title: Transcriptomics of Glp1r-expressing cells in mouse hypothalamus tissue

city: Cambridge

first: Fiona

last: Gribble

state: Cambridgeshire

address: Box 289 Addenbrooke's Hospital, Hills Road

department: Medical Research Laboratories

institute: University of Cambridge

summary: We purified Glp1r-expressing cells from mouse hypothalamus by FACS and identified cell types distinguished by transcripts.

overall_design: Comparative transcriptomic study of cell populations by single cell RNAseq, pooled from 4x male biological replicates.

first: Alice

last: Adriaenssens

first: Christopher

last: Smith


title: Transcriptomic responses of common wheat segregating for Wsm2 locus to wheat streak mosaic virus (WSMV) and mock infection

status: Public on Apr 27 2022

city: Fort Collins

first: Yucong

last: Xie

state: CO

address: 307 Campus delivery

department: Department of Soil and Crop Sciences

country: USA

institute: Colorado State University

type: Expression profiling by high throughput sequencing

summary: To understand transcriptomic responses of WSMV resistant (contains Wsm2 locus or Wsm2+) and susceptible wheat (absent Wsm2 locus or Wsm2-) to WSMV and mock treated conditions and identify possible causative genes underlying Wsm2 locus, we performed a RNA-seq study for differentially expressed genes (DEGs) and de novo transcriptome assembly analysis.

overall_design: Illumina HiSeq 2000 were performed on RNA-seq libraries from four biological replicates of two genotypes (selected double haploids from Snowmass and Antero population contains or absent for Wsm2 locus, annotated as Wsm2+ and Wsm2-) under mock treated condition (Buffer) and WSMV treated (WSMV) condition. Raw reads were filtered to remove adaptor and low-quality reads, and then mapped to wheat reference genome IWGSC ReSeq v1.0. Expression was tabulated with Featurecounts and pairwise comparisons were performed on raw reads with DESeq2. De novo assembly of all cleaned reads were performed with Trinity software.

first: Stephen

last: Pearce


title: Gene expression profiling of osterix-derived cells from control and PTH1R knockout mice in osteoprogenitors

city: Minato-ku

first: Takaharu

last: Kimura

state: Tokyo

address: 4-6-1 Shirokanedai

country: Japan

institute: The University of Tokyo

summary: We used Osx-Cre:GFP / ROSA26-loxP-stop-loxP-tdTomato mice with or without PTH1Rfl/fl. Bone marrow and bone were digested to make single cell suspension, and sorted by fluorescence. Bulk cell RNA sequencing were performed to identify specific transcripts from osterix-derived cells.

overall_design: Freshly isolated 3 subsets of osterix-derived cells from control and conditional PTH1R KO.

first: Joy

middle: Y

last: Wu


title: The single-cell transcriptome profiling of mouse bone marrow LSK cells after resolution of periodontitis

city: Philadelphia

first: George

last: Hajishengallis

state: PA

address: School of Dental Medicine

institute: University of Pennsylvania

summary: Periodontitis induces myeloid lineage skewing of the bone marrow progenitor cells. In this study, we performed single cell RNA-sequencing in LSK (Lin—Sca-1+cKit+ ) cells to determine whether the periodontitis-induced myeloid lineage bias persists after the resolution of periodontal inflammation.

overall_design: C57BL/6 mice were trained by subjecting them to LIP for 21 days followed by ligature removal (to enable inflammation resolution) for 14 days (21dL/14dR, L) or were not ligated during the entire period (NL, N). Hematopoietic progenitors (LSKs; Lin-cKit+Sca1+)  in mice bone marrow were sorted by FACS for single-cell transcriptomics analysis.

first: Xiaofei

last: Li

first: Xiang

last: Yu


title: Reduced adhesion of aged intestinal stem cells contributes to an accelerated clonal drift

city: Ulm

first: Medhanie

middle: Assmelash

last: Mulaw

address: Albert-Einstein-Allee 11

country: Germany

institute: University of Ulm

summary:
To investigate the differential genes expression between young and aged Lgr5 high-ISCs
We performed single-cell gene expression profiling analysis using data obtained from 3 young and 4 old mice

overall_design: Comparative gene expression profiling analysis of sc-RNA-seq data for young and aged Lgr5 high-ISCs from young and aged mice

first: M

middle: A


title: Single cell RNA sequencing of Gipr- and Glp1r-expressing cells in the mouse hypothalamus

country: United Kingdom

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

overall_design: Refer to individual Series


title: Transcriptomics of Gipr-expressing cells in mouse hypothalamus tissue [dataset 2]

summary: We purified Gipr-expressing cells from mouse hypothalamus by FACS and identified cell types distinguished by transcripts.

middle: M


title: Transcriptomics of Gipr-expressing cells in mouse hypothalamus tissue [dataset 1]

overall_design: Comparative transcriptomic study of cell populations by single cell RNAseq, pooled from 24x female biological replicates.


title: The effect of periodontitis - induced trained immunity on the epigenomic profile of mouse bone marrow LSK and GMP cells

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Peripheral inflammation affects hematopoietic stem and progenitor cells (HSPCs) in the bone marrow and induce myeloid lineage skewing of the progenitor cells. In this study, we performed single cell ATAC-sequencing in LSK (Lin—Sca-1+cKit+ ) and GMP (Lin—c-Kit+Sca1—CD16/32+CD34+) cells to determine the impact of ligature-induced periodontitis (LIP) on the epigenomic profile of these BM cells.

overall_design: C57BL/6 mice were trained by subjecting them to LIP for 21 days followed by ligature removal (to enable inflammation resolution) for 14 days (21dL/14dR, L) or were not ligated during the entire period (NL, N). Hematopoietic progenitors (LSKs; Lin-cKit+Sca1+) and granulocyte-macrophage progenitors (GMP; Lin-cKit+Sca1-CD16/32+CD34+) in mice bone marrow were sorted by FACS for single cell ATAC-seq analysis of epigenomic profile.


title: Placental abnormalities are associated with type of embryo transfer in assisted reproductive technologies (ART)

status: Public on Apr 26 2022

first: Yemin

last: Lan

address: 3400 Civic Center Blvd

summary: ART  is associated with abnormalities in DNA methylation and a higher incidence of adverse pregnancy outcomes in humans and mice. However, which specific exposure, among the interventions utilized during ART, is responsible for these outcomes remains unknown. Embryo transfer after vitrification-thaw (frozen ET) into a maternal hormonal environment more closely resembling unassisted conception is increasingly utilized clinically, but reports suggest a higher incidence of large for gestational age (LGA) infants, with male infants potentially more affected than female infants.

overall_design: A prospective cohort study including patients from 2010-2020 was performed and human placentas from singleton pregnancies were obtained after:  fresh IVF cycles and programmed frozen embryo transfer cycles

first: Jayashri

last: Ghosh

first: Carmen

last: Sapienza

first: Monica

last: Mainigi

first: Suneeta

last: Senapati

first: Sneha

last: Mani


title: The effect of periodontitis - induced trained immunity on the transcriptomic profile of mouse LSK cells [RNA-seq]

summary: Hematopoietic stem and progenitor cells (HSPCs) in the bone marrow can sense peripheral inflammation and adapt through expansion and skewing toward the myeloid lineage. In this study, we performed RNA sequencing in LSK (Lin—Sca-1+cKit+ ) cells to determine the impact of ligature-induced periodontitis on the transcriptomic profile of HSPCs in mice bone marrow.

overall_design: C57BL/6 mice were subjected, or not (Non-ligature, NL), to ligature-induced periodontitis (LIP) for 7 days. Mice bone marrow were harvsted and LSKs were FACS-sorted and subjected to RNA-sequencing and transcriptome analysis.


title: The effect of periodontitis - induced trained immunity


title: The Transcript Integrity Index (TII) provides a standard measure of sperm RNA

city: Detroit

first: Grace

middle: Michelle

last: Swanson

state: MI

address: 275 E. Hancock, C.S. Mott Center room 271

institute: Wayne State University

summary: Human sperm RNA-seq was performed and sequences used for the generation of a RNA integrity index capable of assessing samples with high RNA fragmentation.  A two-prong approach was used including both manual and computational analysis.  This identified a series of sperm RNAs that are consistently intact, and acceptable for use in determining sample RNA quality.  These RNAs function in roles related to spermatogenesis and/or fertilization.

overall_design: Samples were human sperm from the male partner of couples seeking fertility treatment.

first: Molly

middle: S

last: Estill

last: Krawetz


title: Expression Data from DMSO or honokiol treated THP-1 and U-937 cells

city: Weifang

first: Yanli

last: Sun

state: Shandong

address: No. 7166, Baotong West Street

department: Department of Clinical Laboratories

country: China

institute: Stem cell institution of Weifang Medical University

summary: We used microarrays to evaluate the effect of honokiol on gene expression in THP-1 and U-937 cells.

overall_design: THP-1 or U-937 cells were treated with honokiol (20 µM or 35  µM) or the negative control (0.1% DMSO) for 24 hours for Total RNA isolation and hybridization on Microarray.

first: Zhenbo

last: Hu


title: RNA-sequencing of RNA extracted from WT and P2Y₂ KO cardiac adipose-derived stem cells (cADSC), untreated or treated with UTP (100 μm)

city: Brussels

first: Didier

last: Communi

address: Route de Lennik 808

department: IRIBHM

country: Belgium

institute: Université Libre de Bruxelles

summary: Our goal was to identify UTP-regulated genes in WT cADSC and P2Y₂ KO cADSC in the frame of the potential therapeutic transplantation of these cells into the mouse ischemic heart after a myocardial infarction. We identified UTP-regulated genes involved in inflammation and the immune response, UTP target genes linked to extracellular matrix organization such as matrix metallopeptidases and various collagen types, UTP-regulated genes related to macrophage chemotaxis and differentiation into pro-fibrotic foam cells and UTP target genes linked to angiogenesis regulation and the response to hypoxia.

overall_design: 8 samples were analyzed; 2 different pools of RNA from 2 untreated WT cADSC cultures, 2 pools of RNA from 2 UTP-treated (100 μm) WT cADSC cultures, 2 pools of RNA from 2 untreated P2Y₂ KO cADSC cultures and 2 pools of RNA from 2 UTP-treated (100 μm) P2Y₂ KO cADSC cultures.

