 for native 5′-UTRs from 4,252 genes, we demonstrate that eIF3 binds specifically to a subset of 5′-UTRs that contain a short unstructured binding motif, AMAYAA. eIF3 binding mRNAs have higher ribosome density in growing cells and are preferentially translated under certain stress conditions, supporting the functional relevance of this interaction. Our results reveal a new class of translational enhancer and suggest a mechanism by which changes in core initiation factor activity enact mRNA-specific translation programs.

overall_design: RNA bind'n'seq (RBNS) of yeast 5'-UTRs binding to eIF3

first: Jiri

last: Koubek

first: Andrei

last: Stanciu

first: Colin

last: Echeverría Aitken

first: Wendy

middle: V

last: Gilbert


title: Expression profiles of mRNAs in the retinas of OIR wildtype, OIR IL-19 knockout and room air control mice.

status: Public on Jan 24 2022

city: Changsha

first: Jingling

last: Zou

state: Hunan

address: No. 139, Middle Renmin Road, furong district

department: Department of Ophthalmology

institute: The Second Xiangya Hospital of Central South University

summary: Seven-day-old C57BL/6 mice and IL-19 knockout (KO) mice were subjected to 75% oxygen for 5 days to induce OIR. To explore the mechanism that caused by IL-19 on retinal neovascularization, Gene expressions among retinas of room air control mice, OIR wildtype mice, and OIR IL-19 knockout mice were measured by RNA-sequencing (RNA-seq).

overall_design: Retinal mRNA profiles of 17-day old oxygen-induced retinopathy (OIR) wild-type (WT), OIR IL-19 knockout (KO) and room air (RA) WT mice.

first: Yedi

last: Zhou


title: Kronos scRT: a uniform framework for single-cell replication timing analysis

status: Public on Mar 29 2022

city: Paris

first: Chunlong

last: Chen

address: 26 rue d’Ulm

institute: Institut Curie

summary: Mammalian genomes are replicated in a cell-type specific order and in coordination with transcription and chromatin organization. Although the field of replication is also entering the singlecell era, current studies require cell sorting, individual cell processing and have yielded a limited number (<100) of cells. Here, we have developed Kronos scRT (https://github.com/CL-CHEN-Lab/Kronos_scRT), a software for single-cell Replication Timing (scRT) analysis. Kronos scRT does not require a specific platform nor cell sorting, allowing the investigation of large datasets obtained from asynchronous cells. Analysis of published available data and droplet-based scWGS data generated in the current study, allows exploitation of scRT data from thousands of cells for different mouse and human cell lines. Our results demonstrate that, although most cells replicate within a close timing range for a given genomic region, replication can also occur stochastically throughout S phase. Altogether, Kronos scRT allows investigating the RT program at a single-cell resolution for both homogeneous and heterogeneous cell populations in a fast and comprehensive manner.

overall_design: Single Cell replication timing using 10X Genomics scCNV platform to test Kronos scRT workflow

first: Stefano

last: Gnan

first: Joseph

last: Josephides

first: Xia

first: Manuela

last: Spagnuolo

first: Dalila

last: Saulebekova

first: Mylène

last: Bohec

first: Marie

last: Dumont

first: Laura

last: Baudrin

first: Daniele

last: Fachinetti

first: Sylvain

last: Baulande

first: Chun-Long


title: Bulk RNA-seq analysis of Etv2 induced EB differentiation.

status: Public on Jan 12 2022

first: Daniel

last: Garry

address: 2231 6th Street SE, 4th Floor

department: Lillehei Heart Institute

institute: University of Minnesota

summary: The vasculature is an essential organ for the delivery of blood and oxygen to all tissues of the body and thus relevant to the treatment of ischemic diseases, injury-induced regeneration, and solid tumor growth. Therefore, the definition of pioneer factors and regulatory pathways that govern the specification and differentiation of endothelial progenitors will serve as a platform for targeted therapies to promote cardiovascular regeneration and treatment of solid organ tumors. Previously, we demonstrated that ETV2 was an essential transcription factor for the development of cardiac, endothelial and hematopoietic lineages.  Here, we report that ETV2 functions as a novel pioneer factor that relaxes closed chromatin and regulates endothelial development. By comparing engineered embryonic stem cell differentiation and reprogramming models with single cell RNA-seq, ATAC-seq and ChIP-seq techniques, we demonstrated that ETV2 was able to bind nucleosomal DNA and function as a pioneer factor independent of the cellular context. We determined that ETV2 executes a pioneering role by recruiting BRG1 to remodel chromatin around endothelial genes and help maintain an open configuration.  ETV2 chromatin binding also resulted in increased H3K27ac deposition.  Collectively, these results will serve as a platform for the development of therapeutic initiatives directed towards cardiovascular and oncological diseases.

overall_design: Total RNA was isolated from iHA-Etv2 MEFs and EBs using the RNeasy kit (Qiagen, Cat# 74104) according to the manufacturer’s protocol. Briefly, cells were lysed in RLT-lysis buffer, followed by a column-based purification process and on-column DNA digestion. Complementary DNA (cDNA) was synthesized using the SuperScript IV VILO kit (Thermo Fisher Scientific, Cat# 11756050) according to the manufacturer’s protocol. Quantitative polymerase chain reaction (qPCR) was performed with ABI Taqman probe sets. Taqman probes used in this study include VIC-labeled GAPDH:4352339E, FAM-labeled Etv2: mm01176581_g1, Flk1: mm00440099_m1, PECAM (Cd31): mm0124616 7_m1, Tie2: mm01256892_ m1, Cdh5: mm00486938_m1, Lmo2: Mm00493153_m1, Sox18: Mm00656049_gH, Smarca4 (Brg1): Mm01151944_m1, Emcn: Mm00497495_m1, Mmp9: Mm00442991_m1, Hopx: Mm00558630_m1, Otor: Mm00498571_m1, Lax1: Mm00556050_m1, Chd8: Mm01316316_m1, Dek: Mm01351566_m1 and Znhit: Mm01201686_m1

first: Wuming

last: Gong

first: Nikita

last: Dsouza


title: Evolutionary and phenotypically plastic response of adult gene expression to larval undernutrition in Drosophila melanogaster

city: Lausanne

first: Tadeusz

middle: J.

