title: Small RNA-seq comparing transcriptome (small RNAs) of spermatozoa from rat fed either a standard-Diet (SD) or a High-Protein-Diet (HPD)

city: Nice Cedex 3

first: Maria

last: Stathopoulou

address: 151 Route de Saint-Antoine de Ginestière

country: France

institute: Inserm

summary: Gene expression analysis of sperm small RNA obtained from rat fed either a standard (SD) or High Protein Diets (HPD). The aim of the study is to identify differentially expressed  small RNA in sperm of SD-fed and HPD-fed males which might be involved in the  epigenetic inheritance of glucose sensitivity induced by a High Protein Diet.

overall_design: Small RNA profiles of spermatozoa from WT rat fed either a Standard (SD) or High Protein (HPD) diet were generated by deep sequencing, in triplicate.

first: Jamileh

last: Movassat

first: Valérie

last: Grandjean

first: Kevin

last: Lebrigand

first: Pascal

last: Barbry


title: Recombinant Anti-thrombin treatment against endotoxemia-induced  acute respiratory distress syndrome (ARDS).

status: Public on May 20 2021

city: Gifu

first: Hideshi

last: Okada

address: 1-1 Yanagido

department: Emergency and Disaster Medicine

country: Japan

institute: Gifu University Graduate School of Medicine

summary: We made LPS induced ARDS model mice, and determined the effect of recombinant anti-thrombin against ARDS.

overall_design: Following 16-h starvation, mice were intraperitoneally injected with LPS (20 mg/kg; MilliporeSigma, Burlington, MA), and in some assays, with saline or recombinant Anti-thrombin (rAT; 750 IU/kg; Kyowa Kirin Co., Ltd, Tokyo, Japan) at 3 and 24 h post-LPS injection. Total RNA was extracted from mouse lung 30h after LPS injection.


title: Long noncoding RNAs in traumatic brain injury accelerated fracture healing

status: Public on May 24 2021

city: chongqin

first: Yao

last: Yang

state: chongqin

address: Gaotanyan

country: China

institute: TMMU

type: Expression profiling by high throughput sequencing

type: Non-coding RNA profiling by high throughput sequencing

summary: It is commonly observed that patients with bone fracture concomitated with traumatic brain injury (TBI) had significantly increased fracture healing while the underlying mechanisms were not fully revealed. Long non-coding RNAs (lncRNAs) were known for paly complicated roles in bone homeostasis while its role in TBI accelerated fracture were rarely reported. The present study was designed to determine the role of lncRNAs in TBI accelerated fracture via transcriptome sequencing and further bioinformatical analysis. Blood samples from 3 fracture only patients, 3 fracture concomitated with TIB patients and 3 healthy controls were harvested and were subsequently subjected to transcriptome lncRNAs sequencing. Differentially expression genes were identified, and pathway enrichment were performed by Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyze. High high-dimensional data portraying by self-organizing map (SOM) machine learning was applied to further interpret the data. A xCell method were then used to predict cellular behavior in all samples based on gene expression profiles and a lncRNA-cell interaction network were generated.  A total of 874 differentially expressed genes were identified, of which about 26% were lncRNAs. Those identified lncRNAs were mainly enriched on traumatic brain injury related- and damage repair related-pathways. SOM analyzes revealed that those differentially expressed lncRNAs could be divided into three major module implications and were mainly enriched on transcriptional regulation and immune related signal pathways, which promote us to further explore cellular behaviors based on differentially expressed lncRNAs. We have predicted that basophils, CD8+ T effector memory cells, B cells, and naïve B cells were significantly down-regulated while microvascular endothelia cells were predicated significantly up-regulated with TBI/Fr group was the lowest or highest, respectively. ENSG00000278905, ENSG00000240980, ENSG00000255670, and ENSG00000196634 were the most differentially expression lcnRNA that related to all changes of cellular behavior. The present study has revealed for the first time that several critical lncRNAs may participate in TBI accelerated fracture potentially via regulating cellular behaviors of basophils, cytotoxic T cells, B cells and endothelia cells.

overall_design: Whole blood lncRNAs profile of  healthy controls, fracture only patients, and fracture concomitated with TIB patients

first: Guoning

last: Guo

first: Tinggang

last: Wang


title: IFN-b acts on monocytes to ameliorate CNS autoimmunity by inhibiting proinflammatory cross-talk between monocytes and Th cells

status: Public on May 19 2021

city: PHILADELPHIA

first: Adam

last: Ertel

state: PA

address: 233 S 10TH ST

department: Cancer Biology

country: USA

institute: Thomas Jefferson University

summary: Multiple sclerosis (MS) has been treated with IFN-b for three decades, but the mechanisms underlying its beneficial effects are not well understood. Our study elucidates, in a mouse model of MS, a potential mechanism whereby IFN-b inhibits cooperation between monocytes and T cells, two key immune cell types in MS. We found that IFN-b acts on monocytes to induce secretion of anti-inflammatory molecules, IL-10 and TGF-b, which act on T cells to suppress their production of pro-inflammatory molecules, GM-CSF, TNF, and FASL. Given that these T cell molecules induce proinflammatory features of monocytes, such as IL-1b production, we concluded that IFN-b therapy suppresses cooperation between monocytes and T cells, which otherwise reciprocally amplifies their harmful roles in MS.

overall_design: WT/IFNAR1-/- mix bone marrow chimera mice were generated, immunized for EAE induction, and treated with IFN-b daily. WT and IFNAR1-/- monocytes were isolated and their transcriptome were analyzed by RNA sequencing

first: Javad

last: Rasouli

first: Paolo

last: Fortina

first: Abdolmohamad

last: Rostami

first: Bogoljub

last: Ciric


title: Neural Cell Adhesion Molecule (NCAM-1) is Required for Ventricular Conduction System Development

city: New York City

first: Alireza

last: Khodadadi-Jamayran

state: NY

address: 550 1st Ave, MSB 304

department: Division of Advanced Research Technologies (DART)

institute: New York University, NYU Langone Medical Center

summary: Cardiac Purkinje cells (PCs) comprise the most distal portion of the ventricular conduction system (VCS) and are essential for synchronous activation of the ventricular myocardium. Contactin-2 (CNTN2), a member of the immunoglobulin superfamily cell adhesion molecules (IgSF-CAMs), was previously identified as a marker of the VCS. Through differential transcriptional profiling we discovered two additional highly enriched IgSF-CAMs in the VCS, NCAM-1 and ALCAM.  Immunofluorescence staining showed dynamic expression patterns for each IgSF-CAM during embryonic and early post-natal stages, but ultimately all three proteins became highly enriched in mature PCs. Mice deficient in NCAM-1, but not CNTN2 or ALCAM, exhibited defects in Purkinje cell gene expression and VCS patterning, as well as cardiac conduction disease. Moreover, using ST8sia2 and ST8sia4 knockout mice, we show that inhibition of post-translational modification of NCAM-1 by polysialic acid (PSA) disrupts trafficking of sarcolemmal intercalated disc proteins to Purkinje cell junctional membranes and abnormal expansion of the extracellular space between apposing Purkinje cells. Taken together, our data provide novel insights into the complex developmental biology of the ventricular conduction system.

overall_design: Extracted RNA from Purkinje cells and Ventricular myocytes of male NCAM-1 WT and KO adult mice in triplicate. WT samples used as control

first: Camila

last: Delgado

first: Glenn

last: Fishman


title: Adhesion-mediated mechanosignaling forces mitohormesis

status: Public on May 22 2021

city: Berkeley

first: Greg

last: Timblin

state: CA

department: MCB

institute: UC Berkeley

summary: Mitochondria control eukaryotic cell fate by producing the energy needed to support life and the signals required to execute programmed cell death. The biochemical milieu is known to affect mitochondrial function and contribute to the dysfunctional mitochondrial phenotypes implicated in cancer and the morbidities of ageing. However, the physical characteristics of the extracellular matrix are also altered in cancer and in aging tissues. We demonstrate that cells sense the physical properties of the extracellular matrix and activate a mitochondrial stress response that adaptively tunes mitochondrial function via SLC9A1-dependent ion exchange and HSF1-dependent transcription. Overall, our data indicate that adhesion-mediated mechanosignaling may play an unappreciated role in the altered mitochondrial functions observed in aging and cancer.

overall_design: RNA-seq of human mammary epithelial cells (MCF10As) cultured on soft-to-stiff physiological stiffnesses (5mM glucose), and soft stiffness with low (5mM) vs. high (25mM) glucose.

middle: A


title: Sustained viremia suppression by SHIVsf162p3cn-recalled effector-memory CD8+ T cells after PD1-based vaccination

city: Hong Kong

first: Ming

last: Yue

address: 21 Sassoon Road, Pokfulam

institute: The University of Hong Kong

summary: HIV-1 functional cure requires sustained viral suppression without antiretroviral therapy. While effector-memory CD8+ T lymphocytes are essential for viremia control, few vaccines elicit such cellular immunity that could be potently recalled upon viral infection. Here, we investigated a program death-1 (PD1)-based vaccine by fusion of simian immunodeficiency virus capsid antigen to soluble PD1. Homologous vaccinations suppressed setpoint viremia to undetectable levels in all vaccinated macaques following high-dose intravenous challenge by the pathogenic SHIVSF162P3CN. Poly-functional effector-memory CD8+ T cells were not only induced after vaccination, but were also recalled upon viral challenge for viremia control as determined by CD8 depletion. Vaccine-induced effector memory CD8+ subsets displayed high cytotoxicity-related genes by single-cell analysis. Vaccinees with sustained viremia suppression for over two years responded to boost vaccination without viral rebound. These results demonstrated that PD1-based vaccine-induced effector-memory CD8+ T cells were recalled by AIDS virus infection, providing a potential immunotherapy for functional cure.

overall_design: Single cell RNA sequencing analysis was done by using FACS purified CD8+ T cells isolated from 3 macaques (B01, B02, and B03) in Group B before immunisation, 4 weeks post, and 12 weeks post immunisation.

first: Yik-Chun

last: Wong

first: Lok-Yan

last: Yim

first: Youqiang

last: Song

first: Zhiwei

last: Chen


title: Transcriptional changes during astrocyte maturation in vivo at single cell resolution II

city: London

first: Michael

last: Lattke

address: Hammersmith Hospital Campus, Du Cane Road

department: Brain Sciences

country: United Kingdom

institute: Imperial College London

summary: Analysis of gene expression during striatal astrocyte maturation in vivo in mouse by scRNA-Seq. This series includes a larger dataset with defined genetic background.

overall_design: In vivo astrocytes were purified from male C57BL/6 mice using anti-ACSA2 MACS at indicated ages. Cells were analysed by 10X-Genomics scRNA-Seq. Similar design as GSE152222 (used in preprint Lattke et al., bioRxiv 2020; doi: https://doi.org/ 10.1101/2020.04.28.066043), but larger dataset for revised manuscript, with consistent defined genetic background and male animals)

first: Francois

last: Guillemot

first: Robert

last: Goldstone


title: Engrailed 1 coordinates cytoskeletal re-organization to induce myofibroblast differentiation

city: Hannover

first: Maximilian

last: Fuchs

address: Carl-Neuberg-Str. 1

department: IMTTS

country: Germany

institute: Hanover Medical School

summary: Transforming growth factor-β (TGFβ) is a key mediator of fibroblast activation in fibrotic diseases including systemic sclerosis. Here we show that Engrailed 1 (EN1) is re-expressed in multiple fibroblast subpopulations in the skin of SSc patients. We characterize EN1 as a molecular amplifier of TGFβ signaling in myofibroblast differentiation: TGFβ induces EN1 expression in a SMAD3-dependent manner, and, in turn, EN1 mediates the pro-fibrotic effects of TGFβ. RNA sequencing demonstrates that EN1 induces a pro-fibrotic gene expression profile functionally related to the cytoskeleton organization and ROCK activation. EN1 regulates gene expression by modulating the activity of SP1 and other SP-transcription factors, as confirmed by ChIP-seq experiments for EN1 and SP1. Functional experiments confirm the coordinating role of EN1 on ROCK activity and the re-organization of cytoskeleton during myofibroblast differentiation both in standard fibroblast culture systems and in in vitro skin models. Consistently, mice with fibroblast-specific knockout of En1 demonstrate impaired fibroblast-to-myofibroblast transition and are partially protected from experimental skin fibrosis.

overall_design: Gene expression profile of human dermal fibroblasts stimulated with TGFβ with siRNA-induced knockdown of EN1, with fibroblasts stimulated with TGFβ and treated with non-targeting siRNA as controls

first: Andrea-Hermina

last: Györfi

first: Alexandru-Emil

last: Matei

first: Chunguang

last: Liang

first: Meik

last: Kunz

first: Jörg

middle: H

last: Distler


title: ATAC-seq_experiment in RKO cell line under low replication stress

city: Toulouse cedex 1

first: Valérie

last: Bergoglio

address: 2 avenue Hubert Curien

institute: INSERM UMR1037

type: Genome binding/occupancy profiling by high throughput sequencing

summary: RKO cells were treated with low doses of aphidicolin (0.2µM) that inhibit replicative DNA polymerases and induce a mild replication stress. ATAC-seq data were analysed on control cells (DMSO) and after 16h of treatment with aphidicolin

overall_design: We used ATAC-seq approach to analyse the impact of low replication stress on chromatin accessiblity comparing DMSO or aphidicolin-treated cells.

first: Valerie


title: Microarray gene expression in RKO cell line under low replication stress

summary:
RKO cells were treated with low doses of aphidicolin (0,2µM) that inhibit replicative DNA polymerases and induce a mild replication stress. Expression data were analysed on control cells (DMSO), 16h of treatment (t0) and after release in complete new culture medium of 13h (N+1) using microarray (affymetrix Clarion S)
We used microarray to analyse the impact of low replication stress on gene expression comparing DMSO or aphidicolin-treated cells at the end of the treatment (t0) and in daughter cells released from the replication stress (N+1).

overall_design: Triplicates of total RNA was extracted  from the same amount of exponentially growing RKO cells in different conditions (DMSO or aphidicolin, t0 or N+1) and hybridized on Affymetrix Microarrays.


title: CRISPR-based targeting of DNA methylation in Arabidopsis thaliana by a bacterial CG specific DNA methyltransferase.

city: Los Angeles

first: Colette

middle: L

last: Picard

state: California

address: 610 Charles E Young Dr East, Room 4045

department: Molecular, Cell and Developmental Biology

institute: University of California - Los Angeles

type: Methylation profiling by high throughput sequencing

summary: CRISPR-based targeted modification of epigenetic marks is an important strategy to regulate the expression of genes and their associated phenotypes in plants and animals. The manipulation of DNA cytosine methylation is particularly attractive in plants since DNA methylation changes are often heritable in subsequent plant generations. Although plants have DNA methylation in all sequence contexts (CG, CHG, CHH, where H can be any nucleotide but G), methylation at symmetrical CG sites is the most important for gene silencing, and is also the most efficiently maintained through miotic and meiotic cell divisions. Thus, DNA methylation targeting tools that directly install CG methylation are highly desirable. Here we have developed two CRISPR-based CG specific targeted DNA methylation systems for plants using the DNA methyltransferase MQ1 (SssI) from the bacterium Mollicutes spiroplasma. To reduce off-target effects, we used a variant of the enzyme with reduced catalytic activity. We demonstrate that methylation added by MQ1 is highly target-specific and can be heritably maintained in the absence of the effector. These tools should be valuable both in crop engineering and in plant genetic research.

overall_design: This study evaluated two new CRISPR-based tools, MQ1v and SunTag-MQ1v, designed to target de novo DNA methylation in plants using MQ1, a CG-specific bacterial methyltransferase. These tools were targeted to the promoter of the gene FWA using three previously-characterized guide RNAs (Papikian et al. 2019).  MQ1v and SunTag-MQ1v were targeted to the FWA promoter in fwa epimutants to test whether they could establish DNA methylation over the promoter and affect FWA expression. Control plants were transformed with dMQ1, a version of MQ1v that has no catalytic activity. MQ1v efficiency was also evaluated in the absence of functional transcriptional (drm1 drm2 background) and post-transcriptional gene silencing (rdr6 background). Transformed plants and their descendants were evaluated by assessing FWA expression (qPCR or RNA-seq), and DNA methylation at the FWA promoter (bisulfite-PCR followed by sequencing, or BS-PCR) and genome-wide (bisulfite sequencing, pr BS-seq). BS-seq and/or BS-PCR was performed for individual transformed plants (T1) and/or their descendants (T2, T3, or T4, corresponding to second-, third-, and fourth-generation descendants of the T1 plants, respectively), as well as appropriate dMQ1, fwa and Col (wild-type) controls. RNA-seq experiments were performed in T4 plants in triplicate.

first: Basudev

last: Ghoshal

first: Brandon

last: Vong

first: Suhua

last: Feng

first: Steven

middle: E

last: Jacobsen


title: Immunological and pathological landscape of dengue serotypes 1-4 infections in immune-competent mice

city: Riverside

first: Juliet

last: Morrison

address: 900 University Avenue

department: Microbiology and Plant Pathology

institute: University of California, Riverside

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

overall_design: Refer to individual Series


title: Immunomodulatory effects of vitamin D supplementation in a deficient population

city: Farmington

first: Mohammed

last: Toufiq

state: Connecticut

address: 10 Discovery Drive, Farmington, CT 06032

department: Research Department

institute: The Jackson Laboratory for Genomic Medicine

summary:
Background: In addition to its canonical functions, vitamin D has been proposed to be an important mediator of the immune system. Despite ample sunshine, vitamin D deficiency is prevalent (> 80%) in the Middle East, resulting in a high rate of supplementation. However, the underlying molecular mechanisms of the specific regimen and potential factors affecting an individual’s response to vitamin D are not well characterized.
Objective: To characterize changes in blood transcriptomic and the potential mechanisms associated with vitamin D3 supplementation and response.
Design: In this intervention study, one hundred vitamin D-deficient women were given a weekly oral dose (50,000 IU) of vitamin D3 for three months. A high-throughput targeted PCR, composed of 264 genes representing important blood transcriptomic fingerprints in health and disease states, was performed on pre- and post-supplementation blood samples to profile the molecular response to vitamin D3. Multivariate, network, gene ontology, and literature mining analyses were used for the interpretation of the transcriptomic profiling results.
Results: We identified 54 differentially expressed genes that were strongly modulated by vitamin D3 supplementation. Network analyses showed significant changes in the immune-related pathways such as TLR4/CD14 and IFN receptors, and catabolic processes related to NFkB, which were subsequently confirmed by gene ontology enrichment analyses. We proposed a model for vitamin D3 response, using the reduced expression of the molecules involved and the receptor-mediated intra-cellular signaling leading to reduce cytokine production.
Conclusions: Blood-transcriptomic profiles of vitamin D3 response were generated using a targeted blood gene panel. Vitamin D has a strong effect on the immune system, G-coupled protein receptor signaling, and the ubiquitin system. We highlighted the major molecular changes and biological processes induced by vitamin D3, which will help to further investigate the effectiveness of vitamin D supplementation among individuals in the Middle East.

first: Mathieu

last: Garand

first: Parul

last: Singh

first: Susie

middle: S

last: Huang

first: Sara

last: Tomei

first: Rebecca

last: Mathew

first: Valentina

last: Mattei

first: Mariam

last: Al Wakeel

first: Elham

last: Sharif

first: Souhaila

last: Al Khodor


title: Regulation of alpha-like globin gene expression in murine primitive erythropoiesis (ChIP-seq)

status: Public on May 18 2021

city: Headington

first: Damien

last: Downes

state: Oxfordshire

address: John Radcliffe Hospital

department: MRC Weatherall Institute of Medicine

institute: The University of Oxford

summary: Haemoglobin, the oxygen-carrying molecule of red blood cells, at all stages of development comprises a tetramer of two α-like and two β-like globin chains. In humans and mice the highly conserved α- and β-globin loci contain genes encoding globin chains specifically expressed only during the first trimester of gestation (termed embryonic globins) in addition to the genes encoding globin chains expressed throughout adult life. In humans the β-globin locus also encodes a β-like chain that is expressed throughout the second and third trimesters of development; this is termed γ-globin and complexes with α-globin to form fetal haemoglobin (HbF). The sequential activation and repression of the genes at each of these loci throughout development to maturity is termed haemoglobin switching and is a poorly understood process. α-thalassemia, which results from insufficient production of α-globin, is a major global health problem with several hundred thousand sufferers world-wide. The embryonically expressed ζ-globin, encoded by the gene HBZ (Hba-x in mice), can functionally substitute for α-globin in adult erythroid cells. Reactivation of this gene is consequently likely to ameliorate the symptoms of α-thalassemia in individuals with a severe phenotype. Little is currently known about the regulation of ζ-globin expression in terms of both the trans- and cis-acting regulatory elements responsible for high-levels of expression of this gene during embryonic erythropoiesis and its maintenance in a transcriptionally inactive state throughout adult life. Using ATACseq, ChIP-seq and NG Capture C and mutant mouse lines this work aims to identify and define the cis-acting regulatory network underlying zeta-globin expression, and define the contribution of each regulatory element.

overall_design: ChIP-seq of epigenetic marks associated with enhancers (H3K4me1), promoters (H3K4me3), active transcription (H3K27ac) and polycomb repression (H2A1119Ub) as well as the boundary transcription factor (CTCF) and the erythroid transcription factor, Gata1.

middle: J

first: Andrew

last: King

first: Siyu

last: Liu

first: Chris

last: Babbs


title: Gene expression profiling of lymph node sub-capsular sinus macrophages in cancer

city: Portland

first: Ferdinando

last: Pucci

address: 2720 S Moody ave

department: Otolaryngology

institute: Oregon Health&Science University

summary: Lymph nodes are key lymphoid organs collecting lymph fluid and migratory cells from the tissue area they survey. When cancerous cells arise within a tissue, the sentinel lymph node is the first immunological organ to mount an immune response. Sub-capsular sinus macrophages (SSMs) are specialized macrophages residing in the lymph nodes that play important roles as gatekeepers against particulate antigenic material. In the context of cancer, SSMs capture tumor-derived extracellular vesicles (tEVs), a form of particulate antigen released in high amounts by tumor cells. We and others have recently demonstrated that SSMs possess anti-tumor activity because in their absence tumors progress faster. A comprehensive profiling of SSMs represents an important first step to identify the cellular and molecular mechanisms responsible for SSM anti-tumor activity. Unfortunately, the isolation of SSMs for molecular analyses is very challenging. Here, we combined an optimized dissociation protocol, careful marker selection and stringent gating strategies to highly purify SSMs. We provide evidence of decreased T and B cell contamination, which allowed us to reveal the gene expression profile of this elusive macrophage subset. Squamous cell carcinomas induced an increase in the expression of Fc receptors, lysosomal and proteasomal enzymes in SSMs. Imaging of mouse and patient lymph nodes confirmed the presence of the top differentially expressed genes. These results suggest that SSMs respond to tumor formation by upregulating the machinery necessary for presentation of tumor particulate antigens to B cells.

overall_design: 16 samples, 2 technical replicates, 2 biological replicates, 2 populations, 2 tissues

first: Danilo

last: Pellin

first: Natalie

last: Claudio


title: Multimeric nanobodies from camelid mice and llamas neutralize SARS-CoV-2 variants

city: bethesda

first: Seolkyoung

last: Jung

state: MD

address: 10 Center Dr

department: NIAMS

institute: NIH

summary: Since the start of the coronavirus disease-2019 (COVID-19) pandemic, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has caused more than 2 million deaths worldwide. Many vaccines have been deployed to date; however, the continual evolution of the viral receptor binding domain (RBD) has recently challenged their efficacy. In particular, SARS-CoV-2 variants originating in South Africa (B.1.351) and the U.K. (B.1.1.7) have reduced plasma neutralization activity and crippled antibody cocktails that received emergency use authorization1-3. Whereas vaccines can be updated periodically to account for emerging variants, complementary strategies are urgently needed to overcome viral escape. One potential alternative are camelid VHHs (also known as nanobodies), which can access conserved epitopes often hidden to conventional antibodies4-6. We here isolate anti-RBD nanobodies from llamas and mice engineered to produce VHHs from alpacas, dromedaries and camels. Through neutralization assays and cryo-electron microscopy we identify two “nanomouse” VHHs that circumvent RBD antigenic drift by recognizing a domain conserved in coronaviruses, away from the ACE2 binding motif. Conversely, llama nanobodies recognize the RBD-ACE2 interphase and as monomers they are ineffective against E484K or N501Y substitutions. Notably, as homotrimers those same VHHs neutralize RBD variants with ultrahigh (pM) affinity, rivaling the most potent antibodies produced to date against SARS-CoV-2. We conclude that multivalent nanobodies can avert SARS-CoV-2 escape mutants and thus they represent promising tools to prevent COVID-19 mortality when vaccines are compromised.

sample_organism:
Lama glama
Mus musculus

platform_organism:
Lama glama
Mus musculus

overall_design: VHH gene sequence from llamas and camelid mice

first: Jianliang

last: Xu

first: Kai

first: Andrea

last: Conte

first: Jenna

last: Lieberman

first: Frauke

last: Muecksch

first: Julio

middle: C

last: Cetrulo Lorenzi

first: Gwo-Yu

last: Chuang

first: Solji

last: Park

first: Zijun

last: Wang

first: Tyler

last: Stephens

first: Lino

last: Tessrollo

first: Paul

middle: D

last: Bieniasz

first: Tatsiana

last: Bylund

first: Theodora

last: Hatzioannou

first: Michel

last: Nussenzweig

first: Peter

last: Kwong

first: Rafael

last: Casellas


title: RNA-Sequencing of  spinach tissues with contrasting oxalate contents

city: Uvalde

first: Vijay

last: Joshi

state: Texas

address: 1619 Garner Field Road

department: Department of Horticultural Sciences

institute: Texas A&M AgriLife Research

summary: Purpose: The goals of this study are to compare differentially expressed transcripts in roots and leaves of spinach cultivars with distinct oxalate contents using transcriptome profiling (RNA-seq)

overall_design: Methods: Leaf and root samples for RNA Seq analysis were collected from three independent spinach plants of cultivar PI175311 and Bloomsdale were flash-frozen in liquid nitrogen for further analysis. Results: The clustering of the index-coded samples was performed on a cBot Cluster Generation System using PE Cluster Kit cBot-HS (Illumina) according to the manufacturer’s instructions. After cluster generation, the libraries were sequenced on an Illumina Hiseq platform, and 150 bp paired-end reads were generated. Raw reads of fastq format were processed to obtain clean reads by removing the adapter, reads containing ploy-N, and low quality reads from raw data. At the same time, Q20, Q30, and GC content, the clean data were calculated. Reference genome and gene model annotation files were downloaded from SpinachBase (http://spinachbase.org/). Index of the reference genome was built using Bowtie v2.2.3, and paired-end clean reads were aligned to the reference genome using TopHat v2.0.12. : HTSeq v0.6.1 was used to count the reads mapped to each gene. Reads Per Kilobase of exon per Megabase of library size (RPKM) were calculated from mapped read by featureCounts. Differential expression analysis of High N and Low N conditions (three biological replicates per tissue per treatment) was performed using the DESeq R package (1.18.0) (Anders and Huber, 2010). Genes with P-value < 0.05 found by DESeq were assigned as differentially expressed. Conclusions: The transcriptomic changes in leaf and root tissues was used for RNA-Seq analysis. A total of six libraries from leaf and root tissue were sequenced for each cultivar using the Illumina HiSeq platform. On average, 60 and million raw reads were generated from leaf tissues for bothe the cultivars. Across all reads , the Q20 and Q30 percentage was more than 98 and 94%, respectively (sequencing error rate was less than 0.02%), and GC content for the libraries was ~43%.  Among all the libraries, the ratio of total mapped reads was above 85%, of which ~81 % reads uniquely mapped to the reference genome . The data generated from all libraries provided a foundation for quality analyses.


title: IL-22 signaling attenuates necrotizing enterocolitis by promoting epithelial cell regeneration

city: Chapel Hill

first: Misty

last: Good

state: NC

address: 101 Manning Drive 4th Floor CB 7596

department: Pediatrics

institute: University of North Carolina at Chapel Hill

summary: Necrotizing enterocolitis (NEC) is a devastating intestinal inflammatory disorder that primarily affects premature infants. Despite decades of research, this disease remains a significant cause of death in the absence of efficient therapeutics. Interleukin (IL)-22 has been shown to play a critical role in maintaining the gut barrier, promoting epithelial regeneration, and controlling intestinal inflammation in adult animal models with an established microbiome. However, the importance of IL-22 signaling in the regulation of gut homeostasis and protection in neonates that lack an established microbiome remains unknown. Therefore, the aim of the current study is to investigate the role of IL-22 in the neonatal intestinal epithelium under homeostatic and inflammatory conditions by using a mouse model of NEC. Our data reveal that Il22 expression in neonatal murine intestine is negligible until weaning. In addition, both human and murine neonates lack IL-22 production during NEC. Mice deficient in IL-22 or mice lacking the expression of IL-22 receptor in intestinal epithelial cells, display a similar susceptibility of neonates to NEC consistent with the lack of endogenous IL-22 at this critical stage of intestinal development. Conversely, treatment with recombinant IL-22 during NEC substantially reduces disease severity. This IL-22-mediated protection is associated with enhanced epithelial regeneration and increased expression of several antimicrobial genes. Strikingly, despite an IL-22-mediated induction of an antimicrobial transcriptional program, the composition of the intestinal microbial communities remains unchanged. Taken together, this study demonstrates that an IL-22 signaling axis promotes protection against neonatal NEC through the induction of epithelial cell regeneration.

overall_design: The aim of the current study was to investigate the role of IL-22 in the neonatal intestinal epithelium under homeostatic and inflammatory conditions by using a mouse model of NEC. All included samples are enteroids from 6 week control (non-NEC) mice.

first: Belgacem

last: Mihi

first: Qingqing

last: Gong

first: Lila

last: Nolan

first: Sarah

last: Gale

first: Martin

last: Goree

first: Elise

last: Hu

first: Wyatt

last: Lanik

first: Jamie

middle: M

last: Rimer

first: Victoria

last: Liu

first: Olivia

middle: B

last: Parks

first: Angela

middle: N

last: Lewis

first: Pranjal

last: Agrawal

first: Marie

last: Laury

first: Pawan

last: Kumar

first: Elizabeth

last: Huang

first: Shay

last: Bidani

first: Cliff

last: Luke

first: Jay

middle: K

last: Kolls


title: Expression data from umbilical cord blood and peripheral blood stem cell-derived monocytes of human

city: Hefei

first: Lin

middle: lin

last: Jin

address: 443 Huangshan Road

institute: University of science and technology of China

summary: To identify  the differences between human umbilical cord blood and peripheral blood monocytes, we performed unsupervised bioinformatic analyses by microarrays.

overall_design: monocytes were isolated from human cord blood mononuclear cells and peripheral blood stem cells at 6 hours after in vitro culture,  three independent experiments were performed using different donors for each experiment.

first: Linlin

first: Haiming

last: Wei


title: Multiplexed functional genomic analysis of somatic 5' untranslated region mutations across the spectrum of human prostate cancer (RNASeq)

status: Public on May 17 2021

city: Seattle

first: Ilsa

last: Coleman

state: WA

address: 1100 Fairview Ave N, E2-112

department: Human Biology

institute: Fred Hutchinson Cancer Center

summary: The functional consequences of genetic variants within 5’ untranslated regions (UTRs) on a genome-wide scale are poorly understood in disease. We developed a high-throughput multi-layer massively parallel sequencing-based method called PLUMAGE (Pooled full-length UTR Multiplex Assay on Gene Expression) to quantify the molecular consequences of somatic 5’ UTR mutations found across 229 prostate cancer patients with localized to metastatic disease. We show that 5’ UTR mutations can control both transcript levels and mRNA translation rates through the creation of new DNA binding elements or RNA-based cis-regulatory motifs. We also discover that single point mutations can simultaneously impact both transcript levels and mRNA translation of the same gene. Using gene editing technology, we validate that a single point mutation in the oncogenic CKS2 5’ UTR can increase mRNA specific translation. Turning to a molecular pathway critical for cancer, we provide evidence that functional 5’ UTR mutations in the MAP kinase signaling pathway can upregulate pathway-specific gene expression and are associated with distinct clinical outcomes. Our study reveals the diverse mechanisms by which the mutational landscape of 5’ UTRs can co-opt multiple levels of gene expression and demonstrates that single nucleotide alternations within leader sequences are functional in cancer.

overall_design: Next-generation sequencing of metastatic castration-resistant prostate cancer (mCRPC) tumors using Illumina TruSeq Stranded mRNA Library prep.

first: Yiting

last: Lim

first: Sonali

last: Arora

first: Samantha

last: Schuster

first: Lukas

last: Corey

first: Matthew

last: Fitzgibbon

first: Cynthia

last: Wladyka

first: Xiaoying

last: Wu

first: Jeffrey

last: Delrow

first: Eva

first: Lawrence

last: True

first: Peter

last: Nelson

first: Gavin

last: Ha

first: Andrew

last: Hsieh


title: Low-energy electron irradiation as a potent alternative to gamma irradiation for the inactivation of (CAR-)NK-92 cells in ATMP manufacturing

city: Leipzig

first: Kristin

last: Reiche

address: Perlickstr. 1

institute: Fraunhofer Institute for Cell Therapy and Immunology

summary: NK-92 cells and their CAR-modified derivatives exhibit strong cytotoxic activity against tumors and are promising "off-the-shelf" therapeutics compared to CAR-T cells. In order to ensure safety and prevent the occurrence of secondary tumors, (CAR-)NK-92 cells need to be inactivated before transfusion. With increasing clinical use, there is a growing demand for enhanced safety and an efficient method that stops cell proliferation, but better maintains the cytotoxic effector functions of the cells compared to commonly used gamma irradiation. Recently, low-energy electron irradiation (LEEI) was successfully applied for inactivation of bacteria and viruses for vaccine production, and was described as a method for NK-92 inactivation for the first time. In the present publication, data on extensive characterization of LEEI and its comparison with gamma irradiation for the inactivation of parental NK-92 cells as well as CD123-directed CAR-NK-92 cells are provided. Our results show that both irradiation methods cause a progressive decrease in cell viability and are therefore suitable for inhibition of proliferation. Notably, cytotoxicity of the NK cells was significantly reduced by gamma irradiation, but not by LEEI three days after irradiation, compared to non-irradiated cells. Both gamma irradiation as well as LEEI led to substantial DNA-damage and an accumulation of irradiated cells in the G2/M phase. In addition, transcriptomic analysis of cells irradiated with both methods revealed approximately 12-fold more differentially expressed genes after gamma irradiation compared to LEEI. Analysis of surface molecules revealed an irradiation-induced decrease in CD56 expression but no changes in the levels of the activating receptors NKp46, NKG2D, and NKp30. Conclusions: The present data show that LEEI efficiently inactivates (CAR )NK-92 cells and sustains their cytotoxic capacity better than gamma irradiation. Taking into account additional logistic advantages of LEEI proves a potent alternative in the manufacturing of (CAR-)NK-92 cells for clinical application.

overall_design: RNA expression of three LEEI-irradiated, three gamma-irradiated and three non-irradiated (control) CAR-NK cell samples

first: Lia

last: Walcher

first: Ann-Kathrin

last: Kistenmacher

first: Charline

last: Sommer

first: Sebastian

last: Böhlen

first: Christina

last: Ziemann

first: Susann

last: Dehmel

first: Armin

last: Braun

first: Uta

last: Kossatz-Böhlert

last: Tretbar

first: Stephan

last: Klöß

first: Axel

last: Schambach

first: Michael

last: Morgan

first: Dennis

last: Löffler

first: Conny

last: Blumert

first: Christoph

last: Kämpf

first: Jana

last: Beckmann

first: Ulla

last: König

first: Bastian

last: Standfest

first: Martin

last: Thoma

first: Gustavo

middle: R

last: Makert

last: Ulbert

first: Anna

last: Dünkel

first: Ulrike

last: Köhl

last: Fricke


title: ATRX promotes heterochromatin formation to protect cells from G-quadruplex DNA-mediated stress [CUT&Tag]

city: Dallas

first: Laura

last: Banaszynski

state: TX

address: 5323 Harry Hines Blvd

department: Green Center for Reproductive Biology Sciences

institute: UT Southwestern Medical Center

summary: We analyze G-quadruplex enrichment in ATRX KO mESC

overall_design: BG4 CUT&Tag in mouse embryonic stem cell lines (WT and ATRX KO)

first: Yu-Ching

last: Teng

first: Aishwarya

last: Sundaresan


title: BANP opens chromatin and activates CpG island regulated genes [RNA-Seq]

city: Basel

first: Dirk

last: Schuebeler

address: Fabrikstrasse 24

country: Switzerland

institute: Friedrich Miescher Institute for Biomedical Research

summary: In mammalian genomes, the vast majority of RNA polymerase II initiation events take place at CpG island promoters. Despite their relevance our understanding of their regulation remains limited. Here we identify Banp as the long sought-after TF that binds the orphan CGCG element in CpG islands by combining single-molecule footprinting with interaction proteomics. We show that Banp drives activity of CpG islands that control essential metabolic genes in the mouse and human genome. Banp binding is strongly repelled by DNA methylation of its motif in vitro and in vivo, which restricts most binding to CpG islands and accounts for its absence at aberrantly methylated CpG islands in cancer cells. Upon binding to an unmethylated motif, Banp opens chromatin and positions nucleosomes. These findings expand our understanding of CpG island gene regulation and put forth a model whereby CpG islands rely for their activity on methylation sensitive TFs capable of opening and organizing chromatin.

overall_design: RNA-seq in mouse ESCs and neurons: ESC wt, DNMT triple-ko (tko) and tko dTag untreated and treated for 1h, 2h, 4h and 6h, neurons  dTag untreated and treated for 1h, 2h, 4h and 6h. All conditions in triplicates, except for ESC wt and tko (6 replicates) and ESC tko dTag 1h (2 replicates). Batch 1: ESC wt replicates 1-3 and ESC tko replicates 1-3. Batch 2: ESC tko replicates 4-6 and ESC tko degron time course. Batch 3: ESC wt replicates 4-6, ESC with Ngn2 construct uninduced and all neuron samples.

first: Ralph

last: Grand

first: Lukas

last: Burger

last: Schubeler


title: BANP opens chromatin and activates CpG island regulated genes [ATAC-Seq]

overall_design: ATAC-seq in mouse ESCs. Wt, dTag untreated, dTag treated for 1h, 2h, 4h, 6h and 20h. All conditions in duplicates.


title: BANP opens chromatin and activates CpG island regulated genes [MNase-seq]

overall_design: MNase-seq in mouse ESCs. dTag untreated, 1h treated and 4h treated. All conditions in duplicates.


title: BANP opens chromatin and activates CpG island regulated genes [SLAM-Seq]

overall_design: SLAM-seq in mouse ESCs. Wt (s4U untreated and treated). dTag untreated and dTag treated for 1h, 2h, 4h, 6h and 20h (all s4U treated). All conditions in triplicates, except dTag untreated (duplicates).


title: ATRX promotes heterochromatin formation to protect cells from G-quadruplex DNA-mediated stress [ChIP-Seq]

summary: We analyze ATRX enrichment in ATRX-GFP and FACT subunits KD mESC

overall_design: ATRX ChIP-seq in mouse embryonic stem cell lines (ATRXWT, ATRX KO, ATRX-GFP, Control, SSRP1KD and SPT16 KD). All inputs sequenced.


title: ATRX promotes heterochromatin formation to protect cells from G-quadruplex DNA-mediated stress [EdU-seq]

summary: We analyze the contribution of ATRX and DAXX to G-quadruplex DNA replication in mESC

overall_design: EdU-seq analysis in mouse embryonic stem cells (WT, ATRX KO, DAXX KO)


title: BANP opens chromatin and activates CpG island regulated genes [ChIP-Seq]

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: ChIP-seq in mouse ESCs, neurons and human HCT116 and HCC1954. 3 replicates Banp IP/IgG control in mouse ESCs wt, in DNMT tko ES cells, in ESCs with Ngn2 construct uninduced and in Ngn2-induced neurons. Duplicates in both HCT116 and HCC1954 cells.


title: Replication timing data for six human cell lines under aphidicolin treatment (t0) and after release (N+1)

first: Valérie

last: Bergoglio

summary: DNA replication is very well orchestrated in mammalian cells due to a tight regulation of the temporal order of replication origin activation, known as the replication timing program. The replication timing of a given replication domain is very robust and well conserved in each cell type. Upon low replication stress, the slowing of replication forks induces delayed replication of some fragile regions leading to DNA damage and genetic instability. Except for these fragile regions, the direct impact of low replication stress on the replication timing in different cellular backgrounds has not been explored in detail. Here we analysed DNA replication timing across the whole genome in a panel of human cell lines in the presence of low replication stress. We demonstrate that low replication stress induced by aphidicolin has a stronger impact on the replication timing of cancer cells than non-tumour cells. Strikingly, we unveiled an enrichment of specific replication domains undergoing a switch from late to early replication in some cancer cells. We found that advances in replication timing correlate with heterochromatin regions poorly sensitive to DNA damage signalling while being subject to an increase of chromatin accessibility in response to aphidicolin. Finally, our data indicate that, following release from replication stress conditions, replication timing advances can be inherited by the next cellular generation, suggesting a new mechanism by which some cancer cells would adapt to cellular or environmental stress.

overall_design:
The 6 human cell lines were treated with aphidicolin 0.2µM or DMSO during adapted duration. T0 correspond to the replication timing of cells under the treatment and N+1 correspond to the replication timing of cells released from the drug during appropriate duration.
Biological replicates for each conditions (37 samples)

first: Jean-Charles

last: Cadoret


title: Iron-dependent apoptosis causes embryotoxicity in inflamed and obese pregnancy

first: Allison

last: Fisher

address: 650 Charles E Young Dr South, CHS 44-100

department: Medicine

institute: UCLA Center for Iron Disorders

summary: Because iron is essential for a healthy pregnancy, iron supplementation during pregnancy is nearly universally recommended, regardless of maternal iron status. A signal of potential harm is the U-shaped association between maternal ferritin, a marker of iron stores, and risk of adverse pregnancy outcomes. However, ferritin is also induced by inflammation and so may overestimate iron stores during inflammation or infection. We used mouse models to determine whether maternal iron loading, inflammation, or their interaction caused poor pregnancy outcomes. Only maternal exposure to both iron excess and inflammation, but not either condition alone, caused embryo malformations and demise. Maternal iron excess potentiated embryo injury during both LPS-induced acute inflammation and obesity-induced chronic mild inflammation. The adverse interaction was dependent on TNFα signaling, caused apoptosis of the placental and embryo endothelium, and was prevented by anti-TNFα or antioxidant treatment in vivo. Our findings raise important questions about the safety of indiscriminate iron supplementation during pregnancy.

overall_design: Endothelial cells were isolated from placentas from pregnant wildtype (C57BL/6) and hepcidin-1 knockout (on the C57BL/6 background) dams after treatment with water (baseline) or 0.5ug/g lipopolysaccharide (LPS) on E15.5 for 6h. For each condition, we pooled endothelial cells from 5 placentas/pregnant mouse, from 4 different mice per group. For each group we have 4 biological replicates. The total number of samples is 16. Total RNA from placental endothelial cells was isolated and subjected to RNA-Seq.

first: Veena

last: Sangkhae

first: Kamila

last: Balušíková

first: Nicolaos

last: Palaskas

first: Tomas

last: Ganz

first: Elizabeta

last: Nemeth


title: ATRX promotes heterochromatin formation to protect cells from G-quadruplex DNA-mediated stress

overall_design: Refer to individual Series


title: Multiplexed functional genomic analysis of somatic 5' untranslated region mutations across the spectrum of human prostate cancer (Ribosome Profiling)

first: Sonali

last: Arora

address: 1100 Fairview Ave N,

institute: FHCRC

summary: The functional consequences of cancer patient-derived genetic variants within the 5’ untranslated regions (UTRs) on a genome-wide scale and their effects on mRNA transcript and translation levels are poorly understood. To systematically interrogate the mutational landscape of 5’ UTRs across cancer patients with localized to metastatic disease, we analyzed the genomes of 226 prostate cancer patients and observed thousands of mutations, many of which impact known cis-regulatory elements. We developed a high-throughput multi-layer massively parallel sequencing-based method called PLUMAGE (Pooled full-length UTR Multiplex Assay on Gene Expression) to simultaneously quantify the effects of 545 5’ UTR somatic mutations across recurrently mutated or cancer-related genes from our patient cohort. Our method enabled unprecedented insights into how 5’ UTR mutations can control multiple levels of gene expression simultaneously. In particular, we identified 190 mutations that significantly altered 5’ UTR function, either by creating new DNA binding elements, disrupting known translation regulatory motifs, or simultaneously impacting both transcript levels and mRNA translation. Furthermore, we also determined that 5’ UTR mutations to the MAP kinase signaling pathway are significantly associated with early metastasis. This study is the first to comprehensively interrogate the functional 5’ UTR mutational landscape of a human cancer revealing the importance of untranslated regions in regulating multiple levels of oncogenic gene expression and provides a high-throughput functional genomics solution applicable to many genetically driven diseases.

overall_design: To investigate the functional impact of 5’ UTR mutations in advanced prostate cancer, we will conduct whole exome sequencing utilizing new 5’ UTR exon capture technology on a cohort of end stage castration resistant prostate cancer patient derived xenografts (PDXs). Derived from rapid autopsy specimens, these PDX models have never been cultured, faithfully recapitulate human disease, and represent a nearly inexhaustible supply of tissues for biological replicates. A number of 5’ UTR somatic mutations, insertions, and deletions were identified. To directly correlate these findings with the translation of specific transcripts genome-wide, we will perform ribosome profiling on biological replicates of the same PDXs. Normal human prostate tissue will also be profiled as a comparison to expression in prostate cancer. This will be the very first functional analysis of the 5’ UTR regulome in a human cancer.

first: Yiting

last: Lim

first: Andrew

last: Hsieh


title: Gene expression changes accompanying vascular tortuosity

status: Public on May 15 2021

city: Hirosaki

first: Toshio

last: Fumoto

state: Aomori

address: 5 Zaifu-cho

department: Department of Neurosurgery

institute: Hirosaki University Graduate School of Medicine

summary: To gain insight into vascular tortuosity, we analyzed gene expression in normal and tortuous vessels. We induced vascular tortuosity in basilar artery using bilateral common carotid artery ligation in One-year old SD rats. Four weeks after the ligation, total RNA was extracted. Equal amounts of RNA from four rats were pooled and 50 ng RNA was subjected to microarray analysis. Pooled RNA from four Sham operated rats were used as a control.

overall_design: We analyzed 1  control sample  and 1 ligated sample using the Clariom S assy.

first: Hiroki

last: Ohkuma


title: Transcriptome analysis of TLR2-activated Th17 cells

city: North Chicago

first: Joseph

last: Reynolds

state: IL

address: 3333 Green Bay Road

department: Microbiology and Immunology

institute: Rosalind Franklin University of Medicine and Science

summary: Naïve CD4, Th17 (TGFB+IL6), Th17 (TGFb+IL6) + PAM3CSK4, and Th17 (TGFB+IL6) + biglycan samples were sequenced after 3d differentiation

overall_design: We report RNA sequencing results for Th17 cells stimulated through the TLR2 pathway. Naïve CD4+ T cells were sorted, TCR was activated, and cells were polarized with TGFb + IL-6 in the absence or presence of 2ug/ml PAM3CSK4 or biglycan

first: Gustavo

last: Martinez


title: ATRX promotes heterochromatin formation to protect cells from G-quadruplex DNA-mediated stress [ATAC-Seq]

first: Laura

last: Banaszynski

summary:
We analyze the contribution of ATRX, DAXX and HIRA to the chromatin accessibility at G-quadruplex in mESC
We analyze the contribution of ATRX and ESET to the chromatin accessibility at G-quadruplex in mESC

overall_design:
ATAC-seq analysis in mouse embryonic stem cell lines (WT, ATRX KO, DAXX KO, HIRA KO).
ATAC-seq analysis in mouse embryonic stem cell lines (ATRX addback into ATRX KO, and ESET KO).

first: Yu-Ching

last: Teng

first: Aishwarya

last: Sundaresan


title: Multiplexed functional genomic analysis of somatic 5’ untranslated region mutations across the spectrum of human prostate cancer

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


title: Extrafollicular IgD+ B cells Generate IgE Antibody Secreting Cells in the Nasal Mucosa

status: Public on May 13 2021

city: Pittsburgh

first: Richard

middle: Paul

last: Ramonell

address: 5859 Phillips Ave

department: Department of Medicine

institute: University of Pittsburgh

summary: Here we describe the dynamics underlying the generation of IgE-antibody secreting cells (ASC) in human nasal polyps (NP), mucosal tissues rich in ASC without germinal centers (GC). Using VH next generation sequencing, we identified an extrafollicular (EF) mucosal IgD+ naïve-like intermediate B cell population with high connectivity to the mucosal IgE ASC. Mucosal IgD+ B cells, express germline epsilon transcripts and predominantly co-express IgM. However, a small but significant fraction co-express IgG or IgA instead which also show connectivity to ASC IgE. Phenotypically, NP IgD+ B cells display an activated profile and molecular evidence of BCR engagement. Transcriptionally, mucosal IgD+ B cells reveal an intermediate profile between naïve B cells and ASC. Single cell IgE ASC analysis demonstrates lower mutational frequencies relative to IgG, IgA, and IgD ASC consistent with IgE ASC derivation from mucosal IgD+ B cell with low mutational load. In conclusion, we describe a novel mechanism of GC-independent, extrafollicular IgE ASC formation at the nasal mucosa whereby activated IgD+ naïve B cells locally undergo direct and indirect (through IgG and IgA), IgE class-switch.

overall_design: Examination of the origin of IgE ASCs by analysis of characteristics of NP-resident B cells. This repository data file contains sequences from two populations of ASCs (CD19+ and CD19-ASCs) isolated from 2 human nasal polyps. There were no replicates and no controls.

first: Alessia

last: Corrado

middle: P

first: F

last: Lee


title: Roles of TET and TDG in DNA demethylation in proliferating and non-proliferating immune cells [BS-PB]

city: Chiba

first: Atsushi

last: Onodera

address: Chuo-ku Inohana 1-8-1

department: Immunology

institute: Chiba University

summary: TET enzymes mediate DNA demethylation by oxidizing 5-methylcytosine (5mC) in DNA to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC). Because these oxidized methylcytosines (oxi-mC) are not recognized by the maintenance methyltransferase DNMT1, DNA demethylation can occur through “passive”, replication-dependent dilution as cells divide. A distinct, replication-independent (“active”) mechanism of DNA demethylation involves excision of 5fC and 5caC by the DNA repair enzyme thymine DNA glycosylase (TDG), followed by base excision repair. Here we used inducible gene-disrupted mice to show that TET enzymes influence both replication-dependent primary T cell differentiation and replication-independent macrophage differentiation, whereas TDG has no effect. Mice with long-term (1 year) deletion of Tdg are healthy and show normal survival and hematopoiesis. In summary, TET enzymes regulate differentiation and DNA demethylation primarily through passive dilution of oxidized methylcytosines in replicating T cells, and active, replication-independent DNA demethylation mediated by TDG does not appear to be essential for immune cell activation or differentiation.

overall_design: The proportions of 5mC/5hmC or 5fC/5caC for each cytosine were analyzed by bisulfite or pyridine borane sequencing, respectively.

first: Edahí

last: Gonzàlez Avalos

first: Chan-Wang

last: Lio

first: Romain

last: Georges

first: Alfonso

last: Bellacosa

first: Toshinori

last: Nakayama

first: Anjana

last: Rao


title: Roles of TET and TDG in DNA demethylation in proliferating and non-proliferating immune cells [RNA-seq]

overall_design: RNA-seq in BMDMs and Th2 cells. Effects of TDG or TET1-3 deletion were examined in Tdg-iKO or TET-iTKO, respectively, as compared to WT control.


title: Roles of TET and TDG in DNA demethylation in proliferating and non-proliferating immune cells


title: Roles of TET and TDG in DNA demethylation in proliferating and non-proliferating immune cells [CMS-Seq]

overall_design: Genome-wide distributions of 5hmC were analyzed by CMS-IP in BMDMs, naïve CD4 T-cells, iTregs, and Th2 cells.


title: Effect of platelets on 3D models of ovarian cancer

city: London

first: Eleni

last: Maniati

address: John Vane Science Centre, Charterhouse Square

department: Barts Cancer Institute

institute: Queen Mary University of London

summary: A 3D multicellular model (MCM) of high grade serous ovarian cancer (HGSOC) consisting of primary human adipocytes, fibroblasts, and mesothelial cells co-cultured with HGSOC cell lines was used to examine the effect of platelets on the ovarian metastasis microenvironment.

overall_design: RNA sequencing was performed on multicellular models with (n = 3) or without addition of platelets (n = 3) as well as co-cultures of mesothelial cells and fibroblasts (n = 3) and adipocyte monocultures (n = 1).

first: Beatrice

last: Malacrida

first: Sam

last: Nichols

first: Oliver

last: Pearce


title: Arginine methylation of METTL14 promotes RNA N6-methyladenosine modification

status: Public on May 14 2021

city: Guangzhou

first: Shan

last: Xiao

address: Southern Shatai Road

department: Developmental Biology

institute: Southern Medical University

summary: Both mRNA and proteins can be modified through addition of methyl groups. For example, addition of N6-methyladenosine (m6A) to mRNAs is critical for human development and health. Post-translational methylation of proteins can impact the dynamic regulation of enzymatic activity. Here we sought to explore the nexus of transcriptional and post-translational methylation by studying the role of methylation of the core methyltransferase METTL3/METTL14 in m6A regulation. We found by mass spectrometry that METTL14 arginine 255 (R255) is methylated (R255me). Global mRNA m6A levels were greatly decreased in METTL14 R255K mutant mouse embryonic stem cells (mESCs). We further found that R255me greatly enhances the interaction of METTL3/METTL14 with WTAP and promotes the binding of the complex to substrate RNA.

overall_design: MeRIP-seq and RNA lifetime sequencing of WT and METTL14 R255K mESCs

first: Xiaona

last: Liu

first: Jinru

last: Cui

first: Linjian

last: Xia


title: Effect of platelets on ovarian cancer cell spheroids

summary: The effect of platelets on malignant cell growth was assessed on high-grade serous ovarian cancer cells grown in spheroids.

overall_design: RNA sequencing on the G164 HGSOC malignant cell line grown in spheroids (n =3) and spheroids with addition of platelets (n = 3).

first: Roanne

last: Jones


title: ADP-ribosylation of STAT1α by PARP-1  is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages [iBMDM_RNA-Seq_PJ34]

first: W. Lee

last: Kraus

address: 5323 Harry Hines Blvd.

summary: A key determinant of the pro-inflammatory responses in macrophages is the Signal Transducers and Activators of Transcription (STAT) family member, STAT1α.  STAT1α activation by interferon-gamma (IFNγ) leads to induction of a transcriptional program that is coordinated in a large part by post-translational modifications such as phosphorylation.  Poly(ADP-ribose) Polymerases (PARPs), which include PARP-1, catalyze the addition of ADP-ribose moieties (ADP-ribosylation) to target proteins and modulate their function.  We found that PARP-1 mediates IFNγ-stimulated transcription by regulating genome-wide binding of STAT1α and its IFNγ-dependent phosphorylation.  We identified STAT1α as a target of PARP-1 and found sites of ADP-ribosylation on its DNA-binding (DBD) and Transcription Activation (TA) domains.  Surprisingly, ADP-ribosylation on the DBD and TA domains had distinct functional consequences on STAT1α transcriptional activity.  Moreover, loss of ADP-ribosylation on either site led to diminished pro-inflammatory responses.  These results suggest that PARP-1-driven ADP-ribosylation of STAT1α is a critical mediator of inflammation in macrophages.

overall_design:
RNA-Seq was perfomed in immortalized bone marrow derived macrophages (iiBMDM) having non-specfic (control) knockdown or Parp1 knockdown and treated with Interferon gamma (IFNγ).
Please note that each processed data file was generated from both replicates and is linked to the corresponding rep1 sample records.

first: Gupte

last: Rebecca

first: Nandu

last: Tulip

first: Kraus

last: W Lee


title: ADP-ribosylation of STAT1α by PARP-1  is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages [BMDM_RNA-Seq_Parp1Ko]

overall_design:
RNA-Seq was perfomed in bone marrow derived macrophages (BMDM) from Wildtype or Parp1 knockout mice and treated with Interferon gamma (IFNγ).
Please note that each processed data file was generated from both replicates and is linked to the corresponding rep1 sample records.


title: ADP-ribosylation of STAT1α by PARP-1  is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages [iBMDM_RNA-Seq_Parp1Ko]

overall_design:
RNA-Seq was perfomed in immortalized bone marrow derived macrophages (iBMDM) having non-specfic (control) knocsh.Parp1own or Parp1 knocsh.Parp1own and treated with Interferon gamma (IFNγ).
Please note that each processed data file was generated from both replicates and is linked to the corresponding rep1 sample records.


title: ADP-ribosylation of STAT1α by PARP-1 is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages.


title: ADP-ribosylation of H3K27acα by PARP-1  is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages [H3K27ac_ChIP-Seq_mutants_iBMDM]

summary: A key determinant of the pro-inflammatory responses in macrophages is the Signal Transducers and Activators of Transcription (STAT) family member, H3K27acα.  H3K27acα activation by interferon-gamma (IFNγ) leads to induction of a transcriptional program that is coordinated in a large part by post-translational modifications such as phosphorylation.  Poly(ADP-ribose) Polymerases (PARPs), which include PARP-1, catalyze the addition of ADP-ribose moieties (ADP-ribosylation) to target proteins and modulate their function.  We found that PARP-1 mediates IFNγ-stimulated transcription by regulating genome-wide binding of H3K27acα and its IFNγ-dependent phosphorylation.  We identified H3K27acα as a target of PARP-1 and found sites of ADP-ribosylation on its DNA-binding (DBD) and Transcription Activation (TA) domains.  Surprisingly, ADP-ribosylation on the DBD and TA domains had distinct functional consequences on H3K27acα transcriptional activity.  Moreover, loss of ADP-ribosylation on either site led to diminished pro-inflammatory responses.  These results suggest that PARP-1-driven ADP-ribosylation of H3K27acα is a critical mediator of inflammation in macrophages.

overall_design:
ChIp-seq was perfomed in immortalized bone marrow derived macrophages (iBMDM) expressing wildtype or ADP-ribosylation deficient mutants.
Please note that each processed data file was generated from both replicates and is linked to the corresponding rep1 sample records.


title: ADP-ribosylation of STAT1α by PARP-1  is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages [BMDM_RNA-Seq_PJ34]

overall_design:
RNA-Seq was perfomed in bone marrow derived macrophages (BMDM) treated with Interferon gamma (IFNγ) with or without PARP inhibitor treatment.
Please note that each processed data file was generated from both replicates and is linked to the corresponding rep1 sample records.


title: ADP-ribosylation of STAT1α by PARP-1  is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages [H3K27ac_ChIP-Seq_BMDM]

overall_design:
ChIP-seq was perfomed in bone marrow derived macrophages (BMDM) treated with Interferon gamma (IFNγ) with or without PARP inhibitor treatment.
Please note that each processed data file was generated from both replicates and is linked to the corresponding rep1 sample records.


title: ADP-ribosylation of STAT1α by PARP-1  is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages [STAT1_ChIP-Seq_Mutants_iBMDM]


title: Multiplexed functional genomic analysis of somatic 5' untranslated region mutations across the spectrum of human prostate cancer (PLUMAGE)

first: Sonali

last: Arora

overall_design: 18 samples

first: Yiting

last: Lim

first: Andrew

last: Hsieh


title: The effect of STAG2 loss in Ewing sarcoma [HiChIP]

city: Cambridge

first: Gabriela

last: Alexe

state: MA

address: 415 Main St.

department: Computational Biology and Bioinformatics

institute: Broad Institute

summary: Ewing sarcoma is an aggressive malignancy characterized by oncogenic rearrangements of the EWS gene with an ETS-family transcription factor, most commonly FLI. Recent comprehensive next-generation sequencing efforts have revealed few other highly recurrent mutations in this disease apart from loss-of-function mutations in STAG2 which occur in 15-20% of tumors. STAG2 is a member of the cohesin complex, which regulates sister chromatid alignment during mitosis and epigenetic regulation of gene expression. While some studies suggest that loss of STAG2 is associated with the development of aneuploidy, this is not the case in Ewing sarcoma. To investigate whether STAG2 loss affects epigenetic regulation of gene expression in Ewing sarcoma, we developed isogenic Ewing sarcoma cell lines with STAG2 knockout. We found that Ewing sarcoma cells engineered for loss of STAG2 maintain an intact cohesion complex that alternately incorporates STAG1.

overall_design: HiChIP SMC1A was performed for the Ewing sarcoma cell line A673 under two conditions: (1) A673 cells treated with non-targeting CRISPR Cas9 guides (STAG2 WT) and  (2) A673 cells treated with STAG2-targeting CRISPR Cas9 guides (STAG2 KO). Cells treated for gene editing were clonally selected and confirmed to either express STAG2 (control condition) or have a loss of STAG2 expression (STAG2 loss condition). For the control condition, we used the cell clones A673.sgNT-1c4 and the STAG2 knockout clone A673.sgSTAG2-1c6. HiChIP was performed for each clone in duplicate. Data analysis was designed as a 2x2 comparison of differential looping for SMC1A HiChIP in A673 STAG2 KO cells vs.  A673 STAG2 WT cells.

first: Kimberly

last: Stegmaier


title: ADP-ribosylation of STAT1α by PARP-1  is Required for IFNγ-stimulated Transcriptional Responses in  Macrophages [STAT1_ChIP-Seq_BMDM]


title: Higher self renewal and oncogenic potential of keratinocytes from Black versus White populations linked to the hydroxysteroid dehydrogenase HSD17B7 gene.

status: Public on May 12 2021

city: Biella

first: Paola

last: Ostano

address: via Malta 3

country: Italy

institute: Fondazione Edo ed Elvo Tempia

summary: Genotyping of 62 foreskin-derived human primary fibroblasts from individuals of Black African versus Caucasian ancestries.

overall_design: All genomic DNAs were assayed on Illumina HumanOmni2.5-8M bead chip arrays to feature 2372784 single nucleotide polymorphism (SNP) markers for each DNA sample.  Samples with low genotyping rate (<95%) were removed. SNPs that have a call rate lower than 0.95 were removed.  PLINK 1.075 and GCTA6 analysis tools were used for data QC and processing. Genetic relationship matrix (GRM) between pairs of individuals were estimated in our samples. “snpStats” (Bioconductor) was used as statistical tool for SNP association studies. “snpgdsPCA”7 (R function) was used to calculate the eigenvectors and eigenvalues for principal component analysis (PCA) based on filtered SNPs. First two principal components were used to create plots.

first: Xiaoying

last: Xu

first: Beatrice

last: Tassone

first: Atul

last: Katarkar

first: Tatiana

last: Proust

first: Berna

last: Ozdemir

first: Jean-Marc

last: Joseph

first: Chiara

last: Riganti

first: Giovanna

last: Chiorino

first: Zoltan

last: Kutalik

first: Karine

last: Lefort

first: Gian

last: Dotto


title: Xenografted breast epithelia reveal differences in growth stimulation among contraceptive progestins related to androgenic properties [human]

city: Lausanne

first: Fabio

last: De Martino

address: EPFL SV ISREC UPBRI Station 19

institute: EPFL

summary: Hormonal contraception exposes women to different synthetic progesterone receptor (PR) agonists, progestins, and transiently increases breast cancer risk. How progestins affect the breast epithelium is poorly understood because we lack adequate models to study this. We hypothesized that individual progestins differentially affect cell proliferation in the normal breast epithelium and hence breast cancer risk. Using mouse mammary tissue ex vivo, we show that testosterone-related progestins strongly induce the PR target and mediator of PR signaling induced cell proliferation Receptor Activator of NF-κB Ligand (Rankl), previously implicated in breast carcinogenesis, whereas other progestins fail to do so. We developed xenografts of normal human breast epithelial cells (HBECSs) to the milk ducts of immunocompromised female mice and show that they remain hormone-responsive. Using HBECSs from 36 women, we show that testosterone-related progestins, desogestrel, gestodene, and levonorgestrel, induce PSA (KLK3) and promote their proliferation, whereas the anti-androgenic progestins, which have shown anti-androgenic properties in reporter assays, chlormadinone acetate and cyproterone acetate, do not. Pharmacological inhibition of the androgen receptor (AR) inhibits PR agonist and levonorgestrel-induced RANKL expression and both pharmacological and genetic AR inhibition reduce levonorgestrel-driven breast epithelial cell proliferation in vivo. Prolonged exposure to androgenic progestins elicits hyperproliferation with cytologic changes.  Thus, different progestins have distinct biological activities in the breast epithelium that should be taken into account for more informed choices in hormonal contraception.

overall_design: 3 samples of human microstructures derived from 3 different patients stimulated with either ETOH, natural progesterone (P4) or promegestone (R5020).


title: Xenografted breast epithelia reveal differences in growth stimulation among contraceptive progestins related to androgenic properties.

overall_design: Refer to individual Series


title: Xenografted breast epithelia reveal differences in growth stimulation among contraceptive progestins related to androgenic properties [mouse]

overall_design: 3 paired biological replicates of mouse organoids stimulated with either ETOH or promegestone (R5020).


title: Higher self renewal and oncogenic potential of keratinocytes from Black versus White populations linked to the hydroxysteroid dehydrogenase HSD17B7 gene [HSD17B7 overexpression]

summary: Transcriptomic profiling of 3 human keratinocytes strains with HSD17B7 overexpression.

overall_design: Transcriptomic profiling of 3 human keratinocytes strains with HSD17B7 overexpression compared to correspondent control samples.

first: Jean


title: PA3027, a novel AraC-type transcriptional regulator activating genes of glycerolipid metabolism in Pseudomonas aeruginosa (PA3027 overexpression RNA-seq)

status: Public on May 11 2021

city: Warszawa

first: Adam

last: Kawałek

address: Pawinskiego 5A

department: Department of Microbial Biochemistry

country: Poland

institute: Institute of Biochemistry and Biophysics, PAS

summary: Pseudomonas aeruginosa encodes a large set of transcriptional regulators which allow to modulate and manage cellular metabolism to survive in variable environmental conditions, including the human body. The AraC family regulators are an abundant group of transcriptional regulators in bacteria, mostly acting as gene expression activators controlling diverse cellular functions e.g. carbon metabolism, stress response and virulence. PA3027 protein from P. aeruginosa has been classified in silico as a putative AraC-type transcriptional regulator. Comparative transcriptome analysis of P. aeruginosa PAO1161 overexpressing PA3027 versus empty vector control revealed a spectacular over 15-fold up-regulation of expression of pa3026-pa3024, PA3464 and PA3342 genes potentially involved in glycerolipid metabolism. Concomitantly, the ChIP-seq analysis using FLAG-PA3027 under mild overproduction conditions revealed at least 22 PA3027 binding sites in the PAO1161 genome, including promoter regions of genes showing a major increase in expression in response to PA3027 excess. The presented data showed that PA3027 acts as the transcriptional regulator in P. aeruginosa activating genes engaged in glycerolipid metabolism.

overall_design: Pseudomonas aeruginosa PAO1161 (leu-, r-, RifR) derivative of PAO1 was used in the experiment (Kawalek et al., 2020; BMC Genomics, 21:14). To determine the impact of an increased PA3027 level on the transcriptome the RNA-seq analysis was performed on RNA isolated from Pseudomonas aeruginosa PAO1161/ pKKB1.11 (lacIQ-tacp-PA3027) (hereafter referred to as PA3027) as well as from P. aeruginosa PAO1161 carrying pAMB9.37 (lacIQ-tacp) cells (empty vector control, hereafter called EV). Cells were grown under selection in L broth with 0.05 mM IPTG till reached the exponential phase of growth (OD600 0.5).The RNA was isolated from exponentially growing cultures (OD600 0.5). Three independent biological replicates of total RNA were isolated for each strain.

first: Karolina

last: Kotecka

last: Kawalek

first: Aneta

last: Bartosik


title: The effect of STAG2 loss in Ewing sarcoma

first: Gabriela

last: Alexe


title: PA3027, a novel AraC-type transcriptional regulator activating genes of glycerolipid metabolism in Pseudomonas aeruginosa  (F-PA3027 ChIP-seq)

overall_design: Pseudomonas aeruginosa PAO1161 (leu-, r-, RifR) derivative of PAO1, was used in the experiment (Kawalek et al., 2020; BMC Genomics, 21:14). Chromatin immunoprecipitation and sequencing (ChIP-seq) analysis was performed to identify PA3027 binding sites in P. aeruginosa. The PAO1161 ΔPA3027 / pKKB1.12 (lacIQ-tacp-flag-PA3027) [hereafter referred to as F-PA3027] and PAO1161 /pABB28.1 (lacIQ-tacp-flag) cells (empty vector control, hereafter called EV-F). Cells were grown under selection in L broth with 0.05 mM IPTG till reached the exponential phase of growth (OD600 0.5). For each biological replicate, two independent cultures of each strain were pooled together. The immunoprecipitation was performed with commercially available polyclonal anti-FLAG antibodies [DYKDDDDK Tag polyclonal  antibodies; PA1-985B; Invitrogen].


title: Gene expression changes upon siCTNNB1 and compound treatment

city: Singapore

first: Gokce

last: OGUZ

address: 60 Biopolis St, GENOME, #02-01

department: Bioinformatics Consulting and Training Platform

country: Singapore

institute: Genome Institute of Singapore

summary: To investigating gene expression changes in HCT116 cells upon siCTNNB1 for 48 hr, or upon treatment with iCRT3, GB1874, GB6853, and GB8679 for 24 hr

overall_design: HCT116 cells are transfected with siNeg or siCTNNB1 (20 nM each) in triplicate for 48 hr. HCT116 cells are also treated with DMSO (1%) or compounds iCRT3, GB1874, GB6853, GB8679 at 50 uM, in triplicate for 24 hr

first: Joo-Leng

last: Low

last: Oguz

first: Adaikalavan

last: Ramasamy

first: Ramanuj

last: DasGupta


title: A Surgical Method for Continuous Intra-Portal Infusion of Gut Microbial Metabolites in Mice

status: Public on May 10 2021

city: Cleveland

first: Lucas

last: Osborn

state: Ohio

address: 9500 Euclid Ave

institute: Cleveland Clinic Foundation

summary: Gut-derived microbial metabolites have been shown to play key roles in human physiology and disease. However, establishing mechanistic links between gut microbial metabolites and disease pathogenesis in animal models presents many challenges. The major route of absorption for microbe-derived small molecules is venous drainage via the portal vein to the liver. In the event of extensive liver first pass- or presystemic hepatic metabolism, the route of administration of these metabolites becomes critical. Here we describe a novel portal vein cannulation technique using a subcutaneously implanted osmotic pump to achieve continuous portal vein infusion in mice. First, the microbial metabolite trimethylamine (TMA) was administered over 4 weeks and compared to a vehicle control. Using a liquid chromatography-tandem mass spectrometry (LC-MS/MS), an increase in peripheral plasma levels of TMA and its host liver-derived co-metabolite trimethylamine-N-oxide (TMAO) were observed in a sexually-dimorphic manner. Next, 4-hydroxyphenylacetic acid (4-HPAA), a structurally-distinct microbial metabolite that undergoes extensive hepatic first pass metabolism, was administered portally to examine  its effects on hepatic gene expression. Peripheral plasma levels showed no difference in free or total 4-HPAA. However, while liver tissue demonstrated no difference in free 4-HPAA, total levels were increased when compared to the control group. More importantly, significant changes were observed in hepatic gene expression using an unbiased RNA sequencing approach. Collectively, this work describes a novel method for administering gut microbe-derived metabolites via the portal vein, mimicking their physiologic delivery in vivo.

overall_design: Comparison of liver mRNA transcription profiles after intraportal administration of either normal saline or 4-hydroxyphenylacetic acid (4-HPAA). N=4 per group.

first: Danny

last: Orabi


title: Generation of kidney ureteric bud and collecting duct organoid

city: Los Angeles

first: Brendan

last: Grubbs

address: 4661 W. Sunset Blvd.

department: Obstetrics and Gynecology

institute: University of Southern California

summary: A platform for generating expandable, branching and gene-editable ureteric bud organoid from primary mouse and human ureteric bud progenitor cells and human pluripotent stem cells, and its maturation into collecting duct organoid.

overall_design: mRNA profiles of mouse ureteric bud (UB) organoids cultured in mUBCM for 5 days, 10 days and 20 days, as well as mouse collecting duct (CD) organoids and adult mouse CD cells purified by FACS from kidneys of 2-month-old Hoxb7-Venus mice.

first: Zhongwei

last: Li

first: Zipeng

last: Zeng

first: Matthew

last: Thornton


title: Next-generation sequencing facilitates quantitative analysis of Ym1+/- monocytes from lung transcriptomes

city: Tokyo

first: Yotsumoto

last: Satoshi

address: 1432-1, Hachioji, Hachioji

institute: Tokyo University of Pharmacy and Life Sciences

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare Lung Ym1+/- monocytes transcriptome profiling (RNA-seq) to microarray.
Methods: Ym1+/- monocytes mRNA profiles of Ym1-Venus mice were generated by deep sequencing, in triplicate, using Illumina NextSeq 500. The sequence reads that passed quality filters were analyzed at the transcript isoform level with methods: TopHat followed by Cufflinks.

overall_design: Reparative monocyte subset mRNA profiles of Ym1-Venus mice.

first: Takumi

last: Shibuya


title: Integrative Pharmacology Strategy Powers the Delineation of the Main Active Components and Exploration the Underlying Mechanism of WBT Acting on Rheumatoid Arthritis

city: Beijing

first: Dan

last: Wu

address: No. 16, Nei Nanxiao Street, Dongzhimen, Dongcheng District

institute: Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences

summary: Wang Bi Tablet (WBT) exhibits effective power in clinic during the treatment of rheumatoid arthritis (RA) in China. We aimed to employ the “integrative pharmacology strategy”, including rapid analysis of chemical composition, pharmacological experiment, and network pharmacology analysis, to delineate the main active components and reveal the underlying mechanism of WBT acting on RA. UPLC-QTOF-MS/MS provided the chemical fingerprint of WBT and UNIFI 1.8 software assisted in chemical composition identification by matching the MSE raw data to the in-house library. The anti-inflammatory effect of WBT was evaluated in TNF-α- stimulated RAW264.7 cells by ELISA and transcriptome sequencing. Network pharmacology analysis was carried out based on Integrative Pharmacology-based Network Computational Research Platform of Traditional Chinese Medicine (TCMIP V2.0, http://www.tcmip.cn/). Functional enrichment and network visualization was performed by DAVID v6.8 and NaviGator v3.0 respectively. Altogether, 293 chemical constituents were preliminarily identified or tentatively characterized in the extract of WBT, and it could effectively against inflammatory response in TNF-α-stimulated RAW264.7 cells. One hundred and thirty-five hub genes were delineated based on the network construction of “Anti-inflammatory Core Genes-Putative Targets-RA genes”, which were mainly involved in inflammation-immune regulation, closely relating to the occurrence and development of RA, such as Cytokine-cytokine receptor interaction, Chemokine signaling pathway, etc., Forty-eight key active constituents were selected according to the frequency binding to hub targets and contents in WBT, corresponding to 13 herbal medicines. In summary, it is the first time to explore the active constituents and underlying mechanism of WBT acting on RA using integrative pharmacology strategy, which may provide an efficient way for the analysis of chemical constituents and exploration of pharmacological mechanism of TCM.

overall_design: Based on RAW264.7 cells (mouse origin), blank group, induction group (inflammatory factors), WBT group.  We want to obtain differentially expressed genes among each group, and further analyze the mechanism of drug efficacy.

first: Yuanyuan

last: JIAO

first: Jia

last: XU

first: Hong

last: CHEN

first: Ping

last: WANG


title: Reactivation of Zeta-Globin by deletion of BCL11A and LRF in murine cells

status: Public on May 09 2021

city: Headington

first: Damien

last: Downes

address: John Radcliffe Hospital

department: MRC Weatherall Institute of Medicine

institute: The University of Oxford

summary: The alpha and beta-globin loci harbour genes that are expressed during development and silenced throughout post-natal life. Learning to reactivate these genes may offer new therapeutic approaches for the hemoglobinopathies, the most common single gene disorders. Here, we address the mechanisms underlying the regulation of the gene encoding zeta-globin, the embryonically expressed alpha-like globin.

overall_design: To determine the role of LRF and Bcl11a in regulating zeta globin in definitive mouse erythroblasts, mouse lines lacking LRF or the BCL11A erythroid enhancer were generated. Lrf-flox (Maeda 2007; PMID: 17495164) mice were crossed to mice expressing Cre under control of Gata1 regulatory elements (Mao 1999; PMID: 10220414), leading to germline excision of floxed regions. Heterozygous Lrf-Flox/- mice were interbred to make genetic combinations. Bcll11a-EnhKO lines lack a 10kb region which contains the mouse erythroid Bcl11a enhancer (Smith 2016; PMID: 27707736); genetic combinations were generated by crossing. Effects of edited variants were determined by ATAC-seq in technical replicates.

first: Andrew

last: King

first: Chris

last: Babbs


title: CD44 alternative splicing senses intragenic DNA methylation in tumors via direct and indirect mechanisms

city: Paris

first: Eric

last: Batsche

address: 7-9, Quai Saint Bernard

department: B2A Biological Adaptation and Ageing - UMR8256 CNRS

institute: IBPS - Institut Biologie Paris Seine

summary:
Exon Level Expression Profiling of HeLa cells depleted of DNMT1 by 2 independent siRNAs
In HeLa cells where depletion of DNMT1 was sufficient to cause reduced levels of DNA methylation inside the body of CD44 and reduced usage of CD44 variant exons, without affecting splicing factors regulating CD44.

overall_design: We analyzed DNMT1-depleted HeLa cells fro using the Affymetrix Human Exon 1.0 ST platform. Array data was processed by Genosplice. Three biological  replicates were performed.

last: Batsché

first: Oriane

last: Mauger

first: Christian

last: Muchardt


title: Single cell RNA-seq of Oncorhynchus mykiss B cells isolated from blood

city: Valdeolmos

first: Pedro

last: Perdiguero

state: Madrid

address: Ctra. de Algete a El Casar Km 8.1

department: Animal Health Research Centre

country: Spain

institute: CISA-INIA

summary: These files represent single cell RNA-Seq data generated on a 10x Chromium genomics platform from Oncorhynchus mykiss B cells isolated from blood.

overall_design: Three healthy rainbow trout were killed by benzocaine overdose. To isolate peripheral blood leukocytes (PBLs) blood was drawn with a heparinized needle from the caudal vein and diluted 10 times with of Leibovitz medium. The resulting cell suspension was placed onto 51% Percoll density cushions collecting the interface cells after centrifugation. Leukocytes were incubated with anti-trout MHC-II APC and YO-PRO in FACS staining buffer. MHC-II+ leukocytes from lymphoid gate were isolated by sorting. A total of 2,500 cells (700 cells/µL) from each fish were loaded to chips from the Chromium Single Cell 5′ Gel Beads Kit (10x Genomics, CA, USA) and subjected to the Chromium Controller Instrument (10x Genomics) to generate single-cell gel beads in emulsion following manufacturer's instructions. Libraries for sequencing were constructed using Chromium Single Cell 5’ Library Construction Kit following manufacturer's instructions. Libraries were sequenced using a NextSeq instrument (Illumina) with 150PE chemistry.

first: Carolina

last: Tafalla

first: Esther

last: Morel

first: Patricia

last: Díaz-Rosales


title: Transcriptional response to overexpressing Vibrio Seventh Pandemic -II island encoded genes

status: Public on May 08 2021

city: Ithaca

first: Tobias

last: Doerr

address: 525 N Campus Rd

department: Microbiology

institute: Cornell University

summary:
Purpose: to characterize the regulatory targets of an AraC-like transcriptional regulator (VC0513) encoded on the Vibrio Seventh Pandemic Island -II (VSP-II) in V. cholerae O1 El Tor N16961
Methods: RNA was isolated from a wild-type N16961 carrying an IPTG-inducible copy of vc0513, vc0515, or an empty vector control
Results: vc0513 induction significantly increased expression of other VSP-II encoded genes relative to the empty vector control
Conclusions: our study represents the first analysis of a transcriptional regulator encoded on the VSP-II island

sample_organism: Vibrio cholerae O1 biovar El Tor str. N16961

overall_design: Examination of transcript profiles of vc0513 or vc0515 expressing V. cholerae relative to an empty vector control

first: Shannon

middle: G

last: Murphy


title: mRNA expression profile in BMCMCs from C57BL/6J and CC027 mice stimulated with or without IgE-ligation.

city: Nishinomiya

first: Kazufumi

last: Matsushita

state: Hyogo pref.

address: 1-1 Mukogawacho

institute: Hyogo College of Medicine

summary: Collaborative Cross (CC) mice are a reproducible yet genetically diverse genetic reference panel derived from eight founder strains. Here, we compared the gene expression profiles of mast cells (MCs) from C57BL/6J (a traditional inbred strain) and CC027 (a CC strain) mice. We generated bone-marrow-derived cultured MCs (BMCMCs) from C57BL/6J and CC027 mice. The cells were sensitized with anti-DNP IgE overnight, then were stimulated with or without DNP-HSA, followed by total RNAs extraction. RNA-seq library construction and sequencing was done by Novogene (https://en.novogene.com/).

overall_design: BMCMCs from C57BL6J and CC027 mice were sensitized with anti-DNP-IgE and stimulated with or without DNP-HSA for 1 hour, followed by RNA extraction. Then, RNA-sequencing was performed.

first: Xin

last: Li

first: Yuki

last: Nakamura

first: Danyue

last: Dong

first: Kaori

last: Mukai

first: Mindy

last: Tsai

first: Stephen

last: Montgomery

last: Galli


title: Increased Systemic HSP70B Levels in Spinal Muscular Atrophy Infants

city: Boston

first: Kathryn

last: Swoboda

address: 185 Cambridge St.

department: Neurology

institute: Massachusetts General Hospital

summary: RNA sequencing in whole blood from spinal muscular atrophy patients and healthy control infants

overall_design: Here, we performed whole-blood RNA sequencing and differential expression analyses in SMA type 1 subjects under 1 year old and age-matched healthy control subjects. Our analysis identified for the first time the Heat Shock Protein Family A Member 7 (HSPA7) as a novel candidate biomarker to track SMA progression in the first year of life, with its circulating protein levels associated with circulating neurofilaments levels in SMA newborns and infants.

last: Eichelberger

first: Christiano

last: Alves

first: Ren

last: Zhang

first: Marco

last: Petrillo

first: Patrick

last: Cullen

first: Wildon

last: Farwell

first: Jessica

middle: A

last: Hurt

first: John

middle: F

last: Staropoli


title: Transcriptional reprogramming by oxidative stress occurs within a predefined chromatin accessibility landscape [ATAC-seq]

status: Public on May 07 2021

city: Duluth

first: Matthew

last: Slattery

state: MN

address: 1035 University Drive, SMed 255

department: Biomedical Sciences

institute: University of Minnesota Medical School

summary: Reactive oxygen species (ROS) are important signalling molecules in many physiological processes, yet excess ROS leads to cell damage and can lead to pathology. Accordingly, cells need to maintain tight regulation of ROS levels, and ROS-responsive transcriptional reprogramming is central to this process. Although it has long been recognized that oxidative stress leads to rapid, significant changes in gene expression, the impact of oxidative stress on the underlying chromatin accessibility landscape remained unclear. Here, we asked whether ROS-responsive transcriptional reprogramming is accompanied by reprogramming of the chromatin environment in MCF7 human breast cancer cells. Using a time-course exposure to multiple inducers of oxidative stress, we determined that the widespread ROS-responsive changes in gene expression induced by ROS occur with minimal changes to the chromatin environment. While we did observe changes in chromatin accessibility, these changes were: (1) far less numerous than gene expression changes after oxidative stress, and (2) occur within pre-existing regions of accessible chromatin. TF footprinting analysis identified 5 TFs or TF families with evidence for ROS-responsive changes in DNA binding: NRF2, AP-1, p53, NFY, and SP/KLF. Importantly, several of these (AP-1, NF- Y, and SP/KLF factors) have not been previously implicated as widespread regulators in the response to ROS. In summary, we have characterized genome-wide changes in gene expression and chromatin accessibility in response to ROS treatment of MCF7 cells, and we have found that regulation of the large-scale transcriptional response to excess ROS is primarily constrained by the cell’s pre-existing chromatin landscape.

overall_design: ATAC-seq of MCF7 cells at 0, 1, 8 and 24 hrs treatment with ethanol vehicle control (EtOH), menadione (MEN) or tert-butyl hydroperoxide (TBOOH), 2 replicates each.

first: Daniel

last: Levings

first: Sarah

last: Lacher


title: Unexpected contribution of fibroblasts to the muscle lineage during foetal myogenesis and role of BMP signalling in this process

status: Public on May 06 2021

first: Estelle

last: Hirsinger

address: 9 quai Saint Bernard

department: LBD

institute: IBPS

summary:
Purpose : Positional information driving limb muscle patterning is considered to be contained in lateral plate mesoderm-derived tissues, such as tendon or muscle connective tissue and not in myogenic cells themselves. The current consensus is that myogenic cells originate from somites, while connective tissue fibroblasts originate from the lateral plate mesoderm. We challenged this model by cell and genetic lineage tracing experiments and identified that a subpopulation of limb myogenic cells did not originate from somite or Pax3 lineage, but rather originated from the lateral plate mesoderm and were derived from Osr1 and Scx lineages.
Results: Analysis of single-cell RNA-sequencing data obtained from limb cells at successive developmental stages identified a subpopulation of cells displaying a dual muscle and connective tissue signature, in addition to independent muscle and connective tissue populations. Active BMP signalling was detected in this junctional cell sub-population and at the tendon/muscle interface in developing limbs. BMP gain- and loss-of-function experiments performed in vivo and in vitro showed that this signalling pathway regulated a fibroblast-to-myoblast conversion.
Conclusions: We propose that localised BMP signalling converts a subset of lateral plate mesoderm-derived fibroblasts to a myogenic fate and establishes a boundary of fibroblast-derived myonuclei at the tendon/muscle interface to control the muscle pattern during limb development and myotendinous formation.

overall_design: Single cell mRNA profiles of whole forelimbs in wild type chicken embryos at three developmental stages (E4, E6 and E10)

first: Joana

last: Esteves de Lima

first: Marie-Ange

last: Bonnin

first: Léa

last: Bellenger

first: Sébastien

last: Mella

first: Delphine

last: Duprez


title: Higher self renewal and oncogenic potential of keratinocytes from Black versus White populations linked to the hydroxysteroid dehydrogenase HSD17B7 gene

first: Paola

last: Ostano

summary: Transcriptomic profiling of 54 foreskin-derived human primary keratinocytes from individuals of Black African versus Caucasian ancestries.

overall_design: 28 African and 26 Caucasian foreskin-derived human primary keratinocytes were transcriptionally profiled using Clariom_D_Human Affymetrix platform.

first: Xiaoying

last: Xu

first: Beatrice

last: Tassone

first: Atul

last: Katarkar

first: Tatiana

last: Proust

first: Berna

last: Ozdemir

first: Jean

last: Joseph

first: Chiara

last: Riganti

first: Giovanna

last: Chiorino

first: Zoltan

last: Kutalik

first: Karine

last: Lefort

first: Gian

last: Dotto


title: CCM3 is a gatekeeper in focal adhesions regulating mechanotransduction and YAP/TAZ signalling

city: Lund

first: Chris

middle: Denis

last: Madsen

address: Scheelevägen 2

department: Laboratory Medicine

country: Sweden

institute: Lund University

summary: We report that loss of CCM3/PDCD10 in fibroblasts induces FAK/Src-paxillin signalling driving actomyosin-dependent mechanotransduction leading to YAP/TAZ signalling. In vivo, loss of CCM3 in fibroblasts drives excessive tissue remodelling leading to the dissemination of breast cancer cells to distant organs.

overall_design: Examination of the varying RNA levels in control human vulval cancer-associated fibroblasts versus CCM3-depleted cells

first: Shan

last: Wang


title: Transcriptional reprogramming by oxidative stress occurs within a predefined chromatin accessibility landscape [RNA-seq]

first: Matthew

last: Slattery

overall_design: RNA-seq of MCF7 cells at 0, 1, 8 and 24 hrs treatment with ethanol vehicle control (EtOH), menadione (MEN) or tert-butyl hydroperoxide (TBOOH), 2 replicates each.

first: Daniel

last: Levings

first: Sarah

middle: E

last: Lacher


title: Genome-wide essential gene identification for Shiga toxin-induced cell death using CRISPR/CAS9 screening system with Vero cells

city: Shinjuku-ku

first: Tsuyoshi

last: Sekizuka

state: Tokyo

address: 1-23-1 Toyama

department: Pathogen Genomics Center

institute: National Institute of Infectious Diseases

summary: Isolation of genes in Vero cell whose knockout confer resistance to Stx-induced cell death

overall_design: A genome-wide CRISPR/CAS9 knockout screen in Vero C1008 cells was performed to identify host factors involved in the regulation of glycan biosynthesis by exploiting Stx sensitivity.

first: Toshiyuki

last: Yamaji

first: Makoto

last: Kuroda


title: Molecular Logic of Cellular Diversification in the Mouse Cerebral Cortex

city: Cambridge

first: Paola

last: Arlotta

address: 7 Divinity Ave

department: Stem Cell and Regenerative Biology

institute: Harvard University

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

overall_design: Refer to individual Series


title: Histone arginine methyltransferase CARM1-mediated H3R26me2 is essential for morula-to-blastocyst transition in pigs

city: Hefei

first: Zubing

last: Cao

state: Anhui

address: 130 changjiang west road

department: Animal Science

institute: Anhui Agricultural University

summary: we show that CARM1 specifically catalyzes H3R26me2 to promote porcine blastocyst formation. The putative histone substrates of CARM1, including H3R2me2, H3R17me2, and H3R26me2, are present in pig early embryos

overall_design: Examination of 6 different samples about non-injected and CARM1-siRNA injected embryos

first: Xu

last: Tong

first: Huiqun

last: Yin

first: Naru

last: Zhou

first: Xiangdong

last: Zhang

first: Mengya

first: Xin

first: Qiuchen

last: Liu

first: Yelian

last: Yan

first: Yangyang

last: Ma

first: Tong

last: Yu

first: Yunsheng

last: Li

first: Yunhai