first: Esteban

last: Diaz Villamil

first: Frédérick

last: Libert


title: Single-Cell versus Single-Nucleus: Transcriptome differences in murine kidney after ischemia-reperfusion injury

city: Hannover

first: Svenja

last: Gaedcke

address: Carl-Neuberg-Str. 1

institute: Hannover Medical School

summary: Goal of the study was to compare single cell (sc) and single nucleus (sn) sequencing of murine kidney after mild (17min) and severe (27min) ischemia reperfusion injury

overall_design: sc and snSeq of murine kidney

first: Julius

last: Sinning

first: Oliver

last: Dittrich-Breiholz

first: Hermann

last: Haller

first: Inga

last: Soerensen-Zender

first: Chieh

last: Liao

first: Alexandra

last: Helmke

first: Payel

last: Sen

first: Sibylle

last: von Vietinghoff

first: David

last: DeLuca

first: Roland

last: Schmitt


title: Recapitulation of endogenous 4R tau expression and formation of insoluble tau in directly reprogrammed human neurons

city: Saint Louis

first: Lucia

last: Capano

state: MO

address: 660 South Euclid Avenue

department: Developmental Biology

institute: Washington University in Saint Louis

summary: Tau is a microtubule-binding protein expressed in neurons and the equal ratio between 4-repeat (4R) and 3-repeat (3R) isoforms are maintained in normal adult brain function. Dysregulation of 3R:4R ratio causes tauopathy and human neurons that recapitulate tau isoforms in health and disease will provide a platform for elucidating pathogenic processes involving tau pathology. We carried out extensive characterizations of tau isoforms expressed in human neurons derived by microRNA-induced neuronal reprogramming of adult fibroblasts. Transcript and protein analyses showed miR-neurons expressed all six isoforms with the 3R:4R isoform ratio equivalent to that detected in human adult brains. Also, miR-neurons derived from familial tauopathypatients with a 3R:4R ratio altering mutation showed increased 4R tau and the formation of insoluble tau with seeding activity. Our results collectively demonstrate the utility of miRNA-induced neuronal reprogramming to recapitulate endogenous tau regulation comparable to the adult brain in health and disease.

overall_design: De-identified human adult dermal fibroblasts from healthy and IVS10+16 patients were transduced with lentivirus to express the brain enriched microRNAs, miR-9/9* and miR-124, cortical-enriched transcription factor MYT1L, and the small molecules ISX9 (in induce NeuroD family of transcription factors) and DAPT (to enhance neuronal reprogramming) to directly convert fibroblasts into miNs as previously established (Yoo et al., Nature 2011; Richner et al., Nat. Prot. 2015; Abernathy, Kim, and McCoy et al., 2017). En masse, cells were collected for both bulk RNA-seq and single cell RNA-seq and prepared for analysis as described in the Protocols.

first: Chihiro

last: Sato

first: Elena

last: Ficulle

first: Anan

last: Yoo

first: Kanta

last: Horie

first: Ji-Sun

last: Kwon

first: Kyle

middle: F

last: Burbach

first: Nicolas

middle: R

last: Barthelemy

first: Susan

middle: G

last: Fox

first: Celeste

last: Karch

first: Randall

middle: J

last: Bateman

first: Henry

last: Houlden

first: Richard

middle: I

last: Morimoto

last: Holtzman

first: Karen

middle: E

last: Duff

first: Andrew


title: Oncogenesis and organ infiltration specific therapy in a preclinical model of Ph-like B cell acute lymphoblastic leukemia [RNA-seq dataset 2]

city: Mainz

first: Federico

last: Marini

address: Obere Zahlbacher Strasse 69

department: Institute of Medical Biostatistics, Epidemiology and Informatics (IMBEI)

institute: University Medical Center - Mainz

summary: We investigated, whether IRF4 deletion might have other collateral oncogenic effects, e.g. by causing signaling pathway alterations and transcriptional changes involved in leukemogenesis. In a first step, we therefore compared gene expression of tumours 10, 11 and 14 described above to those of B220+ mIgM- pro/preB cells that had been sorted from the BM of Irf4–/– mice. The genes that differed more than two-fold between samples were then, in a second step, reduced to only those that we found to be altered by de novo IRF4 expression (Fig. 3E). This approach allowed us to focus on genes that are regulated by IRF4 and at the same time altered in leukemic as compared to non-tranformed pro/preB cells. Finally, as a further level of selection, the resulting genes were matched with a data bank provided by Ochiai et al.30 and selected for genes bound by IRF4 in a ChIP analysis of B cells.

overall_design: Two control (gfp empty) and two test (irf4) samples, in both cases representing biological replicates, are included in this study

first: Dennis

last: Das Gupta

first: Mattthias

last: Klein


title: Inhibition of pyrimidine biosynthesis targets protein translation in AML [RNA-seq_RUNX1]

city: Melbourne

first: Joan

last: So

state: Victoria

address: 305 Grattan St

country: Australia

institute: Peter MacCallum Cancer Centre

summary: Investigation of the transcriptional profile of AML in response to DHODH inhibition by AG636. RUNX1-RUNX1T1 cells were transplanted into Ptprca recipients. Mice bearing tumors were treated with AG636 for one day. Leukemic stem cells (cKit high; CD11b low) from bone marrow and spleen were isolated and RNA sequencing performed.

overall_design: Gene expression profiles of Vehicle or AG636 AML cells.

first: Emily

last: Gruber

first: Alexander

middle: C

last: Lewis


title: Inhibition of pyrimidine biosynthesis targets protein translation in AML [Targeted_CRISPR_screen]

summary: Investigating genes that increase or decrease treatment efficacy of AG636 in MLL-AF9; KrasG12D (MN) AML cells.

overall_design: MN cells were transduced with a custom CRISPR library, and treated with 100 nM or 250 nM AG636 for 10 days and 24 days. Enrichment or dropout analysis was performed.


title: Inhibition of pyrimidine biosynthesis targets protein translation in AML [RNA-seq_I1DN]

summary: Investigation of the transcriptional profile of AML in response to DHODH inhibition by AG636. I1DN model was generated by co-transduction of foetal liver derived HSPCs with constructs encoding IDH1R132H, DNMT3AR882H and NrasG12D. Cells were transplanted into Ptprca recipients. Mice bearing I1DN tumors were treated with AG636 for one day. Leukemic stem cells (cKit high; CD11b low) from bone marrow and spleen were isolated and RNA sequencing performed.


title: Detecting cell-of-origin and cancer-specific methylation features of cell-free DNA from Nanopore sequencing

city: Ein Kerem

first: Benjamin

middle: P

last: Berman

address: Hadassah University Medical Center, Bldg 3 rm 30

country: Israel

institute: Hebrew University of Jerusalem

type: Methylation profiling by high throughput sequencing

summary: The Oxford Nanopore (ONT) platform provides portable and rapid genome sequencing, and its ability to natively profile DNA methylation without complex sample processing is attractive for clinical sequencing. We recently demonstrated ONT shallow whole-genome sequencing to detect copy number alterations (CNA) from the circulating tumor DNA (ctDNA) of cancer patients. Here, we show that cell-type and cancer-specific methylation changes can also be detected, as well as cancer-associated fragmentation signatures. This feasibility study suggests that ONT shallow WGS could be a powerful tool for liquid biopsy, especially real-time medical applications.

overall_design: We reanalyzed 10 plasma cell-free DNA samples from an earlier published study (available at EGAD00001006888). Original data was raw Oxford Nanopore fast5s files, which we mapped and called 5mC levels.

first: Efrat

last: Katsman

first: Shari

last: Orlanski

first: Filippo

last: Martignano

first: Silvestro

last: Conticello


title: Inhibition of pyrimidine biosynthesis targets protein translation in AML [RNA-seq_MN-2]

summary: Investigation of the transcriptional profile and compared with the genome accessibility (using ATAC-seq) in MN cells. Mice bearing MN tumors were treated with AG636 for 2 days. Leukemic stem cells (cKit high; CD11b low) from bone marrow and spleen were isolated. RNA sequencing and ATAC-seq were performed.

overall_design: Gene expression profiles of Vehicle- or AG636-treated MN AML cells.


title: RBM17 Mediates Evasion of Pro-Leukemic Factors from Splicing-coupled NMD to Enforce Leukemic Stem Cell Maintenance

city: La Jolla

first: Gene

last: Yeo

state: CA

address: 2880 Torrey Pines Scenic Dr. Room 3805/Yeo Lab

institute: UCSD

summary: Acute myeloid leukemia (AML) patients suffer from chemo-resistance, high relapse frequency, and low overall survival rate, outcomes driven by leukemic stem cells (LSCs). Understanding the molecular mechanisms that support these primitive leukemic cells is crucial for developing effective AML therapeutics. In the present study, we demonstrate that upregulation of the splicing factor RBM17 preferentially marks and sustains the primitive compartment of AML. RBM17 expression is significantly higher in LSCs and its levels in human AML directly correlate with shortened survival in patients with the disease. RBM17 knockdown in primitive primary AML cells leads to myeloid differentiation and the impairment of their in vitro colony forming and in vivo engraftment capacities. To study the molecular mechanisms underlying the functional roles of RBM17 in AML, we performed global profiling of the RBM17-RNA interactome and proteome changes downstream of RBM17 knockdown. Through these integrative multi-omics analyses, we show that RBM17 repression leads to inclusion of poison exons and production of nonsense-mediated decay (NMD)-sensitive transcripts for pro-leukemic factors such as RBM39 and EZH2, along with the translation initiation factor EIF4A2. We further show that EIF4A2 expression is enriched in LSCs and inhibition of EIF4A2 impairs primary AML progenitor activity. Proteome analysis of AML cells after EIF4A2 knockdown demonstrate that EIF4A2 repression largely recapitulates the biological effect of RBM17 knockdown including the pronounced suppression of proteins involved in ribosome biogenesis. Overall, these results provide a rationale to target RBM17 and/or its downstream NMD-sensitive splicing substrates in primitive leukemic cells for AML treatment.

overall_design: eCLIP-seq against RBM17 in K562 cells.

first: Lina

last: Liu

first: Shashank

last: Sathe

middle: W

first: Yu

last: Lu


title: Inhibition of pyrimidine biosynthesis targets protein translation in AML [ATAC-seq_MN-2]

summary: Investigation of the chromatin accessibility in MN cells. Mice bearing MN tumors were treated with AG636 for 2 days. Leukemic stem cells (cKit high; CD11b low) from bone marrow and spleen were isolated. RNA sequencing and ATAC-seq were performed.

overall_design: Genome binding/occupancy profiling of Vehicle- or AG636-treated MN AML cells by high-throughput sequencing.


title: Inhibition of pyrimidine biosynthesis targets protein translation in AML [RNA-seq_MN]

summary: Investigation of the transcriptional profile of MLL-rearranged AML in response to DHODH inhibition by AG636. Chemo-refractory syngeneic murine AML model driven by doxycycline-inducible expression of MLL-AF9 and constitutive expression of oncogenic Nras (MN) was transplanted into Ptprca recipients. Mice bearing MN tumors were treated with doxycycline or AG636 for one or four days. Leukemic stem cells (cKit high; CD11b low) from bone marrow and spleen were isolated and RNA sequencing performed.

overall_design: Gene expression profiles of Vehicle-, AG636- or Doxycycline-treated MN AML cells.