last: Kawecki

address: Biophore

department: Department of Ecology and Evolution

country: Switzerland

institute: University of Lausanne

summary: This RNAseq study has been performed in the framework of a project using experimental evolution to understand the consequences of genetically-based adaptation to juvenile undernutrition for adult gene metabolism.   Six "Selected" populations of Drosophila melanogaster had been maintained on a very poor larval diet (3.2 g yeast / liter w/v) for 14 years (about 250 generations).   Six "Control" populations of the same origin had been maintained in parallel on the standard diet (12.5 g yeast/liter).  In a factorial design, we performed RNAseq on flies from Selected and Control populations raised both standard and poor diet.    This design separates the effects of the evolutionary differentiation between the two sets of populations, and the effect of the larval diet experienced by the focal individuals.   Adults of all populations were transferred to standard diet within 24 h or emergence and maintained on it for 3 days before being collected for RNAseq.   RNAseq was performed on carcasses of adult mated females (10 carcasses per sample), consisting mainly of the fat body and the abdominal body wall.

overall_design: 2 evolutionary regimes (Selected vs. Control) each with 6 replicate populations (S1-6, C1-6) x 2 larval diet treatments (poor vs. standard); one biological sample of 10 carcasses, all samples multiplexed and seqeunced on two lanes ("lane 2" and "lane 7"; hence two raw data files per sample).

middle: J

first: Berra

last: Erkosar

first: Cindy

last: Dupuis

first: Loriane

last: Savary


title: Etv2-induced MEF reprogramming

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: ChIP-seq analysis of Etv2 induced MEF reprogramming and EB differentiation.

summary:
ChIP-seq analysis of Etv2, Brg1 and H3K27ac post Etv2 induction in MEF and ES/EB.
Methods: EB or MEF cells were harvested as described above. ChIP assays for ETV2 and H3K27ac were performed from 2 x 10e6 MEFs or 1 x 10e7 EB cells, respectively. Briefly, cells were crosslinked with 1% formaldehyde for 10 min at room temperature and the reaction was quenched by glycine at a final concentration of 0.125 M.  For the BRG1 ChIP-seq, cells were first crosslinked in 2 mM disuccinimidyl glutarate (DSG; Life Technologies: Cat. #20593) for 30 min then in 1% formaldehyde for 10 min, quenched with glycine for 5 min.  The remainder of the ChIP protocol was performed following the protocol described by Magli et al19. We used antibodies against H3K27ac (Abcam, ab4729), ETV2 (Abcam, ab181847), and BRG1 (Abcam ab110641) for the respective pulldowns. For both of the pulldowns, protein A Dynabeads were added to the ChIP reactions and incubated for 30 minutes at room temperature. Magnetic beads were washed and chromatin was eluted. After crosslinking reversal, RNase A, and proteinase K treatment, ChIP DNA was extracted with the Min-Elute PCR purification kit (Qiagen). ChIP DNA was quantified with Quant-it PicoGreen dsDNA Assay Kit (Life Technologies). Sequencing libraries were prepared using equal amount of ChIP DNA per sample as per instructions of SMARTer® ThruPLEX® DNA-Seq Kit (Takara R400675) and SMARTer® DNA Unique Dual Index Kit - 24U Set A (Takara R400665). Sequencing was performed at the University of Minnesota Genome Center with the The NextSeq 550 high-throughput benchtop sequencer.

overall_design: ChIP-seq of Etv2, Brg1 and H3K27ac of undifferentiated MEF, 1 day, 2 days and 7 days post Etv2 induction MEFs; ChIP-seq of Etv2, Brg1 and H2K27ac at 3h and 12h post Etv2 induction in EBs


title: hsa_circ_0098181-miRNA-seq

status: Public on Dec 13 2021

city: Shanghai

first: Yuan-Yuan

last: Luo

address: Department of Gastroenterology, Shanghai East Hospital, Tongji University School of Medicine, 150 Jimo Road, Shanghai, 200120, China

institute: Tongji University

type: Non-coding RNA profiling by high throughput sequencing

summary: To explore the potential downstream miRNA of hsa_circ_0098181, we performed a miRNA sequencing.

overall_design: 3 overexpressing hsa_circ_0098181 Huh7 cells and 3 control cells were used to extract total RNA for miRNA sequencing.

first: Xin

last: Zeng

first: Ke-Gong

last: Tao

first: Yi-Ting

last: Lu

first: Bin-Bin

last: Li


title: The Wnt negative regulators RNF43 and ZNRF3 predispose to Hepatocellular carcinoma by impairing liver regeneration and controlling liver fat metabolism

status: Public on Dec 10 2021

city: Dresden

first: Meritxell

last: Huch

address: Pfotenhauerstrasse 108

institute: Max Planck Institute of Molecular Cell Biology and Genetics

summary: The homologous E3 ubiquitin ligases RNF43/ZNRF3 negatively regulate WNT signalling activation. Recently, both genes have been found mutated in several types of cancers. Specifically, loss-of-function mutations result in adenoma formation in mouse small intestine. However, their role in liver pathology and cancer has not been explored yet. Here, we describe that hepatocyte-specific deletion of Rnf43/Znrf3 results in altered lipid metabolism and steatohepatitis, in the absence of exogenous dietary fat supplementation, with a remarkable increase in unsaturated lipids. Upon liver injury, Rnf43/Znrf3 deletion results in impaired hepatocyte regeneration and liver cancer, subsequent to an imbalance between differentiation and proliferation. Using three different liver organoid models (hepatocyte-, hepatoblast- and ductal cell-derived organoids) we demonstrate that the hepatocyte differentiation defects and the lipid metabolic alterations are, at least in part, cell-autonomous. Remarkably, hepatocellular carcinoma patients with mutations in either or both genes or in other WNT pathway components also present altered hepatic lipid metabolism and non-alcoholic steatohepatitis (NASH) profiles. Our findings imply that hyperactivation of WNT signaling through the RNF43/ZNRF3 module predisposes to liver cancer by impairing hepatocyte differentiation and altering the lipid metabolic ground-state of the liver. Both mechanisms combined facilitate the progression towards malignancy. Our findings might aid on the management of those RNF43/ZNRF3 or WNT mutated individuals at risk of developing fatty liver and/or liver cancer.

overall_design: RNAseq of mouse livers WT (24 samples) and KO (24 samples) for Rnf43 and Znrf3 and with chronic damage (11 samples), partial hepatectomy (12 samples) at different time points or without damage (25 samples)

first: Gianmarco

last: Mastrogiovanni


title: Differential brain gene expression in female and male D. melanogaster at 3, 7, and 14 days post-eclosion

status: Public on Mar 25 2022

city: San Diego

first: Goran

last: Bozinovic

address: 3030 Bunker Hill St

institute: Boz Life Science Research and Teaching Institute

summary: This study provides a screen of sexually dimorphic tissue-specific (brain) gene expression in fruit flies age 3, 7, and 14 days post-eclosion. This work can be used as a baseline reference point for comparing sex-specific brain gene expression levels in fruit flies under stress.

overall_design: Effects of age (3, 7, and 14 days post-eclosion) and sex on gene expression in the brain.

first: Jeanne

last: Vu

first: Flannery

last: McLamb

first: Lindsey

last: Griffin

first: Sylvia

first: Zuying

last: Feng


title: Congenital T cell activation blocks transitional to follicular B cell maturation in humans

status: Public on Nov 10 2021

first: Michelle

last: Ramseier

state: MA

address: 400 Main St

department: Harvard-MIT Health Sciences and Technology

institute: Massachusetts Institute of Technology

summary: In patients with CTLA4 deficiency, CD40L-expressing T cells inhibit transitional to follicular B cell maturation in the T cell zone of lymph nodes through the induction of mTORC1 signaling in transitional B cells. We apply scRNA-seq to identify upregulation of PI3K, mTORC1, and oxidative phosphorylation pathways in patients with CTLA4 deficiency, and to characterize resolution of follicular block after treatment with abatacept.

overall_design: scRNA-seq of transitional B cells from healthy donors and patients with CTLA4 deficiency; one set of pre- and post-abatacept treatment from the same CTLA4-deficient patient.

first: Jocelyn

last: Farmer

first: Alex

last: Shalek

first: Shiv

last: Pillai


title: CLIP-seq analysis of KSHV ORF57-associated RNAs

status: Public on Nov 08 2021

city: Frederick

first: Zhi-Ming

last: Zheng

state: Maryland

department: Center for Cancer Research

institute: National Cancer Institute

summary: Kaposi's sarcoma-associated herpesvirus (KSHV) ORF57 is a viral RNA-binding protein essential for viral lytic gene expression. ORF57 binds to target RNA directly via interaction with cellular cofactors. To investigate the entire repertoire of ORF57-associated RNAs we performed UV cross-linking immunoprecipitatin (CLIP) experiment using an affinity-purified,  highly specific anti-ORF57 antibody in KSHV-infected primariy effusion lymphoma BCBL-1 cells undegoing lytic virus replication.

overall_design: Cross-linking Immunoprecipitation (CLIP) using antibody against KSHV ORF57 followed by RNA sequencing

first: Beatriz

last: Alvarado Hernandez

first: Vladimir

last: Majerciak


title: Single-cell transcriptomes and runx2b−/− mutants reveal the genetic signatures of intermuscular bone formation in teleosts

status: Public on Nov 04 2021

city: wuhan

first: yulong

last: chen

state: hubei

address: shizishan street  hongshan district

institute: huazhong agricultural university

summary:
Purpose: Intermuscular bones (IBs) are hard spicules, mainly existing in the myosepta of recent vertebrates. This evolutionary trait has puzzled biologists and consumers, and the molecular basis of IB development remains unclear. The goals of this study are to acquire characteristic single-cell maps of gene expression landscapes and lineages and to elucidate the differentiation trajectory of cell clusters related to IB formation and obtain the key regulated gene of IB formation.
Methods: The scRNA-seq profiles of 60 dpf wild-type zebrafish and 60 dpf runx2b-/- zebrafish were generated, using the 10X Genomics Chromium system. The bulk RNA-seq profiles of 60 dpf wild-type zebrafish and 60 dpf runx2b-/- zebrafish were generated, using mirVana™ miRNA ISOlation Kit. The scRNA-seq data were filted and clsutered by the Seurat package. The cell differentiation trajectorie was constructed, using Monocle analysis. Runx2b knockdown was preformed by RNA interference and runx2b was inducted by neuropeptide substance P. qRT–PCR validation was performed using SYBR Green assays. The mutant zebrafish lines were constructed, using cripsr cas9 technology.The Phenotypic observation was perfomed by Hematoxylin and Eosin (HE), Alizarin red, masson trichrome staining and mico-CT. The nutritent content was also obtained including fatty acid content and anio acid content.
Results: we reveal that teleost fish IBs originate from tendons in the skeletal muscle and that the differentiation trajectory from tendon progenitors and defined runx2b as the key rugulated gene of IB formaiton, using cripsr cas9 technology.
Conclusions: This study is the first to reveal that teleost fish IBs originate from tendons in the skeletal muscle and that the differentiation trajectory from tendon progenitors to the osteoblast lineage is key for IB formation. Large-scale gene function analysis using CRISPR-Cas9 identified the decisive role of runx2b in IB formation. The loss of runx2b significantly restricted osteoblast differentiation and inhibited IB formation, which potentially provides a basis for breeding strains of fish without IBs to improve the safe consumption and economic value of many aquaculture species and in this way, boost the proportion of fish for the supply of animal protein worldwide (Fig. 6). For this reason, we needed to verify that the content of fatty acid in the runx2b−/− mutant was higher than that in runx2b+/+ fish, as well as the expression of key genes related to IBs in the runx2b−/− and runx2b+/+ strains obtained by scRNA-seq data. This study also provides materials and directions for further study to clarify the specific regulatory mechanism of IB formation and will help to discover more specific genetic resources that can be used for IB trait improvement in fish.

overall_design: the scRNA-seq and bulk-seq profiles of 60 dpf wild-type zebrafish and 60 dpf runx2b-/- zebrafish

first: Chun-Hong

last: Nie

first: Shi-Ming

last: Wan

first: Yu-Long

last: Chen

first: Ya-Ming

last: Wu

first: Jia-Jia

last: Zhou

first: Ann

last: Huysseune

first: Paul

last: Witten

first: Alexandre

middle: W

last: Silva Hilsdorf

first: Wei-min

last: Wang

first: Qiang

last: Lin

first: Ze-Xia

last: Gao


title: KLF10 promotes nonalcoholic steatohepatitis progression through transcriptional activation of zDHHC7

first: Shu

last: Yang


title: Protein-RNA interactome analysis of KSHV ORF57 in BCBL-1 cells with lytic virus infection


title: Gene expression profiling in HPV-infected oropharyngeal head and neck cancer

status: Public on Nov 03 2021

city: Milan

first: Loris

last: De Cecco

address: via Venezian 1

country: Italy

institute: IRCSS Istituto Nazionale Tumori

summary: The prognosis of HPV-positive squamous oropharyngeal carcinomas (OPCs)  is more favorable than that of HPV-negative OPCs. However, the prognosis of some of these tumors is dismal and to date validated survival predictors are missing in clinical practice. We designed a study on HPV-positive OPCs to determine whether a previously published gene expression tumor classification model,defined through meta-analysis of publicly available datasets (PMCID: PMC6721309), is able to predict survival in an external patient cohort. The 3 gene expression clusters were tested in a validation set of 286 cases and in  a refined study population of 235 patients and in both confirmed their prognostic value. Five-year OS was 95% in the low risk cluster Cl1, 80% in the intermediate risk cluster (HR=5.74, p=.0057), and 66% in the high risk cluster (HR=9.28, p=.001). Functional/biological cluster characterization identified potentially targetable pathways in the high risk cluster.  This prognostic stratification might be useful for clinical decision making and for planning future trials based on molecular tumor features.

overall_design: In our work, we considered a cohort of the HPV positive OPC patients included in the project “Big Data and Models for Personalized Head and Neck Cancer Decision Support (BD2Decide)”, trial registry ClinicalTrial.gov  number NCT02832102. Patients were treated in seven European referral cancer centers (Italy, The Netherlands and Germany) with expertise in the multidisciplinary management of head and neck cancer patients. Median follow-up was 46.2 months (95% confidence interval 41.2-50) and data collection was concluded in September 2019 (doi:10.1002/hed.26515). OPC patients with loco-regionally advanced non-metastatic disease treated with curative intent were tested for HPV using p16-immunostaining and HPV DNA PCR and when positive included. HPV status was assessed by p16-immunostaining followed by a HPV DNA PCR test on the p16-immunopositive cases. Patients were treated with multimodal strategies (combinations of surgery, radiotherapy, and chemotherapy). For T1N1M0 and T2N1M0 disease single-modality treatment (surgery or radiotherapy) was allowed. To reduce the bias of confounding variables impacting on outcome, we excluded OPC: i) patients with p16-positive but HPV-DNA-negative OPC;  ii) patients receiving unimodal treatment (surgery without post-operative radiation or exclusive radiotherapy without concomitant systemic treatments) for clinical stage T1N>1, T2N>1 and T3-T4 any N disease. TNM7= AJCC/UICC staging system 7th edition; TNM8= AJCC/UICC staging system 8th edition;  subset (n=235)=  exclusion of 51 HPV-DNA negative (n=14) OPCs and suboptimal treatments (n=37); HPV_molecular_clusters= gene-expression stratification based on Locati et al  (PMCID: PMC6721309).


title: Distinct hepatic gene expression patterns characterize progressive disease in NAFLD

status: Public on Oct 15 2021

city: Boston

first: Sonu

last: Subudhi

address: 100 Blossom St, Cox 7

department: Radiation Oncology

institute: Massachusetts General Hospital

summary: The pathogenesis of non-alcoholic fatty liver disease is not fully understood. Transcriptomic analysis of a large cohort of 318 patients provides evidence of gene perturbations related to inflammation, complement and coagulation pathways, and tissue remodeling in distinct states of NAFLD.

overall_design: Using the Nanostring nCounter assay, we quantified expression of 795 genes (and 5 housekeeping genes) hypothesized to be involved in hepatic fibrosis, inflammation, and steatosis in liver tissue from 318 adults. Liver specimens were categorized into 4 distinct NAFLD phenotypes: normal liver histology (NLH), steatosis only (steatosis), non-alcoholic steatohepatitis without fibrosis (NASH F0), and NASH with fibrosis stage 1-4 (NASH F1-F4).

first: Kathleen

last: Corey

first: Georg

middle: M

last: Lauer


title: Type I interferon pathways in SARS-CoV-2-infected individuals from Pakistan

status: Public on Dec 01 2021

city: Karachi

first: Asghar

last: Nasir

state: Sindh

address: Stadium Road

department: Pathology and Laboratory Medicine

country: Pakistan

institute: Aga Khan University

summary: Blood transcriptomes were determined in COVID-19 cases and healthy controls using the Clariom S RNA microarray, Affymetrix assay.

overall_design: COVID-19 cases with a positive respiratory samples SARS-CoV-2 and healthy Controls cases were recruited. Blood transcriptomes were analysed using Clariom S RNA Microarray, Affymetrix Inc.

first: Kiran

middle: I

last: Masood

first: Maliha

last: Yameen

first: Saba

last: Shahid

first: Zahra

last: Hasan


title: ATAC-seq analysis of Etv2 induced MEF reprogramming and EB differentiation.

city: Minneapolis

first: Daniel

last: Garry

summary:
ATAC-seq analysis of Etv2 induced ES/EB, MEFs,Brg1 KO EBs and Brg1 KD MEFs.
Methods: EBs were collected at different time points and disaggregated in 0.25% trypsin at 37C for 3 min incubation with gentle agitation followed by inactivation with culture medium containing 10% FBS. Cells were collected by centrifugation at 500g for 5 minutes, washed once with ice-cold PBS. iHA-Etv2 MEFs were harvested from the culture dishes at different time points by treating the cells with 0.25% trypsin at 37C for 4 min incubation followed by neutralizing and collecting the cells with culture medium containing 10% FBS. ATAC reaction was performed with 50,000 cells as previously described18 using the Tn5 transposase (Illumina) and libraries were created at the University of Minnesota Genome Center. Libraries were then sequenced on a NextSeq Illumina platform (2x50 bp) aiming for 25 million reads per sample.

overall_design: ATAC-seq of undifferentiated MEF, 1 day, 2 days and 7 days post Etv2 induction MEFs; ATAC-seq of 3h and 12h post Etv2 induction in EBs; ATAC-seq of undifferentiated Brg1 KD MEFs, 1 day, 2 days and 7 days post Etv2 inducdtion in Brg1 KD MEFs and Brg1 KO EBs 4 days post Etv2 induction

first: Wuming

last: Gong

first: Nikita

last: Dsouza


title: RNA-seq analysis of KSHV-infected BCBL-1 cells, a primary effusion lymphoma cell line

first: Zhi-Ming

last: Zheng

summary: Primari effusion lymphoma are (PEL) patient-derived transformed B-cells harboring latent Kaposi's sarcoma-associated herpesvirus (KSHV). The treatment of PEL cells with valproic acid (VA) leads to reactivation of KSHV  and viral lytic replication. The aim of this project is to evaluate the effect of KSHV lytic infection on expression of the host transcriptome.

overall_design: A differential expression analysis of host transcritome in BCBL-1 cells with latent infected versus lytic infection

first: Beatriz

last: Alvarado Hernandez

first: Vladimir

last: Majerciak


title: Scalable Production of Mouse Erythroid Cells using an in vitro Embryoid Body differentiation System

status: Public on Sep 22 2021

city: Oxford

first: Mira

middle: Tony

last: Kassouf

state: England

address: John Radcliffe Hospital, Headly way

department: Weatherall Institute of Molecular Medicine

institute: University of Oxford


title: Scalable Production of Mouse Erythroid Cells using an in vitro Embryoid Body differentiation System [ChIP-Seq]

summary: Mouse embryonic stem (mES) cells can be manipulated ex-vivo to recapitulate the process of erythropoiesis, whereby multipotential haematopoietic stem cells undergo lineage specification, differentiation and maturation to produce functional erythroid cells. Although very useful for identifying specific progenitors and precursors, this cell system has not been fully exploited as a source of cells to analyse erythropoiesis, compared with primary erythroid cells or immortalised cell lines. Here we have established a protocol in which erythroid cells can be isolated in a scalable manner from differentiated embryoid bodies (EBs). Using transcriptional and epigenetic analysis, we show that all purified erythroid cells in this system are derived from the primitive haematopoietic lineage. Finally, we show that this system faithfully recapitulates normal erythropoiesis in the embryonic mouse and fully mimics the effects of natural and engineered mutations seen in full mouse models. Data deposited here comprises chromatin profiling of the EB derived CD71+ erythroid  cells; chromatin accessibility (ATAC-Seq), characterisation of the open chromatin by associated histone modifications (ChIP-Seq), chromatin structure (CTCF ChIP-Seq and Capture C interaction data). WT CD71+ cells were compared to either mutant CD71+ cells or other erythroid tissues (mouse adult spleen, E13.5 fetal liver, early embryonic circulating blood) representing both definitve and primitive erythroid lineages.

overall_design: Purified CD71+ erythroid cells derived from differentiated Wildtype (WT) or mutant mouse Embryonic Stem (mES), D3839, deleted for the CTCF boudary elements at HS-38 and HS-39.5 positions at the Alpha Globin locus (Hanssen L et al, Nature Genetics, 2017) were used in these ChIP assays. The profiles produced from WT were used to confirm the primitive nature of the CD71+ erythroid cells derived from EBs. The WT CTCF profile was also used for comparison to that of the D3839 CD71+ cells .

first: Helena

last: Francis

first: Caroline

last: Harrold


title: Scalable Production of Mouse Erythroid Cells using an in vitro Embryoid Body differentiation System [ATAC-Seq]

overall_design: Purified CD71+ erythroid cells derived from differentiated Wildtype (WT) or mutant mouse Embryonic Stem (mES) cells at various sclales (in bulk in 10cm dishes or in wells of 96-well plates) were compared to mouse erythroid cells derived from WT adult spleen (Ter119+), mouse embryonic E13.5 fetal livers (based on CD71+, Ter119+ immunophenotype), or circulating embryonic blood day E9.5, E10.5 and E11.5

first: Matthew

last: Gosden


title: Ex vivo treatment of peripheral blood mononuclear cells (PBMC) with L-sulforaphane

status: Public on Jan 09 2022

city: Melbourne

first: Assam

last: El-Osta

address: 75 Commercial Road

country: Australia

institute: Baker Heart and Diabetes Institute

summary: Blood samples were collected from healthy volunteers, PBMC were collected using standard ficoll centrifugation. RNA was collected from untreated and PBMC treated with 15 µM L-sulforaphane for 24 hours. Libraries were prepared for mRNA-Seq.

overall_design: Four untreated control PBMC samples compared to four PBMC samples treated ex-vivo with 15 µM L-sulforaphane for 24 hours.

first: Scott

last: Maxwell

first: Katherine

last: Ververis

first: Tom

last: Karagiannis

first: Mark

last: Ziemann

first: Ishant

last: Khurana


title: RNA-Seq of kidney glomeruli and renal tubules from Type 2 diabetic (db/db) and non-diabetic mice

status: Public on Sep 29 2021

city: Wuhan

first: Luo

last: Qiang

state: Hubei

address: Zhang Zhidong Road

institute: Renmin hospital of wuhan university

summary: We investigated the gene expression profiles of RNA isolated from kidney glomeruli and renal tubules from aged, 24 week old type-2 diabetic (db/db) and non-diabetic mice

overall_design: In order to investigate the consequences of hyperglycemia on the pathogenesis and progression of diabetic nephropathy Kidney glomeruli and renal tubules, glomeruli and renal tubules  were isolated from 9 db/db mice and 9 db/m mice. Each sample included glomeruli from 3 mice, and three independent biological replicate samples were sequenced from each group. RNA was purified for RNA-Seq analysis on the Illumina HiSeq 4000. The goal of the project was to generate comprehensive list of RNA genes differentially regulated between the two conditions in order to identify novel targets for further study.

first: Qiang

last: Luo

first: ZongWei

last: Zhang

first: Keju

last: Yang

first: Wei

last: Liang


title: Acetylation State of Histone Core Defines Macrophage Dynamics in Diabetic Wounds

status: Public on Sep 14 2021

city: Ulm

first: Pallab

last: Maity

address: Meyerhofstrasse/James Franck Ring

department: Dermatology

institute: University of Ulm

summary: Dysregulation of macrophage populations at the wound site is responsible for the non-healing state of chronic wounds. The molecular mechanisms underlying macrophage dysfunction and its control in diabetes are largely unexplored on an epigenetic level. Here, we report that acetyl histone-H3 (Lys27), an epigenetic mark regulating the macrophage transcriptome, is lost in the hostile tissue microenvironment in diabetes. The diabetic microenvironment, profoundly suppresses the acetylation of histone by activating HDACs-dependent deacetylation pathways. This, in consequence, suppress the STAT1 signaling in macrophages maintained in diabetic conditions. Interestingly, the HDAC inhibitor butyrate - via restoring the acetyl histone-H3 (Lys27)-dependent transcriptome - effectively rescues macrophage functions in a diabetic microenvironment. Butyrate reinstalls the STAT1 mediated transcription program and consequently macrophage activity depicting a unique fingerprint of tissue regeneration and inflammation control even in a hostile diabetic microenvironment. Most interesting, butyrate breaks the vicious cycle of inflammation in diabetic wounds. Our study offers novel pathogenic insight and the unique opportunity to reverse perturbed macrophage function thus holding promise to successfully treat diabetic and other chronic wounds and conditions of unrestrained inflammation.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
For RNA-Seq analysis, 1-5 µg of total RNA from total skin was first rRNA depleted using either Ribominus Eukaryotic system v2 kit (ThermoFisher Scientific). rRNA depleted RNA was subjected to preparation of Illumina compatible RNA-seq libraries using NEBNext Ultra II Directional RNA Library Preparation kit (NEB). RNA-seq libraries were quality controlled through Qiaxcel advanced system (Qiagen) and Bioanalyzer (Agilent) and finally measured by Qubit fluorimeter (ThermoFisher Scientific). Validated libraries were sequenced on Illumia NEXTSEQ 500 and NOVASEQ 6000 system.
Single cell RNA sequencing was performed using Evercode WT v3 single cell whole transcriptome kit (Parse biosciences, USA), as per manufacturer instructions. In brief, single cell suspensions (~50,000 cells from each sample) from unwounded (Day 0) and post-wound (Day 5 and Day 10) mouse skin, were prepared and used for scRNAseq.


title: ChIP-seq analyses of human periodontal ligament fibroblasts (PDLF) transfected with siRNA for PPARg

status: Public on Aug 31 2021

city: Okayama

first: Shigeki

last: Suzuki

state: Okayama

address: 2-5-1 Shikata-cho, Kita-ku

department: Department of Operative Dentistry

country: Japan

institute: Okayama University

summary: Periodontal ligament is soft connective tissue between alveolar bone and cementum, the surface hard tissue of tooth. Periodontal ligament fibroblasts (PDLF) actively express osteo/cementogenic genes, which contribute in periodontal tissue homeostasis. However, the key factors by which PDLF retain such osteo/cementogenic abilities has not been clarified yet. Here, we find PPARg is expressed in vivo periodontal ligament tissue and its distribution pattern is highly correlated with the alkaline phosphate enzymatic activity. Knockdown of PPARg drastically deprives PDLF of osteo/cementogenic abilities in vitro. In contrast, PPARg agonists enhance it. PPARg is required for maintaining acetylation status of H3K9 and H3K27, active chromatin marks, and the supplementation of acetyl-coA, the donor of histone acetylation, is able to restore PPARg knockdown-induced decreased osteo/cementogenic abilities in PDLF. RNA-seq and ChIP-seq combined analyses identify 4 osteogenic transcripts, RUNX2, SULF2, RCAN2, and RGMA, in the PPARg-dependent active chromatin region marked by H3K27ac. Furthermore, the RUNX2 binding sites are selectively enriched in the PPARg-dependent active chromatin region. Together, these findings identify PPARg is the key transcriptional factor of PDLF to retain osteo/cementogenic abilities and reveal that global H3K27ac modification involves in the comprehensive osteo/cementogenic transcriptional alteration mediated by PPARg.

overall_design: Comparison of the H3K27ac peaks between control siRNA- and si-PPARg-transfected PDLF.

first: Hanu

last: Yuan

first: Satoru

last: Yamada


title: RNA-seq analyses of human periodontal ligament fibroblasts (PDLF) transfected with siRNA for PPARg

summary: Purpose : This study was performed to investigate the effects of knockdown of PPARg for osteo/cementogenic differentiation of PDLF through RNA sequencing.  Method : PDLF were transfected wtih either control siRNA or siRNA for PPARg and then cultured in mineralization inducing medium for 6 days. Total RNAs were collected before and after the culture.  Result : Gene ontology analyses of RNA-seq data revealed the genes functionally related with ECM such as collagen synthesis and trimerization, proteoglycans, and fibers were selectively increased by 6-day culture and suppressed biological processes by si-PPARg compared with si-control at day 6 exhibited 5 of 7 upper ranked biological processes were related with ECM organization and metabolism. Moreover, the upper rank of suppressed pathways by si-PPARg compared with si-control at day 6 showed `Endochondral Ossification` was ranked 1st. Conclusions:These results suggested that knockdown of PPARg was sufficient to eliminate osteo/cementogenic and ECM organizing potentials by comprehensive transcriptional dysregulation in PDLF.

overall_design: Examination of the gene expression changes during long-term culture and also between control siRNA- and si-PPARg-transfected PDLF.


title: ChIP-seq and RNA-seq of human periodontal ligament fibroblasts (PDLF) transfected with siRNA for PPARg

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

overall_design: Refer to individual Series


title: SPT5 stabilizes promoter-proximal RNA polymerase II prior to release into gene bodies

city: Chicago

first: Ali

last: Shilatifard

state: IL

address: 320 E Superior St

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University Feinberg School of Medicine

summary:
The regulation of gene expression by RNA polymerase II (Pol II) is a multistep process requiring the concerted action of diverse transcription factors.
Very little is known about in vivo function of transcription factor SPT5 as its deletion results in loss of viability. To circumvent this issue and define in vivo mechanism of action for SPT5, we employed acute degradation of SPT5 and studied its consequence on transcription. We find that SPT5 loss triggers degradation of the core Pol II subunit RPB1, a process which we show to be evolutionarily conserved from yeast to human. This RPB1 degradation requires the E3 ubiquitin ligase Cullin 3, the unfoldase VCP/p97 and a novel form of CDK9 kinase complex. SPT5 specifically stabilizes Pol II at promoter proximal regions, permitting Pol II release from promoters to gene bodies. Our findings provide mechanistic insight into the in vivo function of SPT5 in stabilization of promoter-proximal Pol II prior to release into gene bodies for safeguarding accurate gene expression.

sample_organism:
Saccharomyces cerevisiae
Homo sapiens

platform_organism:
Saccharomyces cerevisiae
Homo sapiens

overall_design: Examination of SPT5 functions in human and yeast cells using acute and rapid depletion system and high-throughput sequencing (ChIP-seq, PRO-seq, TT-seq).

first: Yuki

last: Aoi

first: Marta

last: Iwanaszko


title: Panoramix SUMOylation at chromatin recruits the heterochromatin machinery to piRNA target loci [GRO-seq]

status: Public on Aug 22 2021

city: Vienna

first: Julius

middle: Fabian

last: Brennecke

address: Dr. Bohr Gasse 3

country: Austria

institute: IMBA-Institute for Molecular Biotechnology

summary: Nuclear Argonaute proteins, guided by small RNAs, mediate sequence-specific heterochromatin formation. The molecular principles that link Argonaute–small RNA complexes, once bound to a nascent target RNA, to general heterochromatin effectors are poorly understood. Here, we uncover the mechanistic basis of how the PIWI interacting RNA (piRNA) pathway engages the heterochromatin machinery in Drosophila. Piwi-mediated recruitment of the corepressor complex SFiNX to chromatin leads to SUMOylation of its Panoramix subunit. SUMOylation, together with a hydrophobic LxxLL motif in the intrinsically disordered repressor domain of Panoramix, are necessary and sufficient to recruit Small ovary (Sov), a multi-zinc finger protein essential for general heterochromatin formation and viability. Structure-guided mutations that abrogate the Panoramix-Sov interaction or that prevent Panoramix SUMOylation uncouple Sov from the piRNA pathway, resulting in viable but sterile flies that exhibit de-repression of Piwi-targeted transposons. Thus, coupling the piRNA-guided recruitment of a corepressor to chromatin with its SUMOylation underlies sequence-specific heterochromatin formation.

overall_design: GRO-seq of OSCs.

first: Veselin

last: Andreev

first: Changwei

last: Yu

first: Juncheng

last: Wang

first: Jakob

last: Schnabl

first: Laszlo

last: Tirian

first: Maja

last: Gehre

first: Dominik

last: Handler

first: Grzegorz

last: Sienski

first: Lisa

last: Baumgartner

first: Peter

last: Duchek

first: Karl

last: Mechtler

first: Dinshaw

last: Patel


title: Establishment of Bovine Induced Pluripotent Stem Cells

status: Public on Sep 27 2021

city: Xianyang

first: Young

last: Tang

state: Shaanxi

address: #22 Xinong Road, Yangling District

department: College of Veterinary Medicine

institute: Northwest A&F University

summary: Pluripotent stem cells (PSCs) have been successfully developed in many species. However, generating bovine induced pluripotent stem cells (biPSCs) has been challenging. Here we report the generation of biPSCs by overexpression of lysine-specific demethylase 4A (KDM4A) and repro-gramming factors OCT4, SOX2, KLF4, cMYC, LIN28, and NANOG (KdOSKMLN). These biPSCs exhibited silenced transgene expression at passage 10, and had prolonged self-renewal capacity for over 70 passages. The biPSCs have flat, primed-like PSC colony morphology in combined me-dia of knockout serum replacement (KSR) and mTeSR, but switched to dome-shaped, naïve-like PSC colony morphology in mTeSR medium and 2i/LIF with single cell colonization capacity. These cells have comparable proliferation rate to the reported primed- or naïve-state human PSCs, with three-germ layer differentiation capacity and normal karyotype. Transcriptome analysis revealed a high similarity of biPSCs to reported bovine embryonic stem cells (ESCs) and embryos. The naïve-like biPSCs can be incorporated into mouse embryos, with the extended capacity of in-tegration into extra-embryonic tissues. Finally, 24.6% cloning efficiency could be achieved in nu-clear transfer (NT) experiment using late passage biPSCs as nuclear donors. Our report represents a significant advance in the establishment of bovine PSCs.

overall_design: Comparing mRNA profiles of biPSCs and bMSCs with the previously published data of 4 bovine ESCs from GSE110036 (GSM2976768, GSM2976772, GSM2976773, GSM2976777), 3 bovine 16-cell embayos from GSE52415 (GSM1265770, GSM1265771, GSM1265772), and 3 bovine blastocysts (GSM1265773 from GSE52415, GSM1265774 from GSE52415, GSM2976764 from GSE110036). For these previously published raw data, sequencing adapters and reads with low quality were trimmed using Cutadapt before mapping. The quality of reads after filtering was examined using fastQC. For mapping, bovine genomic sequence and RefSeq gene coordinate (ARS-UCD1.2/bosTau9) were downloaded from the UCSC genome browser. All filtered reads were aligned to bovine reference genome by RNA STAR (Galaxy Version 2.7.8a) with default parameters. The number of reads per gene was counted by featureCounts (Galaxy Version 2.0.1). Differentially expressed genes between different samples were identified using default parameters in DESeq2 (Galaxy Version 2.11.40.6+galaxy1), which generated a principal component analysis plot and the heatmap of the sample-to-sample distance matrix. The most differentially expressed genes (adjusted p-value < 0.05) were extracted from DESeq2 results with an absolute fold change > 5. The normalized counts for those differentially expressed genes and the Z-score of the counts were calculated on the galaxy platform and exhibited as heatmaps by Heatmap2 (Galaxy Version 3.0.1).

first: Yue

last: Su

first: Ling

first: Zhiqiang

last: Fan

first: Ying

last: Liu

first: Jiaqi

last: Zhu

first: Deborah

last: Kaback

first: Julia

last: Oudiz

first: Tayler

last: Patrick

first: Siu

last: Yee

first: Xiuchun

last: Tian

first: Irina

last: Polejaeva


title: Panoramix SUMOylation at chromatin recruits the heterochromatin machinery to piRNA target loci

first: Julius

last: Brennecke


title: Analysis of Amniotic Fluid-Derived Exosomal miRNAs in Fetuses with Tetralogy of Fallot

status: Public on Oct 19 2021

city: Quanzhou

first: Guorong

last: Lyu

state: Fujian

address: No.950, Donghai street

department: Department of Ultrasound

institute: The Second Affiliated Hospital of Fujian Medical University

summary: Tetralogy of Fallot (TOF) is the most common cyanotic congenital heart disease (CHD). Amniotic fluid (AF) contains a higher abundance of biological compounds which could direct reflect the information of fetal health. AF-derived exosomal miRNAs have been investigated as etiological factors in the pathogenesis of TOF. The purpose of this study was to analyze the expression of AF-derived exosomal miRNAs and their roles in TOF development. In this study, we identified differentially expressed (DE)-miRNAs between TOF group and control group. A total of 257 miRNAs were differentially expressed (DE) between TOF and control groups. KEGG pathway enrichment demonstrated that these target genes of DE-miRNAs were involved in Wnt and Notch signaling pathway. Twenty five of 257 DE-miRNAs were overlapped with Notch-regulated and Wnt-regulated miRNAs simultaneously.

overall_design: Investigate the expression of AF-derived exosomal miRNAs

first: Hainan

last: Yang

first: Yadan

last: Li


title: Panoramix SUMOylation at chromatin recruits the heterochromatin machinery to piRNA target loci [RNA-seq]

overall_design: RNA-seq of Piwi KD, Panx KD, Sov KD, HP1 KD or Control KD OSCs.

first: Veselin

last: Andreev

first: Changwei

last: Yu

first: Juncheng

last: Wang

first: Jakob

last: Schnabl

first: Laszlo

last: Tirian

first: Maja

last: Gehre

first: Dominik

last: Handler

first: Grzegorz

last: Sienski

first: Lisa

last: Baumgartner

first: Peter

last: Duchek

first: Karl

last: Mechtler

first: Dinshaw


title: Panoramix SUMOylation at chromatin recruits the heterochromatin machinery to piRNA target loci [CUT&RUN]

overall_design: CUT&RUN of OSCs.

last: Patel


title: Senescent tissue-resident mesenchymal stromal cell is an internal source of inflammation in human osteoarthritic cartilage

status: Public on Aug 16 2021

city: PROVIDENCE

first: ALEXANDER

last: Brodsky

state: Rhode Island

address: RHODE ISLAND HOSPITAL; 593 EDDY ST.,, Aldrich 618

country: USA

institute: RHODE ISLAND HOSPITAL/BROWN UNIVERSITY

summary: RNAseq analysis indicates that the transition from normal cartilage stromal cells (nCSC) to OA-MSC during aging results in activation of SASP gene expression.

overall_design: RNA-seq of osteoarthritic chondrocytes, normal cartilage stromal cells and osteoarthritic cartilage mesenchymal stromal cells.

first: Qian

last: Chen

first: Alexander

first: Wenguang

last: Liu


title: Scalable Production of Mouse Erythroid Cells using an in vitro Embryoid Body differentiation System [Capture-C]

first: Mira

last: Kassouf

country: United Kingdom

overall_design: NG Capture-C combines 3C library preparation with oligonucleotide capture for the desired viewpoint restriction fragments (DpnII fragment), in this case the Alpha Globin R1 enhancer and promoters of Rhbdf1 and Mpg genes.  Purified CD71+ erythroid cells derived from differentiated Wildtype (WT) or mutant mouse Embryonic Stem (mES), D3839, deleted for the CTCF boudary elements at HS-38 and HS-39.5 positions at the Alpha Globin locus (Hanssen L et al, Nature Genetics, 2017) were used in this assay. The profiles produced from WT and D3839 CD71+ cells were then compared.

first: Matthew

last: Gosden


title: Amino acids activate mTORC1 to release roe deer embryos from decelerated proliferation during diapause [Embryo]

status: Public on Aug 15 2021

city: Berlin

first: Vera

last: van der Weijden

address: Ihnestraße 63-73

department: Department of Genome Regulation

institute: Max Planck Institute for Molecular Genetics

summary: mRNA sequencing of diapausing and embryos that resumed a fast rate of development to identify molecular factors involved in the regulation of embryonic diapause

overall_design: Embryonic mRNA profiles of diapausing and reactivated roe deer embryos

middle: A

first: Jochen

middle: T

last: Bick

first: Stefan

last: Bauersachs

first: Anna

middle: B

last: Rüegg

first: Thomas

last: Hildebrandt

first: Frank

last: Goeritz

first: Katarina

last: Jewgenow

first: Pieter

last: Giesbertz

first: Hannelore

last: Daniel

first: Emilie

last: Derisoud

first: Rupert

last: Bruckmaier

first: Barbara

last: Drews

first: Susanne

last: Ulbrich


title: Single-cell analyses unravel cell type-specific responses to a vitamin D analogue in prostatic precancerous lesions

status: Public on Aug 04 2021

first: Gilles

last: Laverny

address: 1 rue Laurent fries

institute: IGBMC