title: Heat Shock Factor 1 (HSF1) supports the ESR1 action in breast cancer (ChIP-seq)

city: Bergen

first: Tomasz

last: Stokowy

address: Nygardsgaten 5

department: IT Division

country: Norway

institute: University of Bergen

summary: Heat shock factor 1 (HSF1) is a key regulator of transcriptional responses to proteotoxic stress. It has been recently linked to signaling of estrogen via ESR1. To study the cooperation of HSF1 and ESR1 in the transcriptional response to estrogen, we established estrogen receptor (ER)-positive breast cancer cell lines with reduced HSF1 levels using specific shRNA or CRISPR/Cas9 approach. HSF1 deficiency led to the inhibition of the mitogenic effect of estrogen in MCF7 and T47D cells. RNA-seq analyses revealed that the stimulatory effect of E2 on the transcriptome was smaller in HSF1-deficient MCF7 cells. This could partially result from the higher basal expression of E2-dependent genes in these cells as a consequence of the enhanced binding of unliganded ESR1 to chromatin, which was revealed by ChIP-seq analyses. Thus, we postulate that some fraction of ESR1 could be released from the inhibitory complex with HSP90 and gain transcriptional competence without E2-stimulation.

overall_design: We sequenced DNA immunoprecipitated from MCF7 cells variants with the normal level of HSF (WT and MIX) and in HSF1-deficient cells (KO#2), using mouse monoclonal anti-ERα antibody. Cells were untreated (control) or stimulated using estrogen (17beta-estradiol) for indicated time. Four ChIP biological replicates were collected and pooled per each sequenced sample. Input samples were included as negative control (no DNA enrichment was observed in mock-IP samples).

first: Natalia

last: Vydra

first: Patryk

last: Janus

first: Wieslawa

last: Widlak


title: Single-cell multi-modal profiling of proteins and chromatin accessibility using PHAGE-ATAC [Intracellular_GFP]

status: Public on Oct 21 2021

city: New York

first: Caleb

last: Lareau

state: New York

address: 417 E 68th St, Zuckerman - ZRC 1132

country: USA

institute: Memorial Sloan Kettering

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Multi-modal measurements of single cell profiles are a powerful tool for characterizing cell states and regulatory mechanisms. While current methods allow profiling of RNA along with either readouts of chromatin or protein, connecting chromatin state to protein levels remains a barrier. Here, we developed PHAGE-ATAC, a method that uses engineered camelid single-domain antibody (‘nanobody’)-displaying phages for simultaneous single-cell measurement of surface proteins, chromatin accessibility profiles, and mtDNA-based clonal tracing through a single-cell and massively parallel droplet-based assay of transposase-accessible chromatin with sequencing (ATAC-seq). We demonstrate PHAGE-ATAC for multimodal analysis in primary human immune cells, for multiplexing, for intracellular protein analysis, and for the detection of SARS-CoV-2 spike protein. Finally, we construct a synthetic high-complexity phage library for selection of novel antigen-specific nanobodies that bind cells of particular molecular profiles, opening a new avenue for protein detection, cell characterization and screening with single-cell genomics.

overall_design: 293T cells expressing cytosolic EGFP were fixed, lysed and then incubated with an anti-EGFP phage nanobody and processed for PHAGE-ATAC

first: Evgenij

last: Fiškin


title: Single-cell multi-modal profiling of proteins and chromatin accessibility using PHAGE-ATAC [ASAP_Phage_comparison]

overall_design: PBMCs were incubated with a panel of phage nanobodies or commercial TotalSeq-A antibodies and processed for PHAGE-ATAC, PHAGE-ASAP or ASAP-seq


title: 1-deoxysphingolipids bind to COUP-TF to modulate lymphatic and cardiac cell development

city: New Haven

first: Fabio

middle: R.

last: Santori

state: CT

address: Yale University Immunobiology Department TAC BLDG RM S640, 300 Cedar Street

department: Immunobiology

institute: Yale University

type: Expression profiling by high throughput sequencing

summary: We identified 1-deoxysphingosine as a ligand for NR2F1 and NR2F2. To prove physiological relevance after in vitro binding assay we used different strategies to manipulate levels of 1-deoxysphingosines and canonical sphingosines in cardiomyocytes derived from human embrynonic stem cells. These manipulations included treating cells with sphingolipid biosynthesis inhibitors (myriocin), adding 1-deoxysphingosines to culture media or induced expression of the Sptlc1 HSAN mutant C133W to enhance 1-deoxysphingosine synthesis by cardiomyocytes.

overall_design: Examination of NR2F1 and NR2F2 targets after treatment with myriocin (sphingolipid biosynthetic inhibitor) or treatment with vehicle, canonical D-erythro-sphingosine or 1-deoxysphingosine. NR2F1 and NR2F2 targets were also analyzed after induction of 1-deoxysphingosine biosynthesis by tetracycline induced expression of SPTLC1 HSAN C133W mutant.

middle: R

middle: B

last: Ivanova


title: Single-cell multi-modal profiling of proteins and chromatin accessibility using PHAGE-ATAC [Spike_mix_pbmcs]

overall_design: A mix of PBMCs and 293T cells expressing either membrane-exposed EGFP-GPI or the SARS-CoV-2 spike protein were incubated with a panel of phage nanobodies and processed for PHAGE-ATAC


title: Analysis of the Chlamydomonas mitochondrial ribosome RNA content

city: Strasbourg

first: Thalia

last: Salinas-Giegé

address: 12 rue du général Zimmer

country: France

institute: CNRS

summary: The goal of this analysis was to identify the ribosomal RNA content of the mitochondrial risome of Chlamydomonas reinhardtii green alga

overall_design: Total RNA extracted from purified mitochondria compared to the RNA extracted from purified mitochondrial ribosome .

last: Salinas


title: Single cell analysis of protein epitopes and chromatin accessibility using PHAGE-ATAC [Species_Mix_GFP]

summary: Single cell multi-omic readouts of both the cellular transcriptome and proteome have significantly enhanced our ability to comprehensively characterize cellular states. Most approaches in this area rely on oligonucleotide barcode-conjugated antibodies that target cell surface epitopes of interest, enabling their concomitant detection with the transcriptome. However, a similar high-throughput measurement of other cellular modalities such as the epigenome in concert with protein levels have not been described. Moreover, detection of epitopes is limited to antigens for which a specific antibody is available. Here, we introduce PHAGE-ATAC, an approach that enables the scalable and simultaneous detection of protein levels and chromatin accessibility data in single cells using the assay of transposase-accessible chromatin with sequencing (ATAC-seq). Quantitative detection of proteins by PHAGE-ATAC is accomplished through the use of engineerable nanobody-displaying phages that are genetically barcoded within the nanobody-encoding phagemids. We demonstrate the utility of PHAGE-ATAC for multimodal single cell genomic analysis in both cell lines and primary human cells. Analogous to phage display approaches, we further establish a synthetic high-complexity library of nanobody-displaying phages and demonstrate its utility to select novel antigen-specific nanobodies for PHAGE-ATAC.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: A 2:1:1 ratio of NIH3T3, EGFP+ HEK293T, and EGFP- HEK29T3 were stained with phages against EGFP carrying a known anti-EGFP nanobody. Cell mixtures were then processed using a modified scATAC-seq protocol.


title: Single cell analysis of protein epitopes and chromatin accessibility using PHAGE-ATAC

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

overall_design: Refer to individual Series


title: Single cell analysis of protein epitopes and chromatin accessibility using PHAGE-ATAC [Human_PBMCs]

overall_design: Human PBMCs were stained with Phages displaying anti-CD4, CD8, or CD16 nanobodies to quantify protein abundances along side accessible chromatin using scATAC-seq


title: Comparing the transcriptome of early and late phase wound macrophages in skin

city: Bonn

first: Joachim

last: Schultze

state: NRW

address: Carl-Troll-Strasse 31

department: Genomics and Immunoregulation

country: Germany

institute: LIMES (Life and Medical Sciences Center Genomics and Immunoregulation)

summary: We inflicted full thickness excisional wounds in the back skin of C57BL/6 wild-type mice. Injury-associated macrophages were isolated from wound tissue during the early inflammatory phase (4 days post injury) and in the late, scar forming phase (14 days post injury), sorted by flow cytometry (CD45+CD11b+F4/80+ wound cells), and subjected to RNAsequencing.

overall_design:
Analysis of the differences in gene expression between early and late phase skin wound macrophages.
Raw data was unavailable at the time of submission because the raw read files had been deleted

middle: L

first: Thomas

last: Ulas

first: Sabine

middle: A

last: Eming

first: Sebastian

last: Willenborg


title: BCR-Seq of Germinal Center B cells from mice infected with LCMV

status: Public on Oct 20 2021

city: Melbourne

first: Andrea

last: Di Pietro

state: VIC

address: 305 Grattan Street

department: Cancer Immunology program

country: Australia

institute: Peter MacCallum Cancer Centre

summary: To study how acute or chronic viral infection (LCMV-WE or Docile) affect somatic hypermutation and/or affinity maturation of Germinal Center B cells, wild-type or Bmi1-deleted mice were infected with either the acute (WE) versus chronic (Docile) strains of LCMV, GC B cells (CD19+IgDloCD95+CD38loCD138-) were isolated 14 days post-infection and thei BCR was sequenced.

overall_design: 8 samples. Each group has at least 2 biological replicates.

first: Kim

last: Good-Jacobson


title: Lysyl hydroxylase 2 deficiency promotes filopodia formation and fibroblast migration

city: Chiba City

first: Ryunosuke

last: Nozaki

state: Chiba Prefecture

address: 1-8-1 Inohana, Chuo-ku, Chiba, Japan

department: Depertment of Oral science

country: Japan

institute: Graduate School of Medecine, Chiba university

summary: Lysyl hydroxylase 2 (LH2) regulates the intermolecular cross-linking of collagen molecules. Accumulation of LH2-modified collagen, which is highly stable and resistant to collagenase cleavage, is one of the causes of fibrosis. In this study, we established LH2-deficient fibroblasts and first analyzed morphology of LH2-deficient fibroblasts by microscopy. LH2-deficient fibroblasts showed a dramatic increase in filopodial dynamics and cell migration, which were supported by time-lapse video microscopy and Immunocytochemistry, and gene expression profiling data. Understanding the precise role of phenotype in LH2-deficient cells may be helpful to define the pathogenesis of fibrosis.

overall_design: Data shows the comparison of differentially expressed genes in LH2 cKO TTFs in culture medium with tamoxifen (LH2 cKO_no.1) vs. LH2 cKO TTFs in culture medium without tamoxifen (LH2 cKO_no.2) after 24h tamoxifen treatent.

first: Atsushi

last: Kasamatsu

first: Joel

last: Moss

first: Katsuhiro

last: Uzawa


title: mRNA expression profile of bovine monocyte derived macrophages infected with Mycobacterium avium subsp. Paratuberculosis (MAP)

city: Changchun

first: Yanhong

last: Bao

state: jilin

address: jilin changchun

country: China

institute: Jilin Agricultural University

type: Non-coding RNA profiling by high throughput sequencing

summary: In order to gain a better understanding of MAP infection in immature macrophages, high-throughput sequencing technology was used to perform an analysis of mRNAs profiles of bovine monocyte derived macrophages upon MAP infection. The total numbers of raw reads collected from the MAP-infected and control MDM cells were 93.6 and 102 million raw reads per library. After removing low-quality sequences, Adaptor sequences and sequences containing more than 5% N (N means that base information cannot be determined), 91.7 and 100 million clean reads were obtained respectively. The analyses of differential expression revealed 618 DE mRNAs in MDM challenged with MAP compared to the control MDM, Among the DE mRNAs, 322 were upregulated and 296 were downregulated.Furthermore, The GO and KEGG pathway enrichment of these mRNA targets were performed.