title: Oncogenesis and organ infiltration specific therapy in a preclinical model of Ph-like B cell acute lymphoblastic leukemia [RNA-seq dataset 1]

summary: We investigated whether IRF4 deletion might have other collateral oncogenic effects, e.g. by causing signaling pathway alterations and transcriptional changes involved in leukemogenesis. In a first step, we therefore compared gene expression of tumours 10, 11 and 14 described above to those of B220+ mIgM- pro/preB cells that had been sorted from the BM of Irf4–/– mice. The genes that differed more than two-fold between samples were then, in a second step, reduced to only those that we found to be altered by de novo IRF4 expression (Fig. 3E). This approach allowed us to focus on genes that are regulated by IRF4 and at the same time altered in leukemic as compared to non-transformed pro/preB cells. Finally, as a further level of selection, the resulting genes were matched with a data bank provided by Ochiai et al. and selected for genes bound by IRF4 in a ChIP analysis of B cells.

overall_design: Examination of primary Irf4–/– leukemia samples in comparison to Irf4–/– bone marrow derived pro/preB cells, as well as leukemia cell lines retrovirally transduced with empty vector control or IRF4 coding retrovirus.

first: Maria

last: Bieringer

first: Lisa

last: Menke


title: H3K27Ac mapping in DLBCL cell lines

city: New York

first: Laura

last: Pasqualucci

state: NY

address: 1130 St Nicholas Avenue

department: Institute for Cancer Genetics

institute: Columbia University

summary: Diffuse large B-cell lymphoma (DLBCL) is the most common B-cell non-Hodgkin lymphoma and remains incurable in ~40% of patients. Coding-genome sequencing efforts identified several genes/pathways altered in this disease, including new potential therapeutic targets. However, the non-coding genome of DLBCL remains unexplored. Here we show that active superenhancers (SEs) are highly and specifically hypermutated in 99% of DLBCL samples. Such aberrant somatic hypermutation (ASHM) displays signatures of Activation Induced Deaminase activity and is linked to genes encoding B cell developmental regulators and oncogenes. To elucidate the functional consequences of SE-ASHM, we explored two recurrent mutational hotspots identified in the SEs linked to the BCL6 and BCL2 proto-oncogenes, but unmutated in normal GC B cells. These mutations prevent the binding and transcriptional downregulation by the transcriptional repressors BLIMP1 and NR3C1, respectively. Genetic correction of selected mutations restored repressor DNA binding as well as target gene transcriptional repression. Notably, this led to counter-selection of cells harboring corrected alleles, indicating oncogenic dependency from the SE mutations. This pervasive SE mutational mechanism reveals a new major set of genetic lesions deregulating gene expression, which expands the involvement of known oncogenes in DLBCL pathogenesis and identifies new deregulated gene targets of therapeutic relevance.

overall_design: CHIP-Seq analysis of H3K27 was performed on 29 DLBCL cell lines using two independent antibodies

first: Antony

middle: B

last: Holmes


title: Transcriptomic analysis of Medicago truncatula calli with MtWOX9-1 overexpression

status: Public on Apr 25 2022

city: St Petersburg

first: Varvara

last: Tvorogova

address: 7/9 Universitetskaya emb

department: Genetics and Biotechnology

country: Russia

institute: St Petersburg State University

summary: Somatic embryogenesis (SE) is the development of embryo-like structures from somatic plant tissues. Recently, weve shown that transcription factor MtWOX9-1 belonging to WOX family is able to stimulate SE in the callus culture in Medicago truncatula. In this research, transcriptomic analysis of highly embryogenic calli with MtWOX9-1 overexpression was performed in comparison with wildtype calli. It was shown that MtWOX9-1 overexpression leads to the activation of several groups of genes, including genes related with cell division and tissue differentiation, and also with seed development. Enriched GO pathways included several groups related with histone methyltransferase activity as well as DNA methylation and chromatin binding, suggesting major epigenetic changes occuring in MtWOX9-1 overexpressing calli.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for calli obtained from Medicago truncatula R108 line plants with MtWOX9-1 (MTR_2g015000) overexpression (2 biological replicates) and calli obtained from Medicago truncatula R108 wildtype plants (2 biological replicates). Wildtype calli were used as control.

first: Elizaveta

last: Krasnoperova

first: Andrei

last: Kudriashov

first: Ksenia

last: Kuznetsova

first: Elina

last: Potsenkovskaia

first: Yulia

last: Fedorova

first: Ludmila

last: Lutova


title: CHD7 and Runx1 interaction provides a braking mechanism for hematopoietic differentiation

first: Bertie

last: Gottgens

address: Jeffrey Cheah Biomedical Centre

summary: We investigated the transcriptional consequences of Chd7 and Runx1 loss in 416B cells, a myeloid progenitor cell line. We engineered the cell line to express Cas9 protein and targeted endogenous Chd7 and Runx1 loci with sgRNAs introduced with lentiviral constructs. We read out the transcriptomes of control and perturbed cells using RNA-Seq following an established SMART-Seq2 protocol (Picelli et al. 2014).

overall_design: 416B cells were transduced with sgRNAs targeting either Chd7 gene (3 different sgRNAs), Runx1 gene (one sgRNA) or GFP (control). Each condition was performed in quadruplicate. Transcriptional changes were assessed using differential expression between treated and the control cells

first: Jingmei

last: Hsu

first: Hsuan-Ting

last: Huang

first: Iwo

last: Kucinski

first: Rebecca

last: Hannah

first: Nicola

last: Wilson

first: Berthold

first: Nancy

last: Speck

first: Leonard

last: Zon


title: Epigenetic reversal of lineage plasticity enhances responsiveness to anti-GD2 therapy in neuroblastoma

city: West Lafayette

first: Nathaniel

middle: Wesley

last: Mabe

state: Indiana

address: 201 S. University St.

department: Medicinal Chemistry and Molecular Pharmacology

institute: Purdue University

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

overall_design: Refer to individual Series


title: Human pluripotent stem cell derived hematopoietic cells using expanded potential stem cell media

city: Cambridge

institute: University of Cambridge

summary: We recently developed a novel chemical media formulation, Expanded Potential Stem Cell Medium (EPSCM), that maintains mouse pluripotent stem cells (PSCs) in a unique cellular state that affords extraembryonic differentiation capacity. Here, we have used EPSCM to stably maintain human PSCs and used EPSCM-maintained iPSCs (EC-EPSCM) to study in vitro hematopoietic differentiation by embryoid body (EB) formation. We found that human PSCs maintained in EPSCM can spontaneously form embryoid bodies and efficiently undergo in vitro hematopoiesis using a simple differentiation protocol. EC-EPSCM PSCs generated primitive-wave and definitive-wave hematopoietic cells, which were chracterized by RNA-sequencing analysis.

overall_design: Characterization of four hematopoietic cell populations (two cell types from two different time points) generated by the embyoid body (EB) differentiation of EPSCM-maintained human iPSC line (EC-EPSCM), which represent primitive-wave hematopoietic cells and definitive-wave hematopoietic cells. Cell populations were purified by fluorescence-activated cell sorting (50-cells per sample) and analyzed by RNA-sequencing using the Smartseq2 protocol.

first: Adam

last: Wilkinson


title: A PDX1 cistrome and single-cell transcriptome resource of the developing pancreas

first: Xiaodun

last: Yang

address: 3451 Walnut St

institute: University of Pennsylvenia

summary: Pancreatic and duodenal homeobox 1 (PDX1) is crucial for pancreas organogenesis, yet the dynamic changes in PDX1 binding in human or mouse developing pancreas have not been examined. To address this knowledge gap, we performed PDX1 ChIP-seq and single-cell RNA-seq using fetal human pancreata. We integrated our datasets with published datasets and revealed the dynamics of PDX1 binding and potential cell-lineage-specific PDX1 bound genes in the pancreas from fetal to adult stages. We identified a core set of developmentally conserved PDX1 bound genes that reveal the broad multifaceted role of PDX1 in pancreas development. Despite the well-known, dramatic changes in PDX1 function and expression, we found that PDX1 bound genes are largely conserved from embryonic to adult stages. This points towards a dual role of PDX1 in regulating the expression of its targets at different ages, dependent on other functionally-congruent or directly-interacting partners. We also showed that PDX1 binding is largely conserved in mouse pancreas. Together, our study reveals PDX1 targets in the developing pancreas in vivo and provides an essential resource for future studies on pancreas development.

overall_design: Single islet cells were dissociated from human fetal pancreata, and loaded on the 10x Genomics (Pleasanton, CA) platform. Single-cell transcriptome libraries were sequenced on the HiSeq platform (Illumina, San Diego, CA).

first: Doris

last: Stoffers

first: Diana

last: Stanescu

first: Jeffrey

last: Raum

first: Junil

last: Kim


title: Inhibition of pyrimidine biosynthesis targets protein translation in AML

first: Joan

last: So


title: Single nucleus RNA-seq gene expression profiling for protein coding and lncRNA genes using livers of male and female mice, with GH infusion, and following TCPOBOP exposure

city: Boston

first: David

middle: J.

last: Waxman

state: MA

address: 5 Cummington Mall

department: Department of Biology and Bioinformatics Program

institute: Boston University

summary: Single nucleus RNA-seq gene expression profiling was carried out for protein coding and lncRNA genes using nuclei extracted from livers from adult male and female mice; from male mice infused with GH continuously for one week, mimicking the female plasma GH pattern; and from male and female mice exposed to TCPOBOP, a xenobiotic agonist ligand of the nuclear receptor CAR, which perturbs sex-biased gene expression in the liver.  Our analysis of these rich single nucleus, mouse liver transcriptomic datasets, comprised of 32,000 liver nuclei representing 9 major liver cell types, revealed: 1) the expression of sex-biased genes and their key GH-dependent transcriptional regulators is primarily restricted to hepatocytes and is largely not a feature of liver non-parenchymal cells; 2) many sex-biased transcripts show sex-dependent zonation within the liver lobule; 3) gene expression is substantially feminized in both periportal and pericentral hepatocytes when male mice are infused with GH continuously; 4) sequencing nuclei increases the sensitivity for detecting thousands of nuclear-enriched long-noncoding RNAs (lncRNAs) and enables determination of their liver cell type-specificity, sex-bias and hepatocyte zonation profiles; 5) the periportal to pericentral hepatocyte cell ratio is significantly higher in male than female liver; and 6) TCPOBOP exposure disrupts both sex-specific gene expression and hepatocyte zonation within the liver lobule. These findings highlight the complex interconnections between hepatic sexual dimorphism and zonation at the single-cell level and reveal how endogenous hormones and foreign chemical exposure can alter these interactions across the liver lobule with large effects on both protein-coding genes and lncRNAs.

overall_design:
Single nuclei suspensions were generated from frozen liver tissue from adult male and female ICR (CD-1) mice that were treated with 3 mg/kg TCPOBOP dissolved in corn oil, or corn oil vehicle, for 27 hr, or adult male mice that were treated with continuous growth hormone given as an infusion (20 ng rat GH per hr per gram body weight) for 7 days. Single nuclei suspensions were processed through the standard 10x Genomics library preparation protocol to generate single nucleus RNA-seq sequencing libraries, used to investigate the sex bias in gene expression, cell type specificity and liver zonation expression profiles of expressed protein coding genes and lncRNAs under five different biological conditions.

Frozen liver tissue (livers from n=4 mice per group, frozen tissue pooled per condition) was homogenized, washed and filtered to generate single nuclei suspensions and subsequently 10x Genomics sequencing libraries for the following six groups: male and female mice injected with vehicle (control); male and female mice given TCPOBOP at 3 mg/kg body weight and then euthanized 27 h later; and male mice treated with growth hormone (GH) given as a continuous infusion using a subcutaneous Alzet osmotic minipump for 7 days, and its untreated male liver control.

first: Christine

middle: N

last: Goldfarb

first: Kritika

last: Karri


title: Plasma cell-free RNA profiling distinguishes cancer from pre-malignant conditions

city: Oxford

first: Breeshey

middle: Jane

last: Roskams-Hieter

address: John Radcliffe Hospital/Headley Way

institute: MRC Weatherall Institute of Molecular Medicine

summary: Cell-free RNA (cfRNA) in plasma reflects gene expression profiles of both localized sites of cancer and at the systemic host response. Here we explore the diagnostic potential of cell-free transcriptomes by mRNA sequencing. We sequenced total cell-free plasma RNA from 90 plasma samples from two independent sample cohorts representing two cancer types, two pre-cancerous conditions and non-cancer donors. We identified distinct gene sets and built classification models that could distinguish cancer patients with specific cancer types from premalignant conditions and non-cancer individuals with high accuracy. Determination of multiple myeloma from its pre-cancerous monoclonal gammopathy of undetermined significance (MGUS) yielded an accuracy of 90% (17/19). Detection of primary liver cancer from its premalignant condition cirrhosis yielded an accuracy of 100% (12/12). This work lays the foundation for developing low cost assays by measuring mRNA transcript levels in plasma using a small panel of genes for detection that can distinguish the presence of cancer from pre-cancerous conditions and non-cancer individuals.

overall_design: We sequenced total cell-free plasma RNA from 90 plasma samples from two independent sample cohorts representing two cancer types, two pre-cancerous conditions and healthy donors: 10 multiple myeloma and 8 liver cancer; 13 pre-malignant conditions including 9 monoclonal gammopathy of undetermined significance and 4 Cirrhosis; and 20 aged and gender matched healthy donor in our pilot cohort.

first: Thuy

middle: T

last: Ngo


title: RNA-seq analysis evaluating effects of Cdk7 inhibitor THZ1 on responses of mouse hearts to transverse aortic artery constriction (TAC)

city: South San Francisco

first: Saptarsi

last: Haldar

address: 1120 Veterans Blvd

institute: Amgen

summary: RNA-seq analysis evaluating effects of Cdk7 inhibitor THZ1 on responses of mouse hearts to TAC

overall_design: Mouse heart mRNA profiles from four groups: 1. Sham+Vehicle; 2. TAC+Vehicle; 3. TAC+THZ1(10mg/kg b.i.d.); 4. Sham+THZ1 (10mg/kg b.i.d.);

first: Tinghu

last: Zhang

first: Qiming

last: Duan


title: RNA-seq analysis evaluating effects of Cdk7 inhibitor THZ1 on responses of neonatal rat ventricular myocytes (NRVM) to adrenergic receptor agonist phenylephrine (PE)

summary: RNA-seq analysis evaluating effects of Cdk7 inhibitor THZ1 on responses of NRVM to adrenergic receptor agonist PE

overall_design: NRVM mRNA profiles from four groups: 1. Vehicle; 2. PE (100 μM); 3. PE (100 μM)+THZ1(10 nM); 4. THZ1 (10 nM);

first: Daniel

last: Day


title: Epigenetic studies of Aspergillus fumigatus

city: Daejeon

first: Kwang-Soo

last: Shin

department: Life Science

country: South Korea

institute: Daejeon University

summary: Transcriptome analysis of deletion mutant which coding histone modification enzymes compared to wild type by NGS

overall_design: Examination of mutant and wild type transcriptome

first: Yong-Ho

last: Choi

first: Min-Woo

last: Lee


title: BAF complex cooperates with c-Myc to orchestrate early fate decisions of CD8 T cells (RNA-seq II)

status: Public on Apr 24 2022

city: Memphis

first: Douglas

last: Green

state: United States

address: 262 Danny Thomas Pl

department: Immunology

institute: St Jude Children's Research Hospital

summary: We found cBAF and c-Myc co-assort asymmetrically to the two daughter nuclei, such that daughter cells with low c-Myc and cBAF display a cell fate trajectory towards memory T cell differentiation. Moreover, cBAF complex and c-Myc physically interact with each other, and act in establishing the chromatin landscape in activated CD8 T cells.

overall_design: RNA-seq analysis of 8 samples:  Activated CD8 T cells (n=4), Activated CD8 T cells treated with Arid1a inhibitor (n=4).

first: Ao

last: Guo

first: Hongling

last: Huang

first: Mark

last: Chen

first: Hongbo

last: Chi


title: Histone H3.3 K27M chromatin functions implicate a network of neurodevelopmental factors including ASCL1 and NEUROD1 in DIPG

city: Sacramento

first: Paul

last: Knoepfler

address: 2425 Stockton Blvd, Room 633

department: Cell Biology and Human Anatomy

institute: University of California Davis

summary:
Background:
The histone variant H3.3 K27M mutation is a defining characteristic of diffuse intrinsic pontine glioma (DIPG)/diffuse midline glioma (DMG). This histone mutation is responsible for major alterations to histone H3 post-translational modification (PTMs) and subsequent aberrant gene expression. However, much less is known about the effect this mutation has on chromatin structure and function, including open versus closed chromatin regions as well as their transcriptomic consequences.
Results:
Recently, we developed isogenic CRISPR-edited DIPG cell lines that are wild- type for histone H3.3 that can be compared to their matched K27M lines. Here we show via ATAC-seq analysis that H3.3K27M glioma cells have unique accessible chromatin at regions corresponding to neurogenesis, NOTCH, and neuronal development pathways and associated genes that are overexpressed in H3.3K27M compared to our isogenic wild-type cell line. As to mechanisms, accessible enhancers and super-enhancers corresponding to increased gene expression in H3.3K27M cells were also mapped to genes involved in neurogenesis and NOTCH signaling, suggesting that these pathways are key to DIPG tumor maintenance. Motif analysis implicates specific transcription factors as central to the neuro-oncogenic K27M signaling pathway including, in particular, ASCL1 and NEUROD1.
Conclusions:
Altogether our findings indicate that H3.3K27M causes chromatin to take on a more accessible configuration at key regulatory regions for NOTCH and neurogenesis genes resulting in increased oncogenic gene expression, which is at least partially reversible upon editing K27M back to wild- type.

overall_design: Total of four different cell lines each in duplicate (total of 8 samples sequenced). Lines notated as SU-DIPG-XVII/XIII-WT are isogenic to SU-SUPG-XVII/XIII and were edited using CRISPR-Cas9 (see Chen et al., 2020). Paired-end 150 sequencing was used.

first: N

middle: A

last: Lewis

first: P


title: RNA polymerase II (RNAPII) ChIP-seq analysis evaluating effects of Cdk7 inhibitor THZ1 on responses of neonatal rat ventricular myocytes (NRVM) to adrenergic receptor agonist phenylephrine (PE)

summary: RNA polymerase II (RNAPII) ChIP-seq analysis evaluating effects of Cdk7 inhibitor THZ1 on responses of NRVM to adrenergic receptor agonist PE

overall_design: Profiles of NRVM RNAPII occupancies from three groups: 1. Vehicle; 2. PE (100 μM); 3. PE (100 μM)+THZ1(10 nM);


title: JAZF1-SUZ12 dysregulates PRC2 recruitment and gene expression during cell differentiation

city: London

first: Maria

last: Vila de Mucha

address: 72 Huntley Street

institute: UCL Cancer Institute

summary: Polycomb repressive complex 2 (PRC2) maintains repression of genes specific for other cell differentiation stages through methylation of histone H3 lysine 27 (H3K27me3) and is regulated by nascent RNA. In endometrial stromal sarcoma, the PRC2 subunit SUZ12 is often fused with the transcription factor JAZF1. How JAZF1-SUZ12 affects PRC2 function and cell differentiation are unknown. Here, we show that JAZF1-SUZ12 disrupts PRC2 recruitment, gene expression and cell differentiation. The loss of the SUZ12 N-terminus in the fusion protein disrupted interaction with JARID2, EPOP and PALI1 and prevented recruitment of PRC2 from RNA to chromatin. JAZF1-SUZ12 occupied PRC2 target genes in undifferentiated cells but relocated to JAZF1 target genes during cell differentiation, altering patterns of H3K27me3 and gene expression and disrupting cell differentiation. We then performed RNA-seq in primary human cells expreesing all proteins, showing that JAZF1-SUZ12 was promoting a specific set of genes upregulated in decidualisation, while blocking immune related genes.  These results reveal the defects in PRC2 function and cell differentiation caused by the JAZF1-SUZ12 fusion protein, which may underlie its role in oncogenesis.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Suz12 GT/GT cell lines stably expressing FLAG-tagged proteins SUZ12, SUZ12D93, JAZF1-SUZ12 and JAZF1 were differentiated into Embryoid Bodies by removal of LIF and forced aggregation. After differentiation, at days 0,4 and 8 we generated calibrated ChIP-seq data using the FLAG epitope to see the binding profile of all 4 proteins. Additionally, we performed calibrated ChIP-seq at days 0 and 8 of H3K27me3 and H4ac. For assays in human endometrial stromal cells (hEnSC), cells were transduced with the same FLAG-tagged proteins, and differentiated for 8 days with medroxyprogesterone and 8?-Br-cAMP to generate decidual cells. Total RNA from days 0 and 8 of treatment was used for bulk RNA-seq to establish changes in gene expression

first: Manuel

last: Tavares

first: Garima

last: Khandelwal

first: Joanne

last: Mutter

last: Beltran

first: Jan

last: Brosens

first: Richard

last: Jenner


title: Localization of a TORC1-eIF4F translation complex during CD8+ T cell activation drives divergent cell fate

summary: Activated CD8+ T lymphocytes differentiate into heterogeneous subsets. Using super-resolution imaging, we found that prior to the first division, dynein-dependent vesicular transport polarized active TORC1 towards the microtubule-organizing center (MTOC) at the proximal pole. This active TORC1 was physically associated with active eIF4F, required for the translation of c-myc mRNA. As a consequence, c-myc translating polysomes polarized toward the cellular pole proximal to the immune synapse, resulting in localized c-myc translation. Upon division, the TORC1-eIF4A complex preferentially sorted to the proximal daughter cell, facilitating asymmetric c-Myc synthesis. Transient disruption of eIF4A activity at first division skewed long-term cell fate trajectories to memory-like function. Using a genetic barcoding approach, we found that first-division sister cells often displayed differences in transcriptional profiles that largely correlated with c-Myc and TORC1 target genes. Our findings provide mechanistic insights as to how distinct T cell fate trajectories can be established during the first division.

overall_design: CTV-labeled GFP-c-Myc OT-I CD8+ T cells were activated on peptide-pulsed APCs for 36 h. First-division GFP-c-Mychigh and GFP-c-Myclow cell populations (CD8+, CTV 2nd peak, highest and lowest 20% GFP-c-Myc) were sorted into medium with or without 200 nM Silvestrol and cultured for additional 2 h.

first: Swantje

last: Liedmann


title: Characteristic DNA methylation profiles of chorionic villi in recurrent miscarriage

city: Chiba

first: Masaki

last: Fukuyo

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

department: Department of Molecular Oncology

institute: Chiba University

type: Methylation profiling by genome tiling array

summary: Genome wide DNA methylation profiling of chorionic villi and decidua from recurrent miscarriage patients and artificial abortion controls. Infinium HumanMethylation450 BeadChip was used.

overall_design: Bisulphite converted DNA from the 20 samples were hybridised to the Infinium HumanMethylation450 BeadChip.

first: Yosuke

last: Matsumoto

first: Keiko

last: Shinjo

first: Yutaka

last: Kondo

first: Mayumi

last: Sugiura-Ogasawara

first: Atsushi

last: Kaneda


title: BAF complex cooperates with c-Myc to orchestrate early fate decisions of CD8 T cells (Microarray)

first: Douglas

last: Green

overall_design: We used microarrary to compare transcriptional profile of sgNTC ('Spike_Smarcd2')- or sgSmarcd2-transduced P14 cells from the dual-color transfer system at day 7.5 post-infection from spleen; we used microarrary to compare transcriptional profile of sgNTC ('Spike_Arid1a')- or sgArid1a-transduced P14 cells from the dual-color transfer system at day 7.5 post-infection from spleen

first: Hongling

last: Huang

first: Ao

last: Guo

first: Mark

last: Chen

first: Hongbo

last: Chi


title: BAF complex cooperates with c-Myc to orchestrate early fate decisions of CD8 T cells (ATAC II)

overall_design: ATAC-seq analysis of 12 samples: First division WT CD8 T cells (n=3), First division Arid1a KO CD8 T cells (ni=3), Activated WT CD8 T cells (n=3), Activated c-Myc KO CD8 T cells (n=3)


title: Genome-wide protein–DNA interaction site mapping using a double strand DNA-specific cytosine deaminase

city: Seattle

first: Joseph

last: Mougous

state: WA

address: 1959 NE Pacific St Box 357710

institute: University of Washington /HHMI

summary: DNA–protein interactions (DPIs) are central to such fundamental cellular processes as transcription and chromosome maintenance and organization. The spatiotemporal dynamics of these interactions dictate their functional consequences; therefore, there is great interest in facile methods for defining the sites of DPI within cells. Here, we present a general method for mapping DPI sites in vivo using the double stranded DNA-specific cytosine deaminase toxin DddA. Our approach, which we term DddA-sequencing (3D-seq), entails generating a translational fusion of DddA to a DNA binding protein of interest, inactivating uracil DNA glycosylase, modulating DddA activity via its natural inhibitor protein, and DNA sequencing for genome-wide DPI detection. We successfully applied this method to four transcription regulators that represent divergent protein families and bind variable numbers of chromosomal locations in two bacterial species. Our data show that 3D-seq offers several advantages over existing technologies, including ease of implementation and the ability to measure DPIs at single-cell resolution.

overall_design: Genome-wide locations of GacA in a parental strain lacking retS and genome-wide locations of GcsR and FleQ in the wild type PAO1 background.

first: James

last: Charity

first: Michael

last: Gebhardt

first: Simon

middle: L

last: Dove


title: Histone deacetylase inhibitor, panobinostat, exerts anti-proliferative effect with partial normalization from aberrant epigenetic states on granulosa cell tumor cell lines

status: Public on Apr 22 2022

city: Kurashiki

first: Takayuki

last: Tsujioka

address: Matsushima 577

department: Department of Laboratory medicine

institute: Kawasaki Medical School

summary:
The prognosis of the patients with inoperable or advanced granulosa cell tumors (GCTs) is still poor, and therefore it is important to establish a novel treatment strategy. Here we investigated the in vitro effects of a histone deacetylase inhibitor, panobinostat (PS) on two GCT cell lines (KGN and COV434).
GCT cell lines were found to be susceptible to PS treatment and it inhibited cell growth mainly by apoptosis. In cell cycle analysis, PS reduced only the ratio of S phase in GCT cell lines. Combined treatment of PS with a deubiquitinase inhibitor, VLX1570 enhanced the expression of p21, cleaved PARP, cleaved caspase-9, heme oxygenase-1, and the acetylation of histone H4 and α-tubulin, leading to an additive anti-proliferative effect on KGN and COV434. The gene set enrichment analysis revealed that PS treatment suppressed DNA replication- or cell cycle-related gene expression which led to chemotherapeutic cell death and in addition, this treatment induced activation of the gene set of adherens junction towards a normalized direction as well as activation of neuron-related gene sets that might imply unexpected differentiation potential due to epigenetic modification by a HDAC inhibitor in KGN cells. Exposure of KGN and COV434 cells to PS increased the expression of E-cadherin, one of the principal regulators associated with adherens junction in quantitative RT-PCR and immunoblotting analysis.   In the present study, we indicate a basis of a novel therapeutic availability of a HDAC inhibitor for the treatment of GCTs and further investigations will be warranted.

overall_design: KGN cells were treated with or without 100 nM of PS for 24 hr for RNA extraction and hybridization on Affymetrix microarrays.


title: BAF complex cooperates with c-Myc to orchestrate early fate decisions of CD8 T cells (ATAC I)

overall_design: ATAC-seq analysis of 9 samples: First division Myc high cells (n=3), First division Myc low cells (n=3), Naïve CD8 T cells (n=3)


title: RNA-seq of Setx KI and WT Spinal Cord

first: Laura

last: Campisi

address: 1468 Madison Avenue

department: Microbiology

institute: Icahn School Of Medicine at Mount Sinai

summary: The goal of this study is to compare differences in splicing between bulk spinal cord samples isolated from Setx KI and WT mice

overall_design: Spinal cords from Setx wildtype (WT; n=4) and Setx knock-in (KI; n=3) mice were isolated

first: Shahab

last: Chizari

first: Sook

middle: Y

last: Ho

first: Ivan

last: Marazzi


title: BAF complex cooperates with c-Myc to orchestrate early fate decisions of CD8 T cells (RNA-seq)

middle: R

overall_design: RNA-seq analysis of 20 samples: Activated CD8 T cells (n=5); Activated c-Myc KO CD8 T cells (n=5); First-division WT CD8 T cells (n=5); First division Arid1a KO CD8 T cells (n=5)


title: RNA-seq of ex vivo isolated, stimulated and re-stimulated CD8 T-cells

summary: The goal of this study is to compare differences in alternative splicing between stimulated and restimulated ex vivo isolated CD8 T cells.

overall_design: Naïve (CD44- CD62L+) CD8 T cells from the spleen of 5-6 weeks WT mice (n=3) were sorted using a BD FACSAria with 5 lasers (BD Bioscience). 105 naive CD8 T cells were stimulated in vitro with 4 ug/ml of purified anti-mouse CD28 and 20 ug/ml of plate-coated, purified anti-mouse CD3 in an incubator at 37ºC and 5% CO2. Recombinant human IL-2 (10 U/ml) was added to the culture at the end of day 1. After 3 days of culture, cells were left untreated or washed and additionally restimulated for 1 hour using a Cell Stimulation Cocktail, PMA/Ionomycin. In vitro activated CD8 T cells were then washed and resuspended in 1 ml TRIzol.


title: SMART-seq of PD1+ and PD1- CD8 T cells derived from Setx L389S (Setx KI) mice

summary: The goal is to understand the nature of PD-1+ CD8 T cells that are detected in the peripheral blood of ALS4 mice (expressing the mutation SetxL389S, also called Setx KI mice). We performed a SMART (Switching Mechanism At the end of the 5’-end of the RNA Transcript)-sequencing analysis of the transcriptome of effector memory CD8 T cells expressing or not PD-1 from the blood of symptomatic KI animals. Amongst the transcripts differentially expressed between these two populations, we detected upregulation of Tox (Thymocyte Selection Associated High Mobility Group Box) in PD-1+ as compared to PD-1- CD8 T cells.

overall_design: 500 CD44+CD62L-PD1+ and 500 CD44+CD62L-PD-1-CD8 T cells from 10-12 month old KI mice were sorted using a BD FACSAria with 3 or 5 lasers (BD Bioscience). Three individual mice were analyzed.

first: Jessica

middle: S


title: Feeding activates FGF15-SHP-TFEB-mediated lipophagy in the gut

city: Urbana

first: Jongsook

middle: Kim

last: Kemper

state: Illinois

address: 524 Burrill Hall  407 S. Goodwin Ave

department: Department of Molecular & Integrative Physiology

institute: University of Illinois at Champaign-Urbana

summary: Lysosome-mediated macroautophagy, including lipophagy, is activated under nutrient deprivation but is repressed after feeding. We show that feeding unexpectedly activates intestinal lipophagy in a manner dependent on both the orphan nuclear receptor, small heterodimer partner (SHP/NR0B2), and the late fed-state gut hormone, fibroblast growth factor-15/19 (FGF15/19). Postprandial intestinal triglycerides (TGs) and apolipoprotein-B48 (ApoB48), the TG-rich chylomicron marker, were elevated in SHP-knockout and FGF15-knockout mice. Genomic analyses in mouse intestine revealed that SHP partners with the key lysosomal activator, transcription factor-EB (TFEB), upregulating autophagy/lipolysis network genes after feeding. In HT29 intestinal cells, FGF19 treatment activated lipophagy in a manner dependent on both SHP and TFEB, reducing TG and ApoB48 levels. Mechanistically, feeding-induced FGF15/19 signaling increases nuclear localization of TFEB and SHP via PKCβ/ζ-mediated phosphorylation, leading to transcriptional induction of Ulk1 and Atgl. Collectively, these results demonstrate that after feeding, FGF15/19-activated SHP and TFEB paradoxically activate gut lipophagy, limiting postprandial TG levels. As excess lipids cause dyslipidemia and obesity, the FGF15/19-SHP-TFEB axis that reduces intestinal TGs via lipophagic activation provides promising therapeutic targets for obesity-associated metabolic disease.

overall_design: Identification of genome-wide intestinal SHP binding sites in SHP-KO and WT mice

first: Sunmi

last: Seok

first: Young-Chae

last: Kim

first: Yang

last: Zhang

first: Bo

last: Kong

first: Grace

first: Jian

last: Ma

first: Byron

middle: K


title: Alterations in gene expression due to aberrant DNA methylation are associated with the effects of anti-EGFR antibodies.

city: Sendai

first: Yasufumi

last: Otsuki

address: 4-1 Seiryo-machi, Aobaku

institute: Tohoku University

summary: Gene expression profiles of nine paired samples collected from formalin-fixed and paraffin embedded primary tumor and normal colonic mucosa in the OCH paired cohort.

overall_design: The purpose of this study was to clarify the significance of aberrant DNA methylation in regulating the expression of genes composing the cetuximab signature.  We collected and analyzed comprehensive DNA methylation and gene expression data of colorectal cancer in three tumor tissue cohorts and three normal colorectal mucosa - tumor tissue paired cohorts.

first: Kota

last: Ouchi

first: Shin

last: Takahashi


title: Transcriptome analysis of human lung andenocarcinoma cell line A549, in which ARAF was depleted and reconstituted with an empty vector control, ARAF wildtype, or a dimer deficient and  kinase inactive mutant.

city: Mainz

first: Federico

last: Marini

address: Obere Zahlbacher Strasse 69

institute: University Medical Center - Mainz

summary:
RNA sequencing analysis of A549 cells that were depleted for ARAF (shARAF) and reconstituted with either empty vector control (+EV) or ARAF wildtype (+ARAF) or dimer deficient ARAF kinase (+R362H) respectively to identify factors that are differentially regulated by ARAF and its kinase activity.
In particular, we discovered that ARAF regulates genes that are involved in proliferation and malignant growth by surpressing gene expression of members of the ERBB3/ AKT signaling pathway. At the same time, a functional interpretation of the differentially expressed (DE) genes with topGO indicated an important role of ARAF in regulation of genes involved in, among others, cell adhesion, cell motility, epithelial cell migration.

overall_design: Two biological replicates of A549 cells that were depleted for ARAF (shARAF), reconstituted with either empty vector control (+EV) or ARAF wildtype (+ARAF) and its kinase deficient mutant (+R362H) were seeded in a 6-well cell culture dish and left for growing (48h) before RNA isolation. After ice cold PBS-wash, cells were either detached with EDTA or directly lysed on plate using High Pure RNA Isolation kit (Cat#11828665001, Roche) according to maufacturer´s instructions. Total RNA was quantified by a Qubit 2.0 fluorometer (Invitrogen) and RNA quality and integrity was determined using a RNA6000 Nano chip on Bioanalyzer 2100 (Agilent). Samples with RNA integrity number (RIN) > 8 were further subjected for RNA library preparation. Barcoded cDNA libraries were prepared from 500 ng of total RNA using the NEBnext Poly(A) mRNA Magnetic Isolation Module and NEBNext Ultra RNA Library Prep Kit for Illumina (NEB) according to the provided instruction. Library quantity was assessed on a Qubit 2.0 using Invitrogen’s Qubit HS assay kit and library size was determined using Agilent’s Bioanalyzer 2100 and a HS DNA assay chip. Barcoded RNA-Seq libraries were onboard clustered using HiSeq Rapid SR Cluster Kit v2 using 8pM and 51 bps were sequenced on an Illumina HiSeq2500 using a HiSeq Rapid SBS kit v2.

first: Juliane

last: Mooz

first: Matthias

last: Klein

first: Krishnaraj

last: Rajalingam


title: Pubching spatial transcriptomics of MDA-MB-231 xenografts

city: Chuo-ku

first: Jun

last: Nakayama

address: Tsukiji 5-1-1

institute: National Cancer Center

summary: Gene expressional analysis with single cell scale by next generation sequencer revealed clonal dissemination in cancer metastasis. To reveal expressional heterogeneity and cell-cell interaction in the primary tumor and the metastasis, we performed transcriptome analysis of micro-tissues dissected from triple negative breast cancer (TNBC) cell line MDA-MB-231 xenograft model by our automated tissue micro-dissection punching technology. This “multiple micro-tissue transcriptome analysis” revealed that there existed three clusters in primary tumor and axillary lymph-node metastasis, two of which were cancer stem cell-like clusters (CD44/MYC-high, HMGA1-high).

first: Kentaro

last: Semba


title: BAF complex cooperates with c-Myc to orchestrate early fate decisions of CD8 T cells

first: Douglas

last: Green

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

overall_design: Refer to individual Series


title: Single-cell analysis reveals an indispensable role of SRSF2 in controlling fate of Myf5-expressing cells during skeletal muscle development

status: Public on Apr 21 2022

city: s

first: ruochen

last: guo

address: 320 Yueyang RD.

institute: Shanghai Institute of Nutrition and Health

summary: Skeletal muscle is an important tissue of animal body, which plays an important role in the process of motion, support and energy metabolism. Skeletal muscles are composed of striated myofibers bundles which formed during embryonic myogenesis and may undergo remodeling in the process of muscle regeneration after birth. SRSF2 is an important member of the splicing factor SR protein family. It plays an active role in transcription elongation, coupling transcription and splicing. A large number of studies have shown that it plays a key role in maintaining the normal development of thymus, heart and bone marrow. Here, we have used single cell RNA-seq and lineage analysis and gene knockout animal model to explore the regulation of splicing factor SRSF2 on myoblast cell fate and differentiation, as well as the mechanism in mouse skeletal muscle development.

overall_design: scRNA-seq: The SRSF2f/f mice were crossed with transgenic C57BL/6J mice expressing Cre recombinase driven by the Myf5 (myogenic factor 5) promoter to generate progeny (SRSF2f/w/Myf5-Cre). The SRSF2f/f mice were crossed with Rosa26-LSL-tdtomato (hereafter tdT) mice. After generated SRSF2f/f/tdT mice (both SRSF2 and tdT are homozygous). Next, mating SRSF2f/f/tdT with SRSF2f/w/Myf5-Cre mice to generate SRSF2f/f/Myf5-Cre/tdT mice. SRSF2f/w/Myf5-Cre/tdT littermates served as controls.

first: Ruochen

last: Guo

first: Ying

last: Feng


title: CITE seq on ALS4 and control CD8 T cells

city: New York

first: Laura

last: Campisi

summary: The goal of this study is to compare the transcriptional program and the TCR sequences of different subsets of CD8 T cells purified from the peripheral blood of 3 patienst with Amyotrophic Lateral Sclerosis 4 (ALS4) and 3 age-matched controls (unreleated). Peripheral blood mononuclear cells (PBMCs) were purified from 3 ALS4 patients and 3 age-matched healthy controls.By combining RNA and surface protein expression followed by multidimensional reduction using the Uniform Manifold Approximation and Projection (UMAP), we could identified 9 clusters corresponding to different CD8 T cell subsets: naïve/naïve like (clusters 0, 2, 5, 8), central memory (cluster 7), effector memory (clusters 4 and 6), TEMRA (clusters 1 and 3), and mucosal associated invariant T (MAIT) cells (cluster 9).   ALS4 patients exhibited a higher proportion of TEMRA,  and a decreased frequency of effector/effector memory cells, as compared to controls. Downregulation of IL7R and CCR7, and upregulation of GZMM and GZMB (encoding for Granzyme M and B, respectively), KLRB1 (Killer Cell Lectin Like Receptor B1), NKG7 (Natural Killer Cell Granule Protein 7), FGFBP2 (Fibroblast Growth Factor Binding Protein 2) and CCL4 (Chemokine (C-C motif) ligands 4), all of which correlate with a TEMRA phenotype are among the top deregulated genes in ALS4 CD8 T cells. The proportion of naïve cells seemed to be increased as well; however only 1 patient has an abnormal high frequency of CCR7+CD45RA+CD28+ cells, while the TEMRA signature was consistent among all 3 ALS4 donors.  More than 90% of expanded clones (≥5) are TEMRA cells in ALS4 patients, in comparison to ~ 60% in healthy controls. By matching UMAP coordinates from RNA and TCR analyses, we observed a colocalization between the most expanded clones, i.e. CD8 T cells in which the same TCR sequence was shared by more than 20 cells, and the TEMRA subset that we found enriched in patients. We quantified this clonal expansion and found that over half of all TEMRA cells comprised of highly expanded clones in patients. Hyperexpanded (>100 clones) TEMRA cells were detected in 2 of the 3 patients, with one clone reaching as high as 407 cells. No hyperexpanded clones were found in other CD8 T cell subsets or in control patients. Comparison of surface protein expression from our CITE-seq analysis in “Low” and “High” (>20) expanded clones from ALS4 patients revealed a stronger TEMRA phenotype in the latter, as indicated by CD27 downregulation and CD45RA and CD57 upregulation.

overall_design: CITEseq and scTCRseq of CD8 T cells isolated from the PBMC of 3 ALS4 and 3 age-matched controls.

first: Shahab

last: Chizari

first: Jessica

last: Ho

first: Ivan

last: Marazzi


title: BAF complex cooperates with c-Myc to orchestrate early fate decisions of CD8 T cells (ATAC III)

summary: We found cBAF and c-Myc co-assort asymmetrically to the two daughter nuclei, such that daughter cells with low c-Myc and cBAF display a cell fate trajectory towards memory T cell differentiation. Moreover, cBAF complex and c-Myc physically interact with each other, and act in establishing the chromatin landscape in activated CD8 T cells.

overall_design: ATAC-seq analysis of 9 samples: Activated WT CD8 T cells (n=3), Activated Arid1a KO CD8 T cells (n=3), Activated WT CD8 T cells treated with Arid1a inhibitor (n=3).

first: Ao

first: Hongling

last: Huang

first: Mark

middle: J

last: Chen

first: Hongbo

last: Chi


title: Effect of Pak4 deficiency on transcriptome in liver of mice

status: Public on Apr 20 2022

city: Jeonju-si

first: Byung-Hyun

last: Park

address: 567 Baekje-daero, Deokjin-gu

institute: Jeonbuk National University Medical School

summary: PAK4 has emerged as a promising target for anti-cancer drug development. However, its role in oxidative stress conditions remains elusive. We investigated the effects of PAK4 signaling on hepatic ischemia/reperfusion (I/R) injury. To elucidate molecular mechanisms, we performed RNA-sequencing analysis using littermate WT and hepatocyte-specific Pak4 KO mice in I/R condition. Transcriptomic analysis enabled us to find the main signaling nodes altered by Pak4 deficiency in close association with antioxidant response.

overall_design: Examination of RNA profile in liver of WT and hepatocyte-specific Pak4 KO mice.

first: Yuancheng

last: Mao

first: Chang

last: Han

first: Eun

last: Bae


title: BAF complex cooperates with c-Myc to orchestrate early fate decisions of CD8 T cells (Cut&Run)

overall_design: Cut&Run analysis of 27 samples: Brg1, Arid1a and c-Myc binding sites were analyzed in first division Arid1a KO and WT CD8 T cells; c-Myc KO and WT activated CD8 T cells. 2 replicates for each group.


title: RNA-seq analysis of transgenic nematodes GMC101 (Aβ1-42) and control strain CL2122

city: Shanghai

first: Hongru

last: Lin

state: Shanghai

address: Siping road 1239

department: life and science technology

institute: Tongji university

summary: We compared the transgenic nematodes GMC101 with control strain 2122.L1 stage nematodes were incubated at 20°C for 42h. Then we transferred the worms to 25°C  for Aβ induction.We chose 8h and 24h of induction as the detection time point.

overall_design: The time point of Aβ induction at 24h was repeated three times and the time point at 8h was repeated twice

first: Hongbing

last: Wang


title: ADAR1 downregulation contributes to p16INK4a upregulation independent of RNA editing during senescence

city: Philadelphia

first: Priyankara

last: Wickramasinghe

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute

summary: Cellular senescence plays a causal role in ageing and, in mouse, depletion of p16INK4a-expressing senescent cells delays ageing-associated disorders. Adenosine deaminases acting on RNA (ADARs) RNA editing enzymes are also implicated as important regulators of human ageing and ADAR inactivation causes age-associated pathologies such as neurodegeneration in model organisms. However, the role, if any, of ADARs in cellular senescence is unknown. Here we show that ADAR1 is post-transcriptionally downregulated by autophagic degradation to promote senescence through upregulating p16INK4a. ADAR1 is downregulated during senescence post-transcriptionally by autophagy-lysosomal pathway and the downregulation is sufficient to drive senescence in both in vitro and in vivo models. Senescence induced by ADAR1 downregulation is p16INK4a dependent and independent of its RNA editing function. Mechanistically, ADAR1 promotes SIRT1 expression by affecting its RNA stability through HuR, an RNA binding protein that increases the half-life and steady state levels of its target mRNAs. And SIRT1, in turn, antagonizes translation of mRNA encoding p16INK4a. Hence, downregulation of ADAR1 and SIRT1 mediates p16INK4aupregulation by enhancing its mRNA translation. Finally, Adar1 is downregulated during ageing of mouse tissues such as brain, ovary, and intestine, and Adar1 expression correlates with Sirt1 expression in these tissues in mice. Together, our study reveals an RNA-editing independent role of ADAR1 in regulating senescence by post-transcriptionally controlling p16INK4a expression.

overall_design:
RNA-seq experiment in ADAR control (Ctrl) and knock-down (KD) conditions in IMR90 cell lines.
HuR RIP-seq in control, shADAR1 and shHuR cell lines.

first: Xue

last: Hao

first: Rugang

last: Zhang


title: The single-cell transcriptional landscape of neuronal fates in the developing mouse cochlea

first: Louis

last: Faure

address: 321 E 61st Street

institute: Memorial Sloan Kettering Cancer Center

summary: The spiral ganglion neurons (SGNs) of the cochlea are essential for auditory perception by transmitting complex auditory information from hair cells (HCs) to the brain, yet the molecular mechanisms generating their diversity are unknown. Here we used single cell RNA sequencing to reconstruct the developmental trajectories of SGN cell fates and identified genes and gene regulatory networks that participate to changes in developmental competence and cell states, and in specification of each major cell types. Our analysis also identified gene modules associated with the sequential binary decisions that delineate neuron fate choices along the diversification tree. Transcriptome analysis of both developing SGN and HC types further revealed cell-state specific cell-cell signaling potentially playing a role in the differentiation and connectivity profile of SGNs as well as human deafness-associated genes, both previously known and novel. Overall, our results identify molecular principles that shape SGN differentiation and will facilitate further studies of SGNs development, physiology and dysfunction.

overall_design: Smart-Seq2 protocol was performed on single isolated cells by Eukaryotic Single Cell Genomics Facility at SciLifeLab, Stockholm. From the Ntrk3Cre;R26tdTOM E14.5 samples, we isolated a total of 135 cells, including 82 neurons and 53 OM. From the PVcre;R26tdTOM mice we isolated a total of 2139 cells: 229 at E15.5 (161 neurons and 68 OM), 661 at E16.5 (580 neurons and 73 OM and 8 HCs), 72 at E17.5 (71 neurons and 1 HC) and 667 at E18.5 (611 neurons and 66 HC). The P3 the transcriptional data was obtained from our previous study.

first: Charles

last: Petipre

first: Phoebe

last: Uhl

first: Paula

last: Fontanet

first: Iva

last: Filova

first: Gabriela

last: Pavlinkova

first: Igor

last: Adameyko

first: Saida

last: Hadjab

first: François

last: Lallemend


title: ScRNA-seq Expression of APOC2 and IFI27 Identifies Four Alveolar Macrophage Superclusters in Cystic Fibrosis and Healthy BALF

status: Public on Apr 19 2022

city: Lebanon

first: Claudia

middle: V

last: Jakubzick

state: NH

address: 1 Medical Center Drive, Rm 626 W Borwell, PO#1250960

department: Microbiology and Immunology

institute: Dartmouth College

summary: Alveolar macrophages (AMs) reside on the luminal surface of the airways and alveoli, ensuring proper gas exchange by ingesting cellular debris and pathogens, and regulating inflammatory responses. Therefore, understanding the heterogeneity and diverse roles played by AMs, interstitial macrophages, and recruited monocytes is critical for treating airway diseases. We performed single-cell RNA sequencing on 113,213 bronchoalveolar lavage cells from four healthy and three uninflamed cystic fibrosis subjects and identified two MARCKS+LGMN+IMs, FOLR2+SELENOP+ and SPP1+PLA2G7+ IMs, monocyte subtypes, DC1, DC2, migDCs, plasmacytoid DCs, lymphocytes, epithelial cells, and four AM superclusters (families) based on the gene expression of IFI27 and APOC2 These four AM families have at least eight distinct functional members (subclusters) named after their differentially expressed gene(s): IGF1, CCL18, CXCL5, cholesterol, chemokine, metallothionein, interferon, and small-cluster AMs. Interestingly, the chemokine cluster further divides with each subcluster selectively expressing a unique combination of chemokines. One of the most striking observations, besides the heterogeneity, is the conservation of AM family members in relatively equal ratio across all AM superclusters and individuals. Transcriptional data and TotalSeq technology were used to investigate cell surface markers that distinguish resident AMs from recruited monocytes. Last, other AM datasets were projected onto our dataset. Similar AM superclusters and functional subclusters were observed, along with a significant increase in chemokine and IFN AM subclusters in individuals infected with COVID-19. Overall, functional specializations of the AM subclusters suggest that there are highly regulated AM niches with defined programming states, highlighting a clear division of labor.

overall_design: We performed single-cell RNA sequencing (scRNA-seq) on 113,213 BAL cells from four healthy donors and three CF subjects. Transcriptional data and TotalSeq technology were used to confirm cell surface markers that distinguish resident AMs from recruited monocytes.

first: Xin

last: Li

first: Fred

last: Kolling

first: Daniel

last: Aridgides

first: Diane

last: Mellinger

first: Alix

last: Ashare


title: Ablation of cDC2 specification by triple mutations in the Zeb2 enhancer [CUT&RUN]

city: Saint Louis

first: Tiantian

last: Liu

address: 660 S. Euclid Ave.

department: Pathology and Immunology

institute: Washington University in St. Louis

summary: The divergence of the common dendritic cell progenitor (CDP) into specified progenitors for the cDC1 and cDC2 dendritic cells subsets is poorly understood. Some transcription factors (TFs) act in commitment of already specified progenitors, such as Batf3 which stabilizes Irf8 autoactivation at the +32 kb Irf8 enhancer, but the mechanism of CDP divergence remains unknown. Here, we report the transcriptional basis of CDP divergence and describe the first requirements for pre-cDC2 specification. Genetic epistasis analysis suggested that Nfil3 acts upstream of Id2, Batf3, and Zeb2 in cDC1 development but has not revealed its mechanism or targets. Analysis of newly generated NFIL3 reporter mice showed extremely transient NFIL3 expression during cDC1 specification. CUT&RUN and ChIP-seq analysis identified endogenous NFIL3 binding in the –165 kb Zeb2 enhancer at three sites that also bind CCAAT-enhancer-binding proteins C/EBPa and C/EBPb.  In vivo mutational analysis using CRISPR/Cas9 targeting showed that these NFIL3/C/EBP sites are functionally redundant, with C/EBPs supporting and NFIL3 repressing Zeb2 expression at these sites, respectively. Mutation of all three NFIL3/C/EBP sites ablated Zeb2 expression in myeloid, but not lymphoid progenitors, causing complete loss of pre-cDC2 specification and mature cDC2 development in vivo. These mice failed to generate TH2 responses against H. polygyrus infection, consistent with cDC2 supporting TH2 responses to helminths. Thus, CDP divergence is controlled by competition between NFIL3 and C/EBPs at the –165 kb Zeb2 enhancer.

overall_design: Examination of endogenous NFIL3 binding in the primary BM cells; NFIL3, C/EBPa and C/EBPb binding in the Hoxb8 cell lines by CUT&RUN.

first: Tian-Tian

first: Sunkyung

last: Kim

first: Kenneth

middle: M

last: Murphy


title: Ablation of cDC2 specification by triple mutations in the Zeb2 enhancer [RNA-seq]

overall_design: mRNA profiles of splenic cDC1 from WT and Nfil3GFP/GFP reporter mice. mRNA profiles of IL-4 and LPS stimulated splenic B cells from WT, Nfil3-/- and Nfil3GFP/GFP reporter mice.


title: Ablation of cDC2 development by triple mutations within the Zeb2 enhancer

summary: The divergence of the common dendritic cell progenitor (CDP) into the conventional type 1 and type 2 dendritic cell (cDC1 and cDC2, respectively) lineages is poorly understood. Some transcription factors act in the commitment of already specified progenitors-such as BATF3, which stabilizes Irf8 autoactivation at the +32 kb Irf8 enhancer-but the mechanisms controlling the initial divergence of CDPs remain unknown. Here we report the transcriptional basis of CDP divergence and describe the first requirements for pre-cDC2 specification. Genetic epistasis analysis suggested that Nfil3 acts upstream of Id2, Batf3 and Zeb2 in cDC1 development but did not reveal its mechanism or targets. Analysis of newly generated NFIL3 reporter mice showed extremely transient NFIL3 expression during cDC1 specification. CUT&RUN and chromatin immunoprecipitation followed by sequencing identified endogenous NFIL3 binding in the -165 kb Zeb2 enhancer at three sites that also bind the CCAAT-enhancer-binding proteins C/EBPα and C/EBPβ. In vivo mutational analysis using CRISPR-Cas9 targeting showed that these NFIL3-C/EBP sites are functionally redundant, with C/EBPs supporting and NFIL3 repressing Zeb2 expression at these sites. A triple mutation of all three NFIL3-C/EBP sites ablated Zeb2 expression in myeloid, but not lymphoid progenitors, causing the complete loss of pre-cDC2 specification and mature cDC2 development in vivo. These mice did not generate T helper 2 (TH2) cell responses against Heligmosomoides polygyrus infection, consistent with cDC2 supporting TH2 responses to helminths. Thus, CDP divergence into cDC1 or cDC2 is controlled by competition between NFIL3 and C/EBPs at the -165 kb Zeb2 enhancer.


title: Dissecting the transcriptome landscape of the human hippocampus

first: Yijing

last: Su

address: Levy 326

department: Department of Oral Medicine

institute: University of Pennsylvania

summary: Here we performed single-nucleus transcriptomic analysis of human hippocampus across the lifespan by split-seq approach

overall_design: Single-nucleus RNA-seq was performed using SPLiT-seq. Taking the advantage of the SPLiT-seq platform, we were able to pool multiple samples together for a run to reduce batch effects. Gene expression data of each individual sample could be demultiplexed and sorted out using provided barcodes.


title: A distinct immature low-density neutrophil population characterizes acute generalized pustular psoriasis [PBMC]

first: Ning

last: Yu

address: Baode Road 1278

institute: Shanghai Skin Disease Hospital

summary: Generalized pustular psoriasis (GPP) is a severe disease featured by neutrophilic pustules and enhanced IL-36 inflammatory pathway in skin. Recently, a transcriptomic analysis of PBMCs and neutrophils in MPO-deficient GPP patients has been performed in stable disease state. However, transcriptomic profiling of PBMCs during acute flare of GPP is unclear. Here, we reported a predominant neutrophil signature in GPP PBMC and a marked increase in the CD66+CD16+ low-density neutrophils (LDNs) within the PBMC fraction of acute GPP patients. Transcriptomic and functional analysis of LDNs revealed a hypoinflammatory phenotype yet enhanced release of neutrophil granule proteases, implicating that LDNs might contribute to the IL-36-mediated inflammation in GPP patients.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for peripheral blood mononulear cells (PBMCs) from 8 generalized pustular psoriasis (GPP) patients, 8 psoriasis vulgaris (PV) patients and 8 healthy donors.

first: Yuling

last: Shi


title: Ablation of cDC2 specification by triple mutations in the Zeb2 enhancer [ChIP-Seq]

overall_design: Examination of NFIL3 binding in the NFIL3-expressing Hoxb8 cell line by ChIP-seq.


title: PIK3CA and p53 mutations promote 4NQO-initated head and neck tumor progression and metastasis in mice

first: Richard

last: Friedman

address: 1130 St. Nicholas Av

department: Medicine

institute: Columbia University Irving  Medical Center

summary: The effect of gain of function mutations in p53, pik3ca, and both genes in mouse tongue subject to 4NQO treatment on gene expression is characterized by RNASeq.

overall_design:
WT: Wild type  squamous epithelium from mouse esophagus  (n=3).
p53 : Heterozygous LSL-p53+/R172H squamous epithelium from mouse tongue (n=3)
E20: Loxp-STOP-Loxp (LSL)-PIK3CAH1047R  squamous epithelium from mouse tongue (n=6)
E20/p53:  LSL- PIK3CAH1047R;LSL-p53+/R172H;K14-Cre squamous epithelium from mouse tongue (n=6)

first: Darío

last: García-Carracedo

first: Yi

last: Cai

first: Wanglong

last: Qiu

first: Kiyoshi

last: Saek

first: Andrew

last: Lee

first: Yinglu

first: Elizabeth

last: Goldberg

first: Elias

middle: E

last: Stratikopoulos

first: Ramon

last: Parsons

first: Chao

last: Lu

first: Argiris

last: Efstratiadis

last: Philipone

first: Angela

last: Yoon

first: Gloria

middle: H


title: Biologic correlates of COVID-19 resolution using single cell RNA sequecing

status: Public on Apr 18 2022

city: Newark

first: Natalie

last: Bruiners

state: NJ

address: 225 Warren street

institute: Rutgers University

summary: We report a case of full resolution of severe COVID-19 due to convalescent plasma transfusion. Following transfusion, the patient showed fever remission, improved respiratory status, and rapidly decreased viral burden in respiratory fluids and SARS-CoV-2 RNAemia. Longitudinal single-cell transcriptomics of peripheral blood cells conducted prior to and at multiple times after convalescent plasma transfusion identified the key biological processes associated with the transition from severe disease to disease-free state. This included post-transfusion disappearance of a subset of monocytes characterized by hyperactivated Interferon responses and decreased TNF-α signaling.

overall_design: We next utilized single-cell RNA sequencing to analyze transcriptional changes occurring in peripheral blood cells collected from the patient on the day of (prior to) the first transfusion with her relative’s plasma (i.e., day 54 post-symptom onset), and multiple times thereafter (day 56, 57, 59 and 61).

first: Maria

last: Gennaro

first: Ryan

last: Dikdan


title: Srebf1-mediated fatty acid metabolism is crucial to sustain sugmented pigmentation SREBF1-mediated metabolic reprogramming through fatty acid metabolism sustains augmented melanogenesis

city: Delhi

first: Vivek

last: Natarajan

state: Delhi

address: Mathura Road

country: India

institute: CSIR-IGIB

summary: Methods: mRNA profiles of B16 mouse melanoma cell line during pigmentation induced by culturing at low density were generated by deep sequencing, in dublicate. Library preparation was done using trueseq 4000 for quencing on Illumina paired end sequencing. The sequence read that passed the quality filter were aligned with GRCm38 as reference genome. Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample and identified 18753 transcripts in the pigmenting B16 melanocytes from D3 to D7 using HTSeq tool. R-values from Corelation plot varies from 0.98-0.99 between two replicates for different days.

overall_design: mRNA prolife of B16 melanocytes during transition from depigmented to pigmented state (D3-D7)

first: Rejesh

last: Gokhale

first: Farina

last: Sultan


title: Mutations of Wiskott-Aldrich Syndrome Protein cause oncogenic cell cycle dysreguation through disruption of alternative Splicing and epigenetic mechanisms

city: San Diego

first: April

middle: Elizabeth

last: Williams

state: California

address: 10010 N Torrey Pines Rd

department: IGC

institute: Salk Institute for Biological Studies

summary: Wiskott-Aldrich syndrome (WAS), which is caused by mutations in the gene encoding WASP, manifests in a wide range of hematologic and immune dysfunctions and predisposition to cancer development 1. WASP is most known as a cytoplasmic effector of actin cytoskeleton rearrangement. However, defective actin polymerization cannot explain many aspects of WAS pathogenesis. Incomplete knowledge of WASP function precludes in-depth understanding of the underlying mechanisms of WAS, and therefore hampers the development of effective therapies. Here we generated induced pluripotent stem cells (iPSCs) from WAS patients (WAS-iPSC) and corresponding isogenic iPSCs wherein the mutations were corrected by targeted genome editing. Hematopoietic cells differentiated from WAS-iPSCs not only recapitulated known disease phenotypes, but also revealed novel nuclear functions of WASP. WASP deficiency causes differential expression of genes involved in many aspects of immune cell function, most prominently cell proliferation. It also leads to a large number of alternative splicing events that are highly enriched in cell cycle regulators. Proteomic analyses revealed that WASP physically interacted with nuclear body components, nuclear structural proteins, chromatin modifying complexes, and many RNA-binding proteins including multiple splicing factors. We show that WASP physically interacts with SRSF2 and is important for nuclear speckle organization. WASP regulates cell cycle progression by physically interacting with EZH2 and thereby influencing epigenetic silencing of its target gene CDKN2A (aka p16). Together, these observations unveil novel functions of WASP in regulation of RNA splicing and cell cycle. The binding of WASP to SRSF2 and EZH2 and the misregulation of these two frequently mutated genes in hematologic malignancies2 in WAS cells provide a possible mechanism for the frequent occurance of malignancy in WAS patients. Furthermore, WAS-associated splicing and proliferation abnormalities could serve as diagnosis tools for patients at risk for cancer and may be exploited as therapeutic targets.

overall_design: Total mRNA was extracted from patient and control samples, respecitively.  RNA-Seq was used to determine the differential gene expressions between patient and control in three different cell types (macrophage, T cell, and B cell).

first: Mo

last: Li

first: Ling

last: Huang


title: Transcriptomic sequencing analysis of glioblastoma cell lines DBTRG and U251.

city: Wuhan

first: Yiqi

last: Wang

state: State...

address: Shizishan Street

institute: Huazhong Agricultural University, China

summary: We showed that DBTRG is more invasive than U251 cell lines by novel brain-stiffness-mimicking matrix gel invasion platform and transwell invasion assay. To understand the molecular mechanisms of DBTRG being more invasive than U251, we performed transcriptomic sequencing analysis of DBTRG and U251 cell lines.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for DBTRG cells and U251 cells.

first: Shuowen

first: Jin

last: Xiong

first: Wendai

last: Bao

first: Yaqi

first: Jun

last: Qin

first: Guang

last: Han

first: Sheng

last: Hu

first: Junrong

last: Lei

first: Zehao

last: Yang

first: Yu

last: Qian

first: Shuang

last: Dong

first: Zhiqiang


title: Next generation sequencing for iPSC-derived macrophages

city: Jeddah

first: Baolei

last: Yuan

state: Makkah

address: Thuwal

country: Saudi Arabia

institute: King Abdullah University of Science and Technology

summary: We performed the next-generation sequencing for an isogenic WASP-KO macrophage model. Above 65 million clean reads per sample were obtained. We found the numerous RNA splicing asscoiated genes were overexpressed and thousands of  mRNA events  were aberrantly spliced in WASP-KO macrophages.

overall_design: RNA proliles of WT and isogenic WASP-KO macrophages derived from iPSCs

first: Xuan

last: Zhou


title: YY1 safeguard multidimensional epigenetic landscape associated with extended pluripotency

city: guangzhou

first: xiaotao

last: dong

address: 190 Kai Yuan Avenue, Science Park

institute: Guangzhou Institutes of Biomedicine and Health Chinese Academy of Sciences

summary: Although extended pluripotent stem cells (EPSCs) have the potential to form both embryonic