overall_design:
Infecting bovine peripheral blood mononuclear macrophages with Mycobacterium avium subsp. paratuberculosis
mRNA-sequencing analysis was performed on an Illumina Hiseq 4000  following the vendor's recommended protocol


title: Klf5 confers embryonic and extra-embryonic bi-potential cell fate in early blastomeres and embryonic stem cells

city: Berkeley

first: Martin

last: Kinisu

state: California

address: 300C Li Ka Shing Building

department: MCB

institute: University of California Berkely

summary: This study examines the transcriptomes of embryonic stem cells following overexpression of Klf5. Additionally, this study examines the transcripomes of single embryos either treated with scramble siRNA or siRNA targeting Klf5.

overall_design: Bulk RNA-seq was performed on mESCs overexpressing Klf5 or GFP as a control. Single embryo RNA-seq was performed on single embryos injected with either a scramble control siRNA, or an siRNA specific to Klf5

first: Claudia

last: Cattoglio

first: Lin

last: He


title: SETDB1-TRIM28 complex suppresses antitumor immunity

city: Philadelphia

first: Priyankara

middle: J

last: Wickramasinghe

state: PA

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute


title: SETDB1-TRIM28 complex suppresses antitumor immunity [CRISPR/Cas9 screen]

summary: Epigenetic mechanism contributes to immune landscapes in cancer.  Here we identify the SETDB1-TRIM28 complex as a critical suppressor of antitumor immunity.  An epigenetic CRISPR-Cas9 screen of 1,218 chromatin regulators identified TRIM28 as a novel suppressor of PD-L1 expression.  We revealed that expression of the SETDB1-TRIM28 complex negatively correlates with infiltration of effector CD8+ T cells.  Inhibition of SETDB1-TRIM28 simultaneously upregulates PD-L1 and activates the cGAS-STING innate immune response to increase infiltration of CD8+ T cells.  Mechanistically, SETDB1-TRIM28 inhibition leads to micronuclei formation in cytoplasm, a known activator of the cGAS-STING pathway.  Thus, SETDB1-TRIM28 inhibition bridges the innate and adaptive immunity.  Indeed, SETDB1 knockout enhances the antitumor effects of immune checkpoint blockade anti-PD-L1 in an ovarian cancer mouse model in a cGAS dependent manner. Our findings establish SETDB1-TRIM28 complex as a regulator of antitumor immunity and its loss activates cGAS-STING innate immunity to boost antitumor effects of immune checkpoint blockades.

overall_design: Epigenetic CRISPR/Cas9 screen

first: Jianhuang

last: Lin

first: Rugang

last: Zhang


title: Transcriptome profiling of HeLa TAF7 (WT) cytoplasmic extracts

status: Public on Oct 22 2021

city: Bethesda

first: Dinah

middle: S.

last: Singer

state: MD

address: Building 10, Room 4B-17

department: Division of Cancer Biology

institute: National Cancer Institute

summary: We perform RNA-Seq to analyze gene expression profiles in HeLa TAF7 (WT) cytoplasmic extracts, which are the inputs of RIP-Seq and PAR-CLIP.

overall_design: We isolated cytoplasmic RNA from HeLa TAF7 (WT) cells, followed by high-throughput RNA-Seq.

middle: S

first: Dan

last: Cheng


title: SETDB1-TRIM28 complex suppresses antitumor immunity [RNA-seq]

overall_design: RNA-seq experiment in SETDB1 WT and KO cells


title: Microarray of total lung cells from young and aged female mice

city: New Brunswick

first: Qi

last: Yang

state: NJ

address: 89 French St

institute: Rutgers RWJMS

summary: Very little is known about the effects of aging on tissues such as the lung, and the resulting effects of immune cells within the tissue. We performed a microarray on total cells from young and aged mice to determine signaling pathways that are altered with aging.

overall_design: Total cells from left lung lobe from 3 young (2-3 months old) and 3 aged (20-22 months old) female C57BL/6 mice were snap frozen, and RNA was extracted using Trizol and used for microarray analysis

first: Shanti

last: D'Souza


title: Heat Shock Factor 1 (HSF1) supports the ESR1 action in breast cancer

first: Tomasz

last: Stokowy


title: RNA-Seq during tumor cell plasticity and Platr18 overexpression/silencing

city: Mobile, AL

first: Simon

last: Grelet

state: Alabama

address: 1660 Spring Hill Ave - USA Drive North,

institute: University of South Alabama - Mitchell Cancer Institute

summary: The study aims: 1-to identify transcripts that are regulated during breast tumor cell plasticity induced by TGF-beta or by the silencing of PCBP1 expression and 2- to analyze the function of Platr18 lincRNA in breast cancer cells transcriptome reprogramming.

overall_design: NMuMG cells were treated with TGF-beta or with a PCBP1-shRNA to induce EMT. Cells over-expressing or silenced for Platr18 were also analyzed

first: Cécile

last: Fréreux

first: Clémence

last: Obellianne

first: Ken

last: Noguchi

first: Breege

last: Howley

first: Annamarie

last: Dalton

first: Philip

last: Howe


title: Optineurin depletion in dendritic cells

city: Hangzhou

first: Lulu

address: Yuhangtang Road 866

institute: Zhejiang Province Key Laboratory of Anti-Cancer Drug Research

summary: Dendritic cells (DCs) are specialized antigen-presenting cells (APCs) linking innate and adaptive immune response. DCs are crucial for initiating adaptive immune responses for elimination of invading pathogens and also in inducing immune tolerance toward harmless components to maintain immune homeostasis. Results provide insight into the function of optineurin and its role in the maturation and function of DCs.

overall_design: Bone marrow-derived dendritic cells (BMDCs) were cultured from Ctrl and DC-conditional Optn KO mice. Total RNA were obtained after LPS treatment for 8h.

first: Jiaying

last: Wang

first: Jincheng

first: Qinjie

last: Weng


title: SUMOylation of Linker Histone H1 Drives Chromatin Condensation and Restriction of Embryonic Cell Fate Identity [ChIP-seq]

city: Rehovot

first: Noa

last: Novershtern

address: Weizmann Institute

department: Molecular Genetics

country: Israel

institute: Weizmann Institute of Science

summary: The fidelity of the early embryonic program is underlined by tight regulation of the chromatin. Yet, our understanding of how the chromatin is organized to prohibit the reversal of the developmental program remains limited. Specifically, the totipotency to pluripotency transition marks one of the most dramatic events to the chromatin, yet the nature of histone changes and regulation underlying this process are only partly characterized. Here we show that linker histone H1 is post-translationally modulated by SUMO2/3, which facilitates its fixation onto ultra-condensed heterochromatin in embryonic stem cells (ESCs). Upon SUMOylation depletion, the chromatin becomes de-compacted and H1 is evicted, leading to totipotency reactivation. Further, we show that H1 and SUMO2/3 jointly mediate the repression of totipotent elements. Lastly, we demonstrate that preventing SUMOylation on H1 abrogates its ability to repress totipotency in ESCs. Collectively, our findings unravel a critical role for SUMOylation of H1 in mediating chromatin repression and desolation of the totipotent identity.

overall_design: ChIP-seq for H3K27ac was measured in Ubc9-depleted mESCs and their isogenic control. In addition, Sumo1 ChIP-seq was measured in v6.5 mouse ESCs.

first: Daoud

last: Sheban

first: Tom

last: Shani

first: Yifat

last: Merbl

first: Jacob

middle: H

last: Hanna


title: Heat Shock Factor 1 (HSF1) supports the ESR1 action in breast cancer (RNA-seq)

status: Public on Oct 19 2021

first: Tomasz

summary: Heat shock factor 1 (HSF1) is a key regulator of transcriptional responses to proteotoxic stress. It has been recently linked to signaling of estrogen via ESR1. To study the cooperation of HSF1 and ESR1 in the transcriptional response to estrogen, we established estrogen receptor (ER)-positive breast cancer cell lines with reduced HSF1 levels using specific shRNA or CRISPR/Cas9 approach. HSF1 deficiency led to the inhibition of the mitogenic effect of estrogen in MCF7 and T47D cells. RNA-seq analyses revealed that the stimulatory effect of E2 on the transcriptome was smaller in HSF1-deficient MCF7 cells. This could partially result from the higher basal expression of E2-dependent genes in these cells as a consequence of the enhanced binding of unliganded ESR1 to chromatin, which was revealed by ChIP-seq analyses. Thus, we postulate that some fraction of ESR1 could be released from the inhibitory complex with HSP90 and gain transcriptional competence without E2-stimulation.

overall_design: Total RNA was isolated from all MCF7 cell variants (untreated and treated with 10 nM E2 for 4 h or heat shocked for 1h at 43°C and recovered for 2 h) using the Direct-ZolTM RNA MiniPrep Kit (Zymo Research) and digested with DNase I (Worthington Biochemical Corporation). For each experimental point, RNAs from three biological replicates were first tested by RT-qPCR for the efficiency of treatments, then pooled and sequenced.

first: Natalia

last: Vydra

first: Patryk

last: Janus

first: Wieslawa

last: Widlak


title: CRISPR recording in zebrafish embryos with orthogonal CRISPR systems

status: Public on Oct 25 2021

city: Salt Lake City

first: James

last: Gagnon

state: UT

address: 1390 Presidents Circle, Crocker Science Center Receiving

institute: University of Utah

summary: The CRISPR-Cas universe continues to expand. The type II CRISPR-Cas system from Streptococcus pyogenes (SpyCas9) is most widely used for genome editing due to its high efficiency in cells and organisms. However, concentrating on a single CRISPR-Cas system imposes limits on target selection and multiplexed genome engineering. We hypothesized that CRISPR-Cas systems originating from different bacterial species could operate simultaneously and independently due to their distinct single-guide RNAs (sgRNAs) or CRISPR-RNAs (crRNAs), and protospacer adjacent motifs (PAMs). Additionally, we hypothesized that CRISPR-Cas activity in zebrafish could be regulated through the expression of inhibitory anti-CRISPR (Acr) proteins. Here, we use a simple mutagenesis approach to demonstrate that CRISPR-Cas systems from Streptococcus pyogenes (SpyCas9), Streptococcus aureus (SauCas9), Lachnospiraceae bacterium (LbaCas12a, previously known as LbCpf1), Acidaminococcus sp. (AspCas12a, previously known as AsCpf1) and Neisseria meningitidis (Nme2Cas9) are orthogonal systems capable of operating simultaneously in zebrafish. We implemented multichannel CRISPR recording using up to three CRISPR systems, and show that LbaCas12a may provide superior information density compared to previous methods. We also demonstrate that type II Acrs (anti-CRISPRs) are effective inhibitors of SpyCas9 in zebrafish. These results indicate that at least five CRISPR-Cas systems and two anti-CRISPR proteins are functional in zebrafish embryos. These orthogonal CRISPR-Cas systems and Acr proteins will enable combinatorial and intersectional strategies for spatiotemporal control of genome editing and genetic recording in animals.

overall_design: amplicon sequencing of genomic barcodes edited with SpyCas9, SauCas9 and/or LbaCas12a, and collected from single embryos.

first: Kimberly

middle: T

last: Truong


title: Transcriptional profiling of CD5lo and CD5hi naïve CD4 T cells at homeostasis and upon removal of tonic TCR signaling [bulk RNA-seq]

status: Public on Oct 18 2021

city: Montreal

first: Judith

last: Mandl

state: Quebec

address: 3649 Promenade Sir William Osler

country: Canada

institute: McGill University

summary: Transcriptional characterization of CD5lo and CD5hi naïve CD4 T cells at homeostasis and upon deprivation of self-pMHC interactions.

overall_design: Bulk RNA-seq profiling of FACS-sorted primary CD5lo and CD5hi naïve CD4 T cells and Foxp3+ Tregs. At homeostasis (CD5lo, CD5hi, and Treg) and 22hr after removing self-pMHC interactions (CD5lo and CD5hi). 4 biological replicates were analyzed for each of the Foxp3+ Tregs, fresh CD5lo and CD5hi, and rested CD5lo and CD5hi naïve CD4 T cell subsets.

middle: N

first: Dakota

last: Rogers


title: Single cell analysis of protein epitopes and chromatin accessibility using PHAGE-ATAC [CD8_Hashing]

first: Caleb

last: Lareau

overall_design: Four donors' CD8 T-cells were hashed using uniquely barcoded anti-CD8 phages, pooled, and then processed using a modified scATAC-seq protocol. Hashing concordance with donor genotype was measured and assessed (see CD8_Human_T-cell_Hashing_barcode-assignments.csv.gz).

first: Evgenij

last: Fiškin


title: Transcriptome wide edits after split-engineered base editor treatment

first: Rahul

middle: M.

last: Kohli

state: Pennsylvania

address: 3610 Hamilton Walk, 502B Johnson Pavilion

department: Department of Medicine

institute: University of Pennsylvania

summary: We report transcriptome wide edits comparison between split-engineered base editors and intact base editors. Our results show that, split-engineered base editors show backgound levels of unique C>U edits when compared to intact base editors.

overall_design: Examination of SNPs

first: Kiara

last: Berríos

middle: M

first: Junwei

last: Shi


title: E. coli SymE-SPA binds DNA in a wide variety of genomic locations in vivo [ChIP-seq]

status: Public on Oct 16 2021

city: Cambirdge

first: Mary

middle: Katherine

last: Thompson

state: MA

address: 31 Ames St Room 68-570

department: Biology

institute: Massachusetts Institute of Technology

summary: The purpose of this experiment was to demonstrate that SymE-SPA binds DNA in vivo when overexpressed, and identify where in the genome SymE-SPA binds. To do this SymE-SPA and an untagged SymE control were expressed, and areas of the genome enriched for SymE-SPA binding were identified.

overall_design: 3 replicates of SPA-tagged SymE samples after 30 minutes of induction, 3 replicates of untagged SymE after 30 minutes of induction, 1 replicate of SPA-tagged SymE sample after 60 minutes of induction, 1 replicate of untagged SymE sample after 60 minutes of induction; 8 samples total

middle: K

first: Michael

last: Laub


title: There is no evidence that E. coli SymE cleaves RNA [RNA-seq]

summary: The purpose of this experiment was to determine whether or not we could detect cleavage in mRNA extracted from cells that expressed high levels of the E. coli protein SymE. This was done by comparing read density across transcripts from +symE and empty vector samples after 0, 30 and 90 minutes of induction.

overall_design: 3 replicates of +symE and 3 replicates of empty vector after 0 minutes of induction, 3 replicates of +symE and 3 replicates of empty vector after 30 minutes of induction, 3 replicates of +symE and 3 replicates of empty vector after 90 minutes of induction. Total: 18 samples

first: Peter

last: Culviner


title: Exploration of RNA-Seq data to identify a potential pathogen of the leaf-mining moth, Stomphastis sp. (Lepidoptera: Gracillariidae)

city: St Lucia

first: Kayvan

last: Etebari

state: QLD

address: Building No 8

department: School of Biological Sciences

institute: The University of Queensland

summary: The leaf-mining moth (Stomphastis sp.) was imported to Australia as a potential biological control agent of a weed, Bellyache bush (Jatropha gossypiifolia). The insect colony has been maintained in the quarantine facility for over eight years. Recently, significant mortality was observed in the population. The larvae demonstrated swollen intersegments with a fragile integument. The infected larvae are cloudy muted green or yellowish whereas a healthy late instar larva is a vivid green. They slowly dehydrate and eventually die and when death occurs, the larval body becomes rubbery and turns to black. We used next generation sequencing approach to identify the cause of mortality in the population. Total RNA was extracted from 20 larvae in two cohorts with and without apparent symptoms of disease for deep sequencing on NovaSeq platform after eukaryote ribosomal RNA depletion. We identified several non-insect sequences belonging to viruses, bacteria and fungi but none of those showed significant abundance or enrichment in the infected dataset. The sequences related to a unicellular yeast, Saccharomyces cerevisiae, are among the highly expressed non-insect contigs and more than 5% of reads in both libraries mapped to the genome of this opportunistic microorganism.

overall_design: Total RNA was extracted from 20 Stomphastis larvae in two cohorts with and without apparent symptoms of unknown disease in the laboratory population.

first: Dianne

last: Taylor

first: M

last: Rahman

first: Kunjithapatham

last: Dhileepan

last: Furlong

first: Sassan

last: Asgari


title: Bulk RNA-seq of MHV-A59 infection in primary murine macrophages

city: Beijing

first: Zeming

last: Zhang

address: 38 Xueyuan Rd , Haidian District,Beijing,China

institute: Institude of Systems Biomedicine

summary: Primary mouse peritoneal macrophages and bone marrow derived macrophages were isolated and infected with MHV-A59.

overall_design: Cells were infected with MHV-A59 at 0.1 MOI for 12 hours. Cells were then lysed with Trizol reagent followed by total RNA isolation and bulk RNA sequencing. Two samples per group.

first: Huawei

last: Xia

first: Fuping

last: You


title: CUT&Tag-BS: an efficient and low-cost method for simultaneous profiling of histone modification and DNA methylation

city: RTP

first: ruifang

last: li

state: NC

address: 111 T.W. Alexander Drive

institute: NIEHS

type: Methylation profiling by high throughput sequencing

summary: It remains a challenge to decipher the complex relationship between DNA methylation, histone modification, and the underlying DNA sequence with limited input material. Here, we developed an efficient, low-input, and low-cost method for simultaneous profiling of genomic binding sites of histone modification and methylation status of the underlying DNA at single-base resolution from the same cells in a single experiment by integrating CUT&Tag with tagmentation-based bisulfite sequencing (CUT&Tag-BS). We demonstrated the validity of our method for both active and repressive histone modifications using 250,000 mouse ESCs. CUT&Tag-BS showed similar enrichment patterns of histone modification to those observed in non-bisulfite-treated control; it further revealed that H3K4me1-marked regions are mostly CpG-poor, lack of methylation concordance, and exhibit prevalent DNA methylation heterogeneity among the cells. We anticipate that CUT&Tag-BS will be widely applied to directly address the genomic relationship between DNA methylation and histone modification, especially in low-input scenario with precious biological samples.

overall_design: CUT&Tag-BS was performed for H3K4me1, H3K9me3, and H3K27me3 using 250,000 (or fewer) mouse E14 embryonic stem cells. The resulting libraries were sequenced with either paired-end 35-bp (run1) or 75-bp reads (run2). To benchmark our method, we also prepared corresponding non-bisulfite-treated CUT&Tag libraries as controls.

first: Ruifang

last: Li

first: Sara

last: Grimm


title: Lnc-SLC15A1-1 Up-regulates CXCL10/CXCL8 Expression in Endothelial Cells by Sponging MicroRNAs (RNA-Seq)

city: Taipei

first: yu chin

last: Huang

address: No.1 Jen Ai road section 1

country: Taiwan

institute: NTU

summary: Cardiovascular disease (CVD) is the leading cause of mortality in diabetes mellitus (DM). However, the molecular factors that cause this disproportiona increase in CVD in the DM/chronic kidney disease (CKD) population are still unknown.Human endothelial cells treated with high glucose to mimic DM and with the uremic toxin indoxyl sulfate (IS) to mimic the endothelial injury associated with CKD. Differentially expressed lncRNAs in these conditions were analyzed by RNA sequencing.Lnc-SLC15A1-1 expression was significantly increased upon IS treatment versus high glucose alone.

overall_design: HUVECs were purchased from Lonza Walkersville, Inc. Cells from the fifth passage were incubated in 30 mM D-glucose (high glucose, HG) or 30 mM D-glucose treated with 0.1 mM IS (HG+IS)  for 96 h.Total RNAs was extractied form the HUVECS with HG and HUVECs.Total RNA was extracted from cultured cells  in these two conditions then RNA-seq was performed.

first: Yu-Chin

first: Liang-Chuan

last: Lai

first: Pin‑Hao

last: Ko


title: Single cell RNA sequencing of lung myeloid cells from aged (20-22 months old) mice treated with Isotype or anti-IFNAR1 antibody for 1 day or 1 week

first: Qi

last: Yang

summary: Monocytes and neutrophils are an important subset of myeloid cells in the lung. However, little is known about how aging affects these lung cells, and the role of Type I interferon signaling. Here, we treat aged (20-22 months old) mice with 500ug anti-IFNAR1 antibody (Clone MAR1-5A3 from BioXcell) intraperitoneally (i.p.) once or for 1 week (total 4 i.p. injections, one every other day) and sorted CD45+ CD11b+ Gr1+ cells from the lung the day after the last injection. We performed single cell RNA sequencing to identify the role of Type I Interferon in these cells from the aging mouse lung.

overall_design: We use 10x Genomics platform to prepare scRNA-seq libraries to detail the different gene expression between CD45+ CD11b+ Gr1+ cells from lungs of aged mice treated with Isotype or anti-IFNAR1 antibody

first: Shanti

last: D'Souza


title: Aging reduces liver resiliency by dysregulating Hedgehog signaling

status: Public on Oct 15 2021

city: Raleigh

first: Raquel

last: Maeso-Diaz

state: United States

address: 3901 Sue Lane

institute: Duke University

summary: Older age is a major risk factor for damage to many tissues, including liver. Aging undermines resiliency (i.e., the ability to recover from injury) and impairs liver regeneration. The mechanisms whereby aging reduces resiliency are poorly understood. Hedgehog is a signaling pathway with critical mitogenic and morphogenic functions during development. Recent studies indicate that Hedgehog regulates metabolic homeostasis in adult liver. The present study evaluates the hypothesis that Hedgehog signaling becomes dysregulated in hepatocytes during aging, resulting in decreased resiliency and therefore, impaired regeneration and enhanced vulnerability to damage. Methods: Partial hepatectomy (PH) was performed on young and old wild type mice and Smoothened (Smo)-floxed mice treated with AAV8-TBG luciferase (control) or AAV8-TBG-Cre vectors to conditionally delete Smo and disrupt Hedgehog signaling specifically in hepatocytes. Changes in signaling were correlated with changes in regenerative responses and compared among groups.  Results: Old livers had fewer hepatocytes proliferating after PH. RNA sequencing identified Hedgehog as a top down-regulated pathway in old hepatocytes before and after the regenerative challenge. Deleting Smo in healthy young hepatocytes before PH prevented Hedgehog pathway activation after PH and inhibited regeneration. GO analysis demonstrated that both old and Smo-deleted young hepatocytes had activation of pathways involved in innate immune responses and suppression of several signaling pathways that control liver growth and metabolism including insulin-like growth factor, Wnt and NOTCH. Hedgehog inhibition promoted telomere shortening and mitochondrial dysfunction in hepatocytes, consequences of aging that promote inflammation and impair tissue growth and metabolic homeostasis.  Conclusion: Hedgehog signaling is dysregulated in old hepatocytes.  This accelerates aging, resulting in decreased resiliency and therefore, impaired liver regeneration and enhanced vulnerability to damage.

overall_design: Global transcriptome profiling of was performed by RNA-seq using freshly isolated hepatocytes from young and old wild type mice or Smo-floxed mice before and at 48 h after PH (n=3wild type mice/group/time point), n=1 Smo-flox-Luci mice/time point and n=2 Smo-flox-Cre mice/time point).

last: Maeso-Díaz

first: George

middle: D

last: Dalton

first: SehHoon

last: Oh

first: Kuo

last: Du

first: Linda

last: Tang

first: Tianyi

last: Chen

first: Jessica

last: Hartman

first: Joel

last: Meyer

first: Anna

last: Diehl


title: Transcriptional analysis of noncanonical and canonical Tregs from ITK deficient mice (ITK KO)

city: Syracuse

first: Mobin

last: Karimi

state: NY

address: 766 Irving Ave Suite 2281

department: Microbiology/Immunology

institute: Suny Upstate Medical University

summary: Regulatory T cells (Tregs) play a critical role in controlling autoimmunity and limiting tissue destruction and inflammation. IL2-inducible T cell kinase (Itk) belongs to the Tec family of tyrosine kinases and is an important component of TCR-mediated signaling. Here we showed that either genetic ablation of Itk signaling, or inhibition of Itk signaling pathways resulted in increased frequency of “noncanonical” CD4+CD25−FoxP3+ Tregs (ncTregs), as well as of “canonical” CD4+CD25+FoxP3+ Tregs (canTregs). Using in vivo models, we showed that ncTregs can prevent the development of acute graft-versus-host disease (GVHD), in part by reducing conventional T cell proliferation, proinflammatory cytokine production, and tissue damage. This reduction in GVHD occurred without disruption of graft-versus-leukemia (GVL) effects. RNA sequencing revealed that a number of effector, cell adhesion, and migration molecules were upregulated in Itk-/-  ncTregs. Furthermore, disrupting the SLP76: ITK interaction using a specific peptide inhibitor led to enhanced Treg development in both mouse cells and primary human cells. This peptide inhibitor also significantly reduced inflammatory cytokine production in primary GVHD patient samples and mouse T cells without causing cell death or apoptosis. We provide evidence that specifically targeting Itk signaling could be a therapeutic strategy to treat autoimmune disorders

overall_design: We analyzed FACS purified canonical (CD25+, FOXP3+) and noncanonical (CD25-, FOXP3+) Tregs of 3 different ITK deficient (ITK KO) mice and canonical (CD25+, FOXP3+) Tregs of 3 different  WT mouse.

first: Mahinbanu

last: Mammadli

first: Adam

last: Waickman


title: Single cell RNA sequencing of myeloid cells from lungs of young and aged mice

summary: Single cell RNA sequencing from lungs of young (2-3 month old) and aged (20-22 month old) mice.

overall_design: Myeloid cells (CD45+ Gr1+ CD11b+) were sorted from lungs of young and aged mice that were perfused. Cells from young mice (2-3 months old) were labeled with TotalSeq-C0951 SA antibody (Barcode sequence AACCTTTGCCACTGC), and cells from aged mice (20-22 months old) were labeled with TotalSeq-C0952 SA antibody (Barcode sequence GTCCGACTAATAGCT). Cells were then pooled for scRNAseq preparation.


title: ADAR1 restricts ZBP1–mediated immune response and PANoptosis to promote tumorigenesis

city: Memphis

first: Geoffrey

last: Neale

state: TN

address: 262 Danny Thomas Place

department: Hartwell Center

institute: St Jude Childrens Research Hospital

summary: Cell death provides host defense and maintains homeostasis. Zα-containing molecules are essential for these processes. ZBP1 activates inflammatory cell death, PANoptosis, while ADAR1 serves as an RNA editor to maintain homeostasis. Here, we identify and characterize ADAR1’s interaction with ZBP1, defining its role in cell death regulation and tumorigenesis. Combining IFNs and nuclear export inhibitors (NEIs) activates ZBP1–dependent PANoptosis. ADAR1 suppresses PANoptosis by interacting with the Zα2 domain of ZBP1 to limit ZBP1 and RIPK3 interactions. Adar1fl/flLysMcre mice are resistant to development of colorectal cancer and melanoma, but deletion of the ZBP1 Zα2 domain restores tumorigenesis in these mice. In addition, treating wildtype mice with IFN-γ and the NEI KPT-330 regresses melanoma in a ZBP1–dependent manner. Our findings suggest that ADAR1 suppresses ZBP1–mediated PANoptosis, promoting tumorigenesis. Defining the functions of ADAR1 and ZBP1 in cell death is fundamental to inform therapeutic strategies for cancer and other diseases.

overall_design: We used microarrays to investigate global expression profiles in bone marrow derived macrophages from WT (Adar1fl/+LysMcre) and Adar1-deficient (Adar1fl/flLysMcre) mice

first: Rajendra

last: Karki

first: Balamurugan

last: Sundaram

first: Bhesh

last: Sharma

first: SangJoon

last: Lee

first: R

last: Malireddi

first: Lam

last: Nguyen

first: Shelbi

last: Christgen

first: Min

last: Zheng

first: Yaqiu

last: Wang

first: Parimal

last: Samir

first: Peter

last: Vogel

first: Thirumala-Devi

last: Kanneganti


title: The FASTK family proteins are master regulators in mitochondrial RNA processing

city: Cambridge

first: Lindsey

last: Van Haute

address: Hills Road

department: MRC Mitochondrial Biology Unit

country: United Kingdom

institute: University of Cambridge

summary: Investigation of roles of the FASTK protein family in non-canonical mitochondrial RNA processing

overall_design: HITS-CLIP to find binding sites of FASTKD4

first: Akira

last: Ohkubo


title: Chromatin landscapes of CD5lo and CD5hi naïve CD4 T cells at homeostasis and upon removal of tonic TCR signaling [ATAC-seq]

first: Judith

last: Mandl

summary: Open chromatin accessibility analysis of CD5lo and CD5hi naïve CD4 T cells at homeostasis and upon deprivation of self-pMHC interactions.

overall_design: Bulk ATAC-seq profiling of FACS-sorted primary CD5lo and CD5hi naïve CD4 T cells. 2 biological replicates were analyzed for each of the CD5lo and CD5hi naïve CD4 T cell subsets at each timepoint.

first: Dakota

last: Rogers


title: Single cell RNA sequencing analysis of the mouse colonic mesenchyme (Drop-seq)

status: Public on Oct 14 2021

city: New Haven

first: Jun

last: Zhao

state: CT

address: 300 George Street

department: Pathology/Immunology

institute: Yale University

summary: Single cell RNA sequencing of the mouse colonic mesenchyme. Mesenchymal/lamina propria cells were isolated from the middle and distal colon of wild type mice and were pooled. The vast majority of intestinal epithelial cells were depleted by Ethylenediaminetetraacetic acid (EDTA) treatment of the tissue samples and mesenchymal/lamina propria cells were isolated after enzymatic treatment with collagenase XI and dispase. Single cell RNA sequencing was performed with the Drop-seq protocol.

overall_design: One sample preparation comprising a pool of three biological replicates

first: Richard

last: Flavell

first: Manolis

last: Roulis


title: Autocrine vitamin D signaling switches off pro-inflammatory programs of Th1 cells

city: Bethesda

first: Behdad (Ben)

last: Afzali Khoshkbijari

address: 10 Center Drive, 10/8D03

department: Kidney Diseases Branch

institute: National Institutes of Diabetes, Digestive, and Kidney Diseases

summary: The molecular mechanisms governing orderly shutdown and retraction of CD4+ T helper (Th)1 responses remain poorly understood. Here, we show that complement triggers contraction of Th1 responses by inducing intrinsic expression of the vitamin D (VitD) receptor (VDR) and the VitD-activating enzyme CYP27B1, permitting T cells to both activate and respond to VitD. VitD then initiated transition from pro-inflammatory IFN-γ+ Th1 cells to suppressive IL-10+ cells. This process was primed by dynamic changes in the epigenetic landscape of CD4+ T cells, generating super-enhancers and recruiting several transcription factors, notably c-JUN, STAT3 and BACH2, which together with VDR shaped the transcriptional response to VitD. Accordingly, VitD did not induce IL-10 in cells with dysfunctional BACH2 or STAT3. Bronchoalveolar lavage fluid CD4+ T cells of COVID-19 patients were Th1-skewed and showed de-repression of genes down-regulated by VitD, either from lack of substrate (VitD deficiency) and/or abnormal regulation of this system.

overall_design:
Memory CD4+ cells were isolated by negative selection and treated with 1,25-VitD (10nM) or carrier alone for set amounts of time. Cells were prepared for RNA-seq of poly-adenylated RNA, CUT&RUN against H3K27Ac, c-JUN, and non-specific targeting (IgG), or CUT&Tag against VDR, STAT3, BACH2, and IgG.

RNA-sequencing and analysis
Memory CD4+ T cells, including from BACH2 WT or L24P patients, were activated with VitD or carrier.  600,000 cells were pelleted at 300g for 5 min and RNA extracted by RNAqeous Micro (Thermo Fisher Scientific). 1µg total RNA for each sample was subjected to NEBNext Poly(A) mRNA Magnetic Isolation (E7490) and resulting mRNA prepared for RNA-seq by NEB Ultra II (New England Biolabs) and sequenced by HiSeq (Illumina).

The expression levels of all genes in RNA-seq libraries were quantified by ‘rsem-calculate-expression’ in RSEM/v1.3.1 with parameters ‘--bowtie-n 1 --bowtie-m 100 --seed-length 28 --bowtie-chunkmbs 1000’. The bowtie index for RSEM alignment was generated by ‘rsem-prepare-reference’ on all RefSeq genes, downloaded from UCSC table browser in April 2017. EdgeR/v3.26.8 was used to normalize gene expression among all libraries and identify DEGs among samples. Microarray analysis sourced from GSE119416 was carried out using Partek Genomics Suite (Partek, Inc.). DEGs were defined using the following criteria: at least 1.5-fold change in either direction at p-value<0.05 for microarray; at least 1.75-fold change in either direction at FDR<0.05 for RNA-seq.

H3K27Ac and c-JUN CUT&RUN
CUT&RUN sequencing was performed on memory CD4+ T cells activated as above in the presence of carrier or VitD for 0.75h, 48h and 72h using the published protocol of Skene et al (Nature Protocols 2018). Antibodies against H3K27Ac (ab4729, Abcam), c-JUN (60AB, Cell Signaling Technologies), and non-specific IgG (31235, Thermo Fisher Scientific) were used with pAG-MNase (123461, Addgene) on 50,000 cells per target.  CUT&RUN (and CUT&Tag below) buffer components were purchased from Sigma-Aldrich and Thermo Fisher Scientific. Post-CUT&RUN, short DNA fragments were prepared for paired-end sequencing (NEB Ultra II, New England Biolabs) with NovaSeq (Illumina). Amplified libraries were quantified by high sensitivity fluorometry (DeNovix) and sized via the 4200 TapeStation (Agilent Technologies).

H3K27Ac reads aligned to the human reference genome (GRCh37; hg19) using Bowtie2 with parameters ‘--local --maxins 250’ with sorting and indexing of aligned reads by samtools/v1.10, reads with mapping quality <30 removed. H3K27Ac peaks were called by MACS2 with parameter ‘-f BAMPE --nomodel’. JUN reads were mapped without ‘--local’ parameter.  <120 bp fragments were further sorted and indexed by samtools. JUN induced peaks called by MACS2 parameters ‘--nomodel -g hs -f BAMPE -q 0.01 --SPMR --keep-dup all’. IgG BAM files were included as controls for MACS2. Peaks for H3K27Ac and JUN experiments were screened against previously characterized hypersequencable-regions (Hg19 Blacklist, CutRunTools). H3K27ac and JUN mean binding intensities in peak regions calculated by UCSC bigWigAverageOverBed. Peaks, combined from two conditions, with mean binding intensity greater than 0.2 in at least one condition and fold change of signal>1.5 were differential. Known motifs +/−100bp from differential peak summits were identified (HOMER, findMotifsGenome.pl, parameter ‘-size given’). De novo motif footprinting for c-JUN was performed (CutRunTools). Average cuts per bp proximal to JUN peaks was plotted as a histogram relative to IgG. CUT&RUN tracks and heatmaps were visualized using IGV (Broad Institute) and deepTools, respectively.

VDR, STAT3, and BACH2 CUT&Tag
VDR, STAT3, and BACH2 genome-wide binding in memory CD4+ T cells activated in the presence of 10nM VitD or carrier for 96h was detected by CUT&Tag as per the published protocol (CUT&Tag V.3, dx.doi.org/10.17504/protocols.io.bcuhiwt6) using fixed nuclei (2min fixation of isolated nuclei, RT) with 75,000 nuclei/target and antibodies targeting either VDR (D2K6W, Cell Signaling Technologies), STAT3 (D3Z2G, Cell Signaling Technologies), BACH2 (D3T3G), or non-specific IgG (31235) diluted 1:50 accordingly and incubated with fixed-nuclei for 1h, RT. Secondary antibody (1:100) (ABIN101961, antibodies-online) was bound for 0.5h, RT. Preloaded pA-Tn5 (C01070001, Diagenode) was tethered to antibodies for 1h,  RT. Libraries were sequenced paired-end by Novaseq. Resulting reads were processed and mapped to Hg19 (CutRunTools), with peaks called (SEACR/v1.2, stringent, all mapped fragments) and hypersequencable-regions subtracted. Known motif enrichment was determined +/−200bp from peak summits and data visualized as in CUT&RUN.

first: Daniel

last: Chauss

first: Tilo

last: Freiwald

first: Reuben

last: McGregor

first: Bingyu

last: Yan

first: Luopin

first: Estefania

last: Nova-Lamperti

first: Dhaneshwar

last: Kumar

first: Zonghao

last: Zhang

first: Heather

last: Teague

first: Erin

last: West

first: Kevin

last: Vanella

first: Marcos

last: Ramos-Benitez

first: Jack

last: Bibby

first: Audrey

last: Kelly

first: Amna

last: Malik

first: Alexandra

middle: F

last: Freeman

first: Daniella

last: Schwartz

first: Didier

last: Portilla

last: Chertow

first: Susan

last: John

first: Paul

last: Lavender

first: Michail

last: Lionakis

first: Nehal

last: Mehta

first: Claudia

last: Kemper

first: Nichola

last: Cooper

first: Giovanna

last: Lombardi

first: Arian

last: Laurence

first: Majid

last: Kazemian

first: Behdad

last: Afzali


title: Microarray analysis of Tm6sf2 E167K knockin mouse liver

city: Ann Arbor

first: Haocheng

last: Lu

state: Michigan

address: 2800 plymouth road

department: Internal Medicine

institute: University of Michigan

summary:
The rs58542926 C>T variant of the transmembrane 6 superfamily member 2 gene (TM6SF2), encoding an E167K amino acid substitution, is associated with liver disease and cardiovascular disease.
Therefore, we determined the long-term effects of this coding variant of TM6SF2 on glucose metabolism in mice with CRISPR/Cas9-mediated knock-in of TM6SF2 E167K (Tm6sf2 KI) and compared to littermate wild-type mice.

overall_design: To elucidate the molecular mechanisms that mediate the effects of the Tm6sf2 E167K variant on glucose and lipid metabolism, we performed microarray analysis in liver samples from the Tm6sf2 KI and Wt male mice fed a a high fat-diet (HFD) for 20 weeks.

first: Yanbo

last: Fan


title: The histone lysine acetyltransferase HBO1 (KAT7) regulates hematopoietic stem cell quiescence and self-renewal

city: Parkville

first: Waruni

last: Abeysekera

state: Victoria

address: 1G Royal Parade

institute: Walter and Eliza Hall Institute of Medical Research

summary: In this study, we have used inducible and tissue-specific genetic deletion to investigate the function of HBO1 in the hematopoietic system. RNA-seq was used to examine the dependence of gene expression on the presence of HBO1(KAT7). Two different conditional expression sytems were used to induce Cre recombinase and delete a floxed allele of HBO1 in this study.  This was to control for the treatement effect during Cre induction (interferon induction of Mx1-cre vs. tamoxifen induction of Rosa26-CreERT2).  This data set indicates that HBO1 promotes expression of multiple genes encoding a hematopoietic transcription factor network essential for HSC quiescence and self-renewal in adult hematopoiesis.

overall_design: 4 induced homozygous mutant vs. 4 induced heterozygous controls per specific  cre-recombinase and per cell type (2 methods of deletion, 2 cells populations) => (4+4)x2x2=32 samples

first: Yuqing

last: Yang

first: Tim

last: Thomas

first: Anne

last: Voss

first: Gordon

last: Smyth


title: Transcriptional and chromatin landscape profiles of naïve CD4 T cells

first: Judith

last: Mandl

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

overall_design: Refer to individual Series


title: Copy number variation analysis of desmoplastic malignant mesothelioma

city: Tokyo

first: Rei

last: Noguchi

state: Tokyo

department: Division of Rare Cancer Research

institute: National Cancer Center Research Institute

type:
Genome variation profiling by SNP array
SNP genotyping by SNP array

summary: Desmoplastic malignant mesothelioma is a rare tumor. Due to the rarity, genomic profile of desmoplastic malignant mesothelioma is not unveiled. To elucidate genomic profile of desmoplastic malignant mesothelioma, we used illumina infinium omini exomeexpress in an established patient-derived cell line of desmoplastic malignant mesothelioma.

overall_design:
A sample was analyzed on Illumina infinium OmniExpressEsome genotyping bead array. Desmoplastic malignant mesothelioma cells from one patient with desmoplastic malignant mesothelioma were examined.
Genomic DNA extracted from patient-derived cell line  derived desmoplastic malignant mesothelioma was genotyped using Infinium OmniExomeExpress-8 Chips (Illumina).

first: Tadashi

last: Kondo


title: Fully characterized mature human iPS- and NMP-derived motor neurons thrive without neuroprotection in the spinal contusion cavity [SMN V3]

city: Albany

first: Janet

last: Paluh

address: CNSE, Nanofab East, 257 Fuller Road

department: Nanobioscience

institute: SUNY Polytechnic Institute

summary: Unpublished findings related to Frontiers 2021 article: Fully characterized mature human iPS- and NMP-derived motor neurons thrive without neuroprotection in the spinal contusion cavity

overall_design:
RNA-Seq of day10 NSC rosettes; day19, 25, Motor Neuron Progenitors; Day 28, 32, 37, 42 Spinal Motor Neurons of humanAfrican American hiPSC line F3.5.2.

The Paluh-HKYW5AFXY.featureCounts.tsv contains processed data for both GSE178600 and GSE178601.

first: Zachary

last: Olmsted


title: CXCL10 regulates heterogenity of the CD8+ T cell response and viral set point during chronic infection

status: Public on Oct 13 2021

city: Charlestown

first: Aleksandra

last: Ozga

address: 149 13th Street

institute: Massachusetts General Hospital and Harvard Medical School

summary: CD8+ T cell responses to chronic infection are sustained by stem-like cells, which differentiate into effector-like cells mediating some level of pathogen control; however persistent antigen progressively impairs their effector functions. Understanding the molecular control of stem-like cell differentiation to improve durability of functional T cell responses has important therapeutic implications. Here, we found that the chemokine receptor CXCR3 was highly expressed on viral-specific stem-like CD8+ T cells and that one of its ligands, CXCL10, regulated the persistence and heterogeneity of responding CD8+ T cells in spleens of mice chronically infected with lymphocytic choriomeningitis virus. CXCL10 was produced by inflammatory monocytes and fibroblasts of the splenic red pulp where it granted stem-like cells access to signals promoting differentiation and limited their exposure to pro-survival niches in the white pulp. Consequently, the magnitude of functional CD8+ T cell responses was greater in Cxcl10-/- mice and was associated with a lower viral set point.

overall_design: Four CD8+ T cell subsets distinguished based on the Ly108, CX3CR1 and CD101 expression sorted from WT or Cxcl10-/-  are analyzed. Each subset has at least 3 biological replicates

first: Melvyn

last: Chow

first: Mateus

middle: E

last: Lopes

first: Rachel

last: Servis

first: Mauro

last: Di Pilato

first: Philippe

last: Dehio

first: Jeffrey

last: Lian

first: Thorsten

last: Mempel

first: Andrew

last: Luster


title: The transcriptome data of primary microglia cells from contrl and over expression CYP11A1 rat's offspring

city: chengdu

first: li

last: tongtong

state: sichuan

address: sichuan

institute: xinong

summary: NOT PROVIDED; REQUESTED

overall_design: over expression CYP11A1 in female rats, and extract microglia from their offspring, and perform transcriptome sequencing

first: Xu

last: Wenming


title: Identification of enhancer-promoter contacts in squamous cancer cells

status: Public on Oct 17 2021

city: Salt Lake City

first: Xiaoyang

last: Zhang

address: 1950 CIrcle of Hope

institute: University of Utah


title: Effects of murine Oncostatin M on OP9 stromal cells and NIH/3T3 fibroblasts

city: Freiburg

first: Justus

last: Duyster

address: Hugstetter Str. 55

department: Department of Hematology and Oncology

institute: Freiburg University Medical Center

summary: The IL-6 cytokine Oncostatin M (OSM) is involved in cell development, growth, hematopoiesis, inflammation and cancer. Intriguingly, OSM has proliferative and antiproliferative effects de-pending on the target cell. The molecular mechanisms underlying these opposing effects are not fully understood and its role in tumorigenesis remains controversial. In the present study, we investigated the effect of murine OSM (mOSM) on proliferation in two cell types of distinct origin. We found that mOSM impairs the proliferation of murine bone marrow (BM) stromal cell lines and primary BM stromal cells, whereas murine fibroblasts responded to mOSM with increased proliferation. When we set out to reveal the mechanisms underlying these opposing effects of OSM on proliferation, we detected increased expression of the OSM receptors OSMR and LIFR in stromal cells. Interestingly, Osmr knockdown and Lifr overexpression attenuated the OSM-mediated effect on proliferation in both cell lines indicating that mOSM affected the pro-liferation of both cell lines signaling mainly through the OSMR. Furthermore, mOSM induced activation of the JAK-STAT, PI3K-AKT and MAPK-ERK pathways in OP9 and NIH/3T3 cells with differences in total protein levels between the two cell lines. Our findings offer new insights into the regulation of proliferation by mOSM.

overall_design: We included 12 samples in total. 3x untreated OP9 cells, 3x OP9 cells + 10 ng/mL of OSM, 3x untreated NIH/3T3 cells, 3x NIH/3T3 cells + 10 ng/mL of OSM.

first: Lena

last: Jakob

first: Dietmar

last: Pfeifer


title: Gene expression profiling in Nlrp3 deficient mice infected with Francisella tularensis

city: Greenville

first: Weihua

last: Huang

state: North Carolina

address: 600 Moye Blvd, Brody 7S-24

department: Pathology

institute: East Carolina University

summary: This study investigated gene expression profile in wild type and nlrp3 deficient mice infected with F. tularensis LVS. RNA-sequencing was performed to determine the transcript levels of genes using total RNA extracted from lungs of wild type C57BL/6 and Nlrp3-/- mice infected with 1 × 104 CFU of F. tularensis LVS on days 3 and 7 post-infection.

overall_design: Wild type C57BL/6 and Nlrp3-/- mice infected intranasally with 1 × 104 F. tularensis LVS were sacrificed on days 3 and 7 post-infection. Lungs were harvested aseptically from these mice and placed in excess of trizol. Organs were snap-frozen in liquid nitrogen and stored at -80°C until required. When ready, the organs frozen in trizol were thawed on ice and homogenized in 2mL of fresh trizol using a Tissue-tearor.  The homogenate was centrifuged at 15,000×g for 15 minutes at 4°C to pellet the tissue debris and limit RNA degradation from tissue-resident nucleases. Total RNA was extracted from the clarified supernatants using the “Purelink RNA mini kit” (Invitrogen) following the manufacturer’s instructions. DNA was eliminated from the samples by DNAse treatment using “RNAse-Free DNAse” (Qiagen). Purified RNA was used for RNA-sequencing.

first: Varadharajan

middle: R

last: Suresh

first: Meenakshi

last: Malik

first: Chandra

last: Bakshi


title: Sex-biased ZRSR2 mutations in myeloid malignancies impair plasmacytoid dendritic cell activation and apoptosis [cell line]

city: Boston

last: Lane

address: 450 Brookline Ave

institute: Dana-Farber Cancer Institute

summary: Blastic plasmacytoid dendritic cell neoplasm (BPDCN) is an aggressive leukemia of plasmacytoid dendritic cells (pDCs). BPDCN occurs at least three times more frequently in men than women, but the reasons for this sex bias are unknown. Here, studying genomics of primary BPDCN and modeling disease-associated mutations, we link acquired alterations in RNA splicing to abnormal pDC development and inflammatory response through Toll-like receptors. Loss-of-function mutations in ZRSR2, an X chromosome gene encoding a splicing factor, are enriched in BPDCN and nearly all mutations occur in males. ZRSR2 mutation impairs pDC activation and apoptosis after inflammatory stimuli, associated with intron retention and inability to upregulate the transcription factor IRF7. In vivo, BPDCN-associated mutations promote pDC expansion and signatures of decreased activation. These data support a model in which male-biased mutations in hematopoietic progenitors alter pDC function and confer protection from apoptosis, which may impair immunity and predispose to leukemic transformation.

overall_design: RNA sequencing of CAL1 BPDCN cells with shRNA knockdown of ZRSR2 or control shRNA at 0, 2, and 7 days after shRNA induction by doxycycline


title: An AraC/XylS Family Transcriptional Regulator Modulates the Oxidative Stress Response of Francisella tularensis

summary: Francisella tularensis is a Gram-negative bacterium that causes a fatal human disease known as tularemia. The Centers for Disease Control have classified F. tularensis as Category A Tier-1 Select Agent. The virulence mechanisms of Francisella are not entirely understood. Francisella possesses very few transcription regulators, and most of these regulate the expression of genes involved in intracellular survival and virulence. The F. tularensis genome sequence analysis reveals an AraC (FTL_0689) transcriptional regulator homologous to the AraC/XylS family of transcriptional regulators. In Gram-negative bacteria, AraC activates genes required for L-arabinose utilization and catabolism. The role of the FTL_0689 regulator in F. tularensis is not known. In this study, we characterized the role of FTL_0689 in gene regulation of F. tularensis and investigated its contribution to intracellular survival and virulence. The results demonstrate that FTL_0689 in Francisella is not required for L-arabinose utilization. Instead, FTL_0689 specifically regulates the expression of the oxidative and global stress response, virulence, metabolism, and other key pathways genes required by Francisella when exposed to oxidative stress. The FTL_0689 mutant is attenuated for intramacrophage growth, and mice infected with the FTL_0689 mutant survive better than wild-type F. tularensis LVS infected mice. Based on the deletion mutant phenotype, FTL_0689 was termed osrR (oxidative stress response regulator). Altogether, this study elucidates the role of the osrR transcriptional regulator in tularemia pathogenesis.

sample_organism: Francisella tularensis subsp. holarctica LVS

overall_design: Comparison of transcript abundance in Francisella tularensis with or without the osrR gene when grown in the absence or the presence of oxidative stress induced by a superoxide generating compound, menadione.

first: Dina

last: Marghani

first: Zhuo

last: Ma


title: dsRNA mimics treatment of Melanoma and Human lymphatic cells

city: Madrid

first: David

last: Olmeda

state: Madrid

address: Melchor Fernandez Almagro 3

department: Molecular Oncology

country: Spain

institute: CNIO

summary: The crosstalk between cancer cells and the lymphatic vasculature has long been proposed to define competency for metastasis. Nevertheless, the discovery of selective blockers of lymphovascular niches has been compromised by the paucity of experimental systems for whole-body analyses of tumor progression. Spatio-temporal analyses in autochthonous melanomas and patient-derived xenografts identified double stranded RNA mimics (dsRNA nanoplexes) as potent repressors of lymphangiogenesis and metastasis. Mechanistically, dsRNA nanoplexes were found to suppress lymphangiogenic drivers in both tumor cells and their associated lymphatic vasculature (via MIDKINE and Vegfr3, respectively). This dual inhibitory action, driven by type I interferon, was not shared by FDA-approved antimelanoma treatments or by lymphangiogenic blockers in clinical testing. These results underscore the power of Vegfr3-lymphoreporters for pharmacological testing in otherwise aggressive cancers

overall_design: Human Lymphatic Microvascular Endothelial Cells (HMVEC-dLy) (Lonza, MD) were grown in EGM™-2 medium supplemented with MV BulletKit (Lonza). SK-Mel-147 melanoma cells were grown in DMEN 10% FBS.   2.5 x 10^5 cells were seeded in MW6 plates 24h before treatment.  When indicated, the cells were treated for 10h with 1 µg/ml BO-110 or vehicle control. Cells were then harvested, and total RNA was extracted. 500ng of total RNA samples were used. Each condition was analyzed in triplicates.  Sequencing libraries were prepared with the "QuantSeq 3‘ mRNA-Seq Library Prep Kit (FWD) for Illumina" (Lexogen, Cat.No. 015) by following manufacturer instructions.

first: Maria

last: Soengas


title: Cut&Tag of ZGA staged embryos

first: Michael

last: Rauer

address: Breisacherstr. 153

department: Medical Faculty

institute: University of Freiburg

summary: During embryogenesis, the genome shifts from transcriptionally quiescent to extensively active in a process known as Zygotic Genome Activation (ZGA). In Drosophila, the pioneer factor Zelda is known to be essential for the progression of development, still, it regulates the activation of only a small subset of genes at ZGA. However, thousands of genes do not require Zelda, suggesting that other mechanisms exist. By conducting GRO-seq, HiC and ChIP-seq in Drosophila embryos, we demonstrate that up to 65% of zygotically activated genes are enriched for the histone variant H2A.Z and independent of Zelda. H2A.Z enrichment precedes ZGA and Polymerase II loading onto chromatin. In vivo knock-down of maternally contributed Domino, a histone chaperone and ATPase, reduces H2A.Z deposition at transcription start sites, causes global downregulation of housekeeping genes at ZGA, and compromises the establishment of the 3D chromatin structure. We infer that maternal factors and H2A.Z are master regulators of the de novo establishment of transcriptional programs during ZGA via chromatin reorganization.

overall_design: Cut&Tag libaries for Ctrl and DomKD ZGA embryos, two replicates per condition

first: Dafne

last: Morales

first: Nicola

last: Iovino


title: Characterization of genomic rearrangements in RPE-1 cells with and without DSB in the ABCB1 promoter

city: Amsterdam

first: Anna

last: Gonzalez Manjon

email: a.gonzalez@nki.nl, li.koob@nki.nl

address: Plesmanlaan 121

country: Netherlands

institute: Netherlands Cancer Institute

summary: The discovery of the Clustered Regularly-Interspaced Short Palindromic Repeats (CRISPR) and its development as a genome editing tool has revolutionized the field of molecular biology. In the DNA damage field, CRISPR has brought an alternative to induce endogenous double-strand breaks (DSB) at desired genomic locations and study the DNA damage response and its consequences. Many systems for sgRNA delivery have been reported in order to efficiency generate this DSB, including lentiviral vectors. However, some of the consequences of these systems are yet not well understood. Here we report that lentiviral-based sgRNA vectors can integrate into the endogenous genomic target location, leading to undesired activation of the target gene. By generating a DSB in the regulatory region of the ABCB1 gene using a lentiviral sgRNA vector, we can induce the formation of taxol-resistant colonies. We show that these colonies upregulated ABCB1 via integration of the EEF1A1 and the U6 promoters from the sgRNA vector. We believe that this is an unreported CRISPR/Cas9 artefact that researchers need to be aware of when using lentiviral vectors for genome editing.

overall_design: We have performed Targeted Locus Amplification (TLA), a chromosome conformation capture-based technique to selectively aplify and sequence DNA flanking the ABCB1 promoter. We employed two cell lines: RPE-1 parental and an RPE-1 clone (sg6C9) with a Cas9-DSB induced in ABCB1 (Sample 1 and 2). We have also performed ChIP seq of RPE-1 parental and seven clones with a DSB-induced in ABCB1 which are taxol resistant: sg6C1, sg6C9, sg11C3, sg11C4, sg11C6, sg12C5, sg11C4 (Samples 3 to 11).

first: Rene

last: Medema


title: Repeated cocaine or methamphetamine treatment alters astrocytic CRF2 and GLAST expression in the ventral midbrain: RNA sequencing data

city: Oklahoma City

last: Beckstead

state: OK

address: 825 NW 13th Street

department: Aging and Metabolism

institute: Oklahoma Medical Foundation

summary: We describe the effects of cocaine and methamphetamine treatment on astrocytic expression of the corticotropin releasing factor 2 receptor and the excitatory amino acid transporter 1 in the ventral midbrain. The RNA-sequencing data confirms the expression of CRF2R on astrocytes using the Aldh1l1-NuTRAP model. This allowed for astrocyte mRNA isolation and characterization, confirming CRF2R expression on ventral midbrain astrocytes.

overall_design: mRNA profiles of naïve Aldh1l1-NuTRAP ventral midbrain astrocytes

first: Amanda

last: Sharpe

first: Marta

last: Trzeciak

first: Nicole

last: Eliason

first: Harris

last: Blankenship

first: Bre'Ana

last: Byrd

first: Phillip

last: Douglas

first: Willard

last: Freeman


title: Interleukin-10 causes interferon γ-dependent emergency myelopoiesis

city: Porto

first: Ana

middle: Raquel

last: Maceiras

address: Rua Alfredo Allen, 208

country: Portugal

summary: In emergency myelopoiesis (EM), expansion of the myeloid progenitor compartment and increased myeloid cell production is observed, often mediated by the pro-inflammatory cytokine IFN-γ. IL-10 inhibits IFN-γ secretion, largely by its effects on macrophages and dendritic cells, but, paradoxically, its therapeutic administration to humans causes hematologic changes similar to those observed in EM. In this work we used different in vivo systems, including a humanized immune system mouse model, to show that IL-10 triggers EM, with a significant expansion of the myeloid progenitor compartment and production of myeloid cells. Hematopoietic progenitors display a prominent IFN-γ transcriptional signature, and we show that IFN-γ mediates IL-10-driven EM. We also found that IL-10, unexpectedly, induces IFN-γ production by all T cell subsets in vivo. Therefore, in addition to its established anti-inflammatory properties, IL-10 can induce IFN-γ production and EM, opening new perspectives for the design of IL-10-based immunotherapies.

overall_design: AmpliSeq Transcriptome was performed in sorted bone marrow CD4 and CD8 T cells from three pMT-10 overexpressing IL-10, three C57BL/6 (control), and three control pMT-10.


title: Nanostring CAR T Characterization panel comparing expression in cord blood-derived transgenic T cells cultured with IL-2 or with IL-7 and IL-15, peripheral blood-derived transgenic T cells cultured with IL-2

city: Besançon

first: Chrystel

last: Marton

address: 8 rue du Dr Jean-François-Xavier Girod

institute: UMR1098 INSERM

summary:
The purpose of the experiment was to transcriptionally characterize transgenic T cells derived from cord blood (CB) and cultured with IL-2 or IL-7 and IL-15, or from peripheral blood (PB) and cultured with IL-2.
The sample replicates were generated using peripheral and cord blood mononuclear cells, obtained from healthy human donors and cord blood units that were not compliant with banking standards, respectively. T cells were then transduced with a retroviral vector encoding an HLA-DRB1*04-restricted TCR targeting the human papillomavirus oncoprotein E7.

overall_design: There are 4 sample replicates for the peripheral blood condition, all of which were cultured with IL-2, and 8 sample replicates for the cord blood condition, among which 4 were cultured with IL-2 and 4 were cultured with IL-7 and IL-15. All experiments were independent.


title: Transcriptional profiling of individual naïve CD4 T cells sorted across the full spectrum of self-reactivity [scRNA-seq]

city: Montreal

first: Judith

last: Mandl

address: 3649 Promenade Sir William Osler

institute: McGill University

summary: Transcriptional characterization of naïve CD4 T cells across the spectrum of self-reactivity at the single cell level

overall_design: scRNA-seq profiling of FACS-sorted primary naïve CD4 T cells  at homeostasis. Individual naïve CD4 T cells were sorted across the full spectrum of self-reactivity using CD5 as a surrogate marker for strength of tonic TCR interactions.

first: Dakota

last: Rogers


title: Microglial-expressed Apoe does not modulate plaque pathogenesis

city: Irvine

first: Kim

last: Green

state: CA

address: 3208 Biological Sciences 3

department: Neurobiology and Behavior

institute: University of California, Irvine

summary: This dataset allows for the exploration of the effect of microglial-expressed Apoe knockout in a 5xFAD Alzheimer's Disease model.

overall_design: Mice hippocampi were sequenced in the following groups at 4 and 12 months of age: 1. Apoe(fl/fl) 2. Apoe(fl/fl) Csf1r-cre 3. Apoe(fl/fl) 5xFAD 4. Apoe(fl/fl) 5xFAD Csf1r-Cre. Microglial-expressed Apoe was knocked out of mice that are homozygous for Apoe(fl/fl) and hemizygous for Csf1r-cre. n=3-8 for each group

first: Caden

last: Henningfield

first: Neelakshi

last: Soni


title: H2A.Z RELACS of ZGA staged embryos

first: Michael

last: Rauer

overall_design: Corresponding Inputs and H2A.Z RELACS ChIP-seq in Ctrl and DomKD ZGA embryos, 2 replicates per condition

first: Dafne

last: Ibarra Morales

first: Nicola

last: Iovino


title: Histone variant H2A.Z regulates zygotic genome activation

overall_design: Refer to individual Series


title: MNase-seq of Ctrl and DomKD ZGA embryos

overall_design: MNase-seq of Ctrl and DomKD ZGA embryos, 2 replicates per condition


title: HiC of Ctrl and DomKD ZGA embryos

overall_design: HiC of Ctrl and DomKD ZGA embryos, 2 replicates per conditon


title: ChIP-seq of ZGA staged embryos

overall_design: Corresponding inputs and ChIP-seqs of H2A.Zand Rpb3 in pre-ZGA embryos(c9-13), 2 replicates per stage. Corresponding inputs and ChIP-seqs of H2A.Z and Rpb3 in ZGA embryos in Ctrl and DomKD embryos, two replicates per condition. Corresponding inputs and ChIP-seq of H3K4me3 and H3K36me3 in ZGA Ctrl embryos, 2 replicates each.


title: RNA-seq of staged embryos

overall_design: RNA-seq of Ctrl (3 replicates) and DomKD (4 replicates) ZGA embryos and RNA-seq of unfertilized Ctrl eggs, 4 replicates.


title: Enhanced glucose metabolism through activation of HIF-1α covers the energy demand in a rat embryonic heart primordium after heartbeat initiation.

city: Sapporo

first: Tatsuya

last: Sato

state: Hokkaido

address: South-1, West-17, Chuo-ku

department: Department of Cellular Physiology and Signal Transduction

institute: Sapporo Medical University School of Medicine

summary: The initiation of heartbeat is an essential step in cardiogenesis in the heart primordium, but it remains unclear how intracellular metabolism responds to increased energy demands after heartbeat initiation.  In this study, embryos in Wistar rats at embryonic day 10, at which heartbeat begins in rats, were divided into two groups by the heart primordium before and after heartbeat initiation and their metabolic characteristics were assessed.  Metabolome analysis revealed that increased levels of ATP, a main product of glucose catabolism, and reduced glutathione, a by-product of the pentose phosphate pathway, were the major determinants in the heart primordium after heartbeat initiation.  Glycolytic capacity and ATP synthesis-linked mitochondrial respiration were significantly increased, but subunits in complexes of mitochondrial oxidative phosphorylation were not upregulated in the heart primordium after heartbeat initiation.  Hypoxia-inducible factor (HIF)-1α was activated and a glucose transporter and rate-limiting enzymes of the glycolytic and pentose phosphate pathways, which are HIF-1α-downstream targets, were upregulated in the heart primordium after heartbeat initiation.  These results suggest that the HIF-1α-mediated enhancement of glycolysis with activation of the pentose phosphate pathway, potentially leading to antioxidant defense and nucleotide biosynthesis, covers the increased energy demand in the beating and developing heart primordium.

overall_design: Gene expression of Embryonic heart primordium was measured at pre-heartbeat and post-heartbeat E10. The experiment was carried out with three biological replicates in each group.

first: Noritsugu

last: Tohse

first: Takeshi

last: Kobayashi

first: Nobutoshi

last: Ichise


title: Single-cell architecture and functional requirement of alternative splicing during hematopoietic stem cell formation

city: Beijing

first: Jia

last: Yu

address: Dong Dan San Tiao 5, Dongcheng District

institute: Peking Union Medical College

summary: Hematopoietic stem cells (HSCs) are well known to generate from a rare hemogenic population of embryonic endothelia cells through a dynamic fate transition, accompanied by widespread changes in gene regulatory networks. Nevertheless, RNA splicing, a striking feature in shaping transcriptome and enabling the generation of vast transcripts and diverse proteins, has never been illustrated in this transient endothelial to HSC conversion. Here, we constructed an isoform-based transcriptome landscape during HSC development at single-cell resolution, which enables the identification of hemogenic signature isoforms, as well as stage-specific splicing events. Combining with functional screening, we showed that the inclusion of such hemogenic-specific exons was essential for hemogenic function in vitro. Moreover, their gradual appearance along with HSC specification from endothelia cells was orchestrated by the splicing regulator Srsf2. Therefore, early Srsf2 deficiency from endothelia cells dramatically impaired the HSC generation, which was accompanied by changes in splicing pattern of several master hematopoietic regulators. These results redefine our understanding of the dynamic transcriptome diversity and demonstrate that the elaborate control of splicing governs cell fate in HSC formation.

overall_design: Single-cell full-length RNA-sequencing of two HSC-associated cell populations, and RNA-sequencing of Srsf2 f/+ or Srsf2 f/f and Vec-Cre;Srsf2 f/f cells.

first: Fang

last: Wang

first: Puwen

last: Tan

first: Pengcheng

last: Zhang

first: Yue

last: Ren

first: Jie

last: Zhou

first: Dong

first: Bin

last: Liu

first: Yu

last: Lan


title: ATAC-seq of ZGA staged embryos

overall_design: ATAC-seq for Ctrl and DomKD ZGA embryos, two replicates per condition


title: Single cell transcriptome of human alveolar epithelial cells

city: San Francisco

first: Jaymin

last: Kathiriya

address: 513 Parnassus Ave

department: Medicine

institute: University of California San Francisco

summary: To determine the ability of human AEC2s to transdifferentiate in vitro, we performed single cell mRNA sequencing using 10X Genomics platform of freshly sorted AEC2s and several timepoints in a 3D organoid co-culture (AEC2s with primary human mesenchyme) system.

overall_design: Freshly sorted human AEC2s were co-cultured with primary human mesenchyme in a 3D organoids. At day 7, day 14, and day 21, epithelial cells and mesenchymal cells were harvested and sequenced via 10X Genomics.

first: Chaoqun

first: Harold

last: Chapman

first: Tien

last: Peng


title: Unique functions for Notch4 in murine embryonic lymphangiogenesis

city: Chicago

first: Jan

last: Kitajewski

state: IL

address: 909 S. Wolcott Ave. MC 901

department: Dept. of Physiology and Biophysics

institute: University of Illinois at Chicago

summary: To assess DLL4/Notch LEC signaling, we seeded HdLECs on DLL4FC-coated or control FC-coated plates and collected RNA after 6 hours which was subjected to mRNA sequencing. Relative to the HdLECs seeded on FC-coated plates, DLL4FC significantly altered the expression of 675 genes with a padj <0.05. 69 genes were induced 1.2-fold while 68 genes were suppressed 1.2-fold. Analysis of the top 30 upregulated genes revealed DLL4/Notch signaling upregulated the expression of known direct effectors of Notch signaling, Hey1, EphrinB2, Hes4, Dll4, and Hes1, as well as key lymphangiogenic genes, Gja4 (Cx37), Cxcr4, Gja1 (Cx43), Ccl2, Ackr3, and Sema3g). DLL4/Notch signaling also downregulated lymphangiogenic genes, including Apln and Adm . GO: Biological Pathway (BP) analysis indicates the LEC DLL4/Notch signaling induces genes responsible for pattern specification, neurogenesis and chemotaxis.

overall_design: RNAseq screens from in-vitro activation of Lymphatic Endothelial Cells (LEC)  by tethered DLL4 (6h) or Ig-Fc (6h).

first: Ajit

last: Muley

first: Minji

last: Uh

first: Glicella

last: Salazar-DeSimone

first: Bhairavi

last: Swaminathan

first: Jennifer

last: James

first: Aino

last: Murtomaki

first: Seock-Won

last: Youn

first: Joseph

last: McCarron

first: Chris

first: Maria

last: Gnarra

first: Gloria

last: Riitano

first: Yoh-suke

last: Mukouyama

first: Carrie

last: Shawber


title: Next Generation Sequencing of Ruminicoccus Gnavus after pregnelonone exposure

city: Bellinzona

first: Andrea

last: Alimonti

address: Via vela 6

country: Switzerland

institute: Institute of oncology research

summary: Examination of Ruminicoccus Gnavus gene expression profile after pregnelonone exposure

overall_design: pregnelonone exposure

first: Martina

last: Troiani

first: Marco

last: Bolis


title: Calcitriol ameliorates damage in high-salt diet-induced hypertension: evidence of communication with the gut–kidney axis

status: Public on Oct 12 2021

city: Shanghai

first: Ruifeng

last: Ding

address: No. 415 Fengyang Road, Huangpu District

department: Department of Anesthesiology

institute: Changzheng Hospital, Second Affiliated Hospital of Naval Medical University

summary: Several studies have established a link between high-salt diet, inflammation, and hypertension. Vitamin D supplementation has shown anti-inflammatory effects in many diseases; gut microbiota is also associated with a wide variety of cardiovascular diseases, but potential role of vitamin D and gut microbiota in high-salt diet-induced hypertension remains unclear. Therefore, we used rats with hypertension induced by a high-salt diet as the research object and analyzed the transcriptome of their tissues (kidney and colon) and gut microbiome to conduct an overall analysis of the gut–kidney axis. We aimed to confirm the effects of high salt and calcitriol on the gut–kidney immune system and the composition of the intestinal flora. We demonstrate that consumption of a high-salt diet results in hypertension and inflammation in the colon and kidney and alteration of gut microbiota composition and function. High-salt diet-induced hypertension was found to be associated with seven microbial taxa and mainly associated with reduced production of the protective short-chain fatty acid butyrate. Calcitriol can reduce colon and kidney inflammation, and there are gene expression changes consistent with restored intestinal barrier function. The protective effect of calcitriol may be mediated indirectly by immunological properties. Additionally, the molecular pathways of the gut microbiota-mediated BP regulation may be related to circadian rhythm signals, which needs to be further investigated. An innovative association analysis of the microbiota may be a key strategy to understanding the association between gene patterns and host.

overall_design: mRNA profiles of kidney and colon from three groups (normal control (NC) group, high salt model (HS) group, and high salt diet plus calcitriol supplementation (CAL) group) were generated by RNAseq, with n=4~6, using Illumina NEXTSEQ 500 platform.

first: Zilong

last: Xiao

first: Yufeng

last: Jiang

first: Yi

last: Yang

first: Yang

last: Ji

first: Xunxia

last: Bao

first: Kaichen

last: Xing

first: Xinli

last: Zhou

first: Sibo

last: Zhu


title: ACC1-expressing pathogenic T helper 2 cell populations facilitate lung and skin inflammation

city: Kisarazu

first: Toshio

last: Kanno

state: Chiba

address: 2-6-7 Kazusa-kamatari

institute: KAZUSA DNA Research Institute

summary: T cells possess distinguishing effector functions and drive inflammatory disorders. We have previously identified IL-5-producing Th2 cells as the pathogenic population predominantly involved in the pathology of allergic inflammation. However, the cell-intrinsic signaling pathways that control the pathogenic Th2 cell function are still unclear. We herein report the high expression of acetyl-CoA carboxylase 1 (ACC1) in pathogenic CD4+ T cell population in the lung and skin. The genetic deletion of CD4+ T cell-intrinsic ACC1 dampened eosinophilic and basophilic inflammation in the lung and skin by constraining IL-5 or IL-3 production. Mechanistically, ACC1-dependent fatty acid biosynthesis induces the pathogenic cytokine production of CD4+ T cells via metabolic reprogramming and the availability of acetyl-CoA for epigenetic regulation. We thus identified a distinct phenotype of the pathogenic T cell population in the lung and skin, and ACC1 was shown to be an essential regulator controlling the pathogenic function of these populations to promote type 2 inflammation.

overall_design: Related to Fig 5, RNA-seq was performed to investigate inflammatory cytokine genes that are associated with Th cells.  Related to Fig S3, RNA-seq was performed to investigate inflammatory cytokine genes that are associated with Th2 and Th17 cells.

first: Takahiro

last: Nakajima

first: Satoru

last: Yokoyama

first: Shigemi

last: Sasamoto

first: Hikari

last: Asou

first: Damon

last: Tumes

first: Osamu

last: Ohara

first: Toshinori

last: Nakayama

first: Yusuke

last: Endo


title: Interleukin-10 causes interferon-γ-dependent emergency myelopoiesis

first: Ana

last: Maceiras

overall_design:
AmpliSeq Transcriptome was performed in different populations of bone marrow progenitors from pMT-10 overexpressing IL-10 and BL/6 (control).
Targeted RNA-seq


title: The histone lysine acetyltransferase HBO1 regulates hematopoietic stem cell quiescence and self-renewal

city: Parkville

first: Waruni

last: Abeysekera

address: 1G Royal Parade

department: Bioinformatics

institute: Walter and Eliza Hall Institute of Medical Research

summary: KAT7 (HBO1) is a histone acetyltransferase required for histone H3 lysine 14 acetylation (H3K14ac) and normal levels of gene expression in hematopoietic stem and progenitor cells (HSPCs).  The loss of H3K14ac was detected by western blot in whole bone marrow and by flow cytometry in specific HSPC populations. In order to determine the normal distribution of H3K14ac in the HSPC population a CUT&Tag experiment was performed using lineage negative, cKit positive Sca1 positive cells (LSK) and lineage negative, cKit positive Sca1 negative cells (progenitor cells). HBO1 was deleted using the poly(I:C)  inducible MX1-cre recombinase transgene and compared to Kat7 heterozygous control cells, in which the MX1-cre was likewise induced.  Analysis was performed at a stage when H3K14ac was no longer detectable by western blot.  In order to control for the effects of PCR amplification and normalisation of library size before sequencing, equal numbers of Drosophila Schneider 2 cells were added to each sorted mouse cell population (“spike in”). After sequencing-read quality control, reads were mapped to a combined Mus musculus (Mm10) and Drosophila melanogaster (DmelR635) genome. To each mouse sample a normalisation factor based on the recovery of Drosophila reads within the same sample was applied. Genomic distribution of KAT7 (HBO1) was determined in control progenitor cells by CUT&Tag.

overall_design: 2 mutant (conditional knock out) vs. 2 control haematopoietic stem were assessed for the genomic distribution of histone H3 lysine 14 acetylation (H3K14ac) by CUT&Tag. Control progenitor cells were assessed for the genomic distribution of KAT7.

first: Tim

last: Thomas

first: Anne

last: Voss

first: Yuqing


title: Epithelial phenotype restoring drugs suppress macular 1 degeneration phenotypes in an iPSC model

city: Bethesda

first: David

last: McGaughey

state: MD

address: 10 Center Dr

department: NEI

institute: NIH

summary: Age-related Macular Degeneration (AMD), a blinding eye disease, is characterized by pathological protein- and lipid-rich drusen deposits underneath the retinal pigment epithelium (RPE) and atrophy of the RPE monolayer in advanced disease stages - leading to photoreceptor cell death and vision loss. Currently, there are no drugs to stop drusen formation or RPE atrophy in AMD. We developed an iPSC-RPE AMD model that recapitulates drusen and RPE atrophy. Drusen deposition is dependent on AMD-risk-allele CFH(H/H) and anaphylatoxin triggered alternate complement signaling via the activation of NF-B and downregulation of autophagy pathways. High-throughput screen identified two drugs, L-745,870, a dopamine receptor antagonist, and aminocaproic acid, a protease inhibitor that reduce drusen deposits and restore RPE epithelial phenotype in anaphylatoxin challenged iPSC-RPE with or without the CFH(H/H) genotype. This comprehensive iPSC-RPE model replicates key AMD phenotypes, provides molecular insight into the role of CFH(H/H) risk-allele in AMD, and discovers two candidate drugs to treat AMD.

overall_design: Examination of whole RNA seq across three different iPSC-RPE with CC-HS treatment and in presence of drugs.

first: Ruchi

last: Sharma

first: Aman

last: George

first: Malika

last: Nimmagada

first: Davide

last: Ortolan

first: Kapil

last: Bharti


title: Bulk ATAC-seq on whole brain across DGRP lines (homozigous fly lines)

city: Leuven

first: Gert

last: Hulselmans

address: Herestraat 49 PO Box 602

department: Center for Brain and Disease Research

country: Belgium

institute: VIB

summary: Bulk ATAC-seq performed on whole adult brains across multiple homozigous fly lines (DGRP) in order to find caQTLs. Young adults (1-3 days) were used for all genotypes.

overall_design: omniATAC-seq on one male and one female fly per experiment

first: S

last: Aibar

first: J

last: Janssens

first: I

last: Taskiran

last: Aerts


title: For lncRNA, circRNA and mRNA sequencing of thyroid-associated ophthalmopathy patients and normal people

first: Zifan

last: Yue

address: No. 415 Fengyang Road, Hangpu District

department: Ophthalmology

institute: Shanghai Changzheng Hospital

type:
Expression profiling by array
Non-coding RNA profiling by array

summary:
We identified the differentially expressed mRNAs and lncRNAs/circRNAs of the case group and the control group through high-throughput sequencing technology, performed a functional enrichment analysis of the differentially expressed genes, and used protein-protein interaction (PPI) analysis to identify the hub genes.
A novel ceRNA network was successfully established, and the network components may serve as promising diagnostic biomarkers or therapeutic targets for thyroid-associated ophthalmopathy in the future.

overall_design: Orbital adipose/connective tissue was obtained from 3 thyroid-associated ophthalmopathy patients with no history of thyroid disease, inflammatory disease, autoimmune disease, or orbital disease. All the thyroid-associated ophthalmopathy patients underwent orbital decompression for proptosis correction, and control subjects underwent cosmetic upper and lower blepharoplasty. The 3 control groups were obtained from the tissue remains of healthy people after plastic surgery.

first: Fei

last: Tong


title: Transcriptome analysis of LPS stimulated PECs (WT and Zc3h12a S513A)

city: Kyoto

first: Kotaro

last: Akaki

address: Sakyo-ku, Yoshidakonoecho

department: Department of Medical Chemistry

institute: Kyoto University

summary: Regnase-1 is an endoribonuclease crucial for controlling inflammation by degrading mRNAs encoding cytokines and inflammatory mediators in mammals. However, it is unclear how Regnase-1-mediated mRNA decay is controlled in interleukin (IL)-1β or Toll-like receptor (TLR) ligand-stimulated cells. Here, by analyzing the Regnase-1 interactome, we found that IL-1β or TLR stimulus dynamically induced the formation of Regnase-1-β-transducin repeat-containing protein (βTRCP) complex. Importantly, we also uncovered a novel interaction between Regnase-1 and 14-3-3 in both mouse and human cells. Strikingly, both interactions occur in a mutually exclusive manner, underscoring the importance of modulating Regnase-1’s activity. Additionally, we show that in IL-1R/TLR-stimulated cells, the Regnase-1-14-3-3 interaction is mediated by IRAK1 through a previously uncharacterized C-terminal structural domain. Phosphorylation of Regnase-1 at S494 and S513 is critical for Regnase-1-14-3-3 interaction, while a different set of phosphorylation sites of Regnase-1 are known to be required for the recognition by βTRCP and proteasome-mediated degradation. 14-3-3 stabilizes Regnase-1 but abolishes its activity by inhibiting Regnase-1-mRNA association. Furthermore, nuclear-cytoplasmic shuttling of Regnase-1 is abrogated by 14-3-3 interaction. Taken together, the results suggest that a novel inflammation-induced interaction of 14-3-3 with Regnase-1 stabilizes inflammatory mRNAs by sequestering Regnase-1 in the cytoplasm to prevent mRNA recognition.

overall_design: Transcriptome analysis of thioglycollate-elicited PECs derived from WT and Regnase-1 (Zc3h12a) S513A knock-in mice. Cells were stimulated with LPS for 0, 4, or 8 hours.

first: Fabian

last: Hia

first: Nobuyuki

last: Kakiuchi

first: Seishi

last: Ogawa

first: Osamu

last: Takeuchi


title: Myeloid Niches Sustain Antitumor T Cells and License PD-1 Blockade via CD28 Costimulation

city: Lausanne

first: Aspram

last: Minasyan

address: Bugnon 25A

institute: Ludwig Institute for Cancer Research / CHUV

summary: The local mechanisms regulating exhaustion of tumor-infiltrating lymphocytes (TILs) and responsiveness to PD-1 blockade remain partly elucidated. In human ovarian cancer we show that tumor-reactive intraepithelial (ie)CD8+ TILs engaged by antigen are polyfunctional and upregulate PD-1, which restrains their effectiveness. PD-1+ TILs exhibit a continuum of TCR-engaged/exhausted states with variable effector fitness related to CD28 costimulation, which they receive in intraepithelial niches involving myeloid antigen-presenting cells (mAPC). Following PD-1 blockade, activation of TILs requires CD28 costimulation mediated in situ by tumor mAPCs, which is locally enhanced by CTLA-4 blockade. CD40 ligand also amplifies TIL responses in situ, especially in tumors in which mAPCs are not activated. Thus, dysfunctional and exhausted TILs, in a state of TCR engagement but without proper CD28 costimulation by mAPCs in situ, are unlikely to fully benefit from PD-1 blockade.

overall_design: scRNA-seq of CD8+ TILs from 17 ovarian tumor-digest cultures.

first: Riccardo

last: Turrini

first: Noémie

last: Fahr

first: Denarda

last: Dangaj

first: Marie-Agnès

last: Doucey

first: George

last: Coukos


title: Supermeres, a new functional extracellular nanoparticle replete with disease biomarkers and therapeutic targets

city: NASHVILLE

first: Marisol

last: Ramirez

state: TENNESSEE

address: 2220 Pierce Avenue, 571 Preston Research Building

department: Center for Quantitative Sciences, Department of Biostatisticss

institute: Vanderbilt University Medical Center

summary: RNA-Seq of cellular and extracellular samples from DiFi cells. The distribution of extracellular RNA among cells, sEV pellet, exomeres and supermeres  is distinct.

overall_design: The extracellular release of RNA from colonrectal cancer cell line DiFi is examined based on separation of the samples into distinct extracellular compartments.

first: Qin

last: Zhang

first: Dennis

last: Jeppesen

first: James

last: Higginbotham

first: Qi

last: Liu

first: Robert

last: Coffey


title: Transcripitonal analysis of genes in the mouse liver that regulated by extracellular nanoparticles by Next-Generation Sequencing

summary: We examined the effects on the liver following systemic delivery of extracellular nanoparticles exomeres and supermeres. Mice were injected with supermeres or exomeres via tail vein.To discern possible differences in the effects of supermeres and exomeres, we performed RNA-seq analysis of the liver tissue.GSEA analysis revealed that there were pathways significantly downregulated by both nanoparticles including cholesterol homeostasis, fatty acid metabolism, oxidative phosphorylation and adipogenesis. However, despite these overall similarities, there were significant differences in gene expression between the two groups.

overall_design: mRNA profiles of mouse liver tissue treated with PBS control, exomeres and supermeres

first: Youngmin

last: Lee


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

overall_design: Refer to individual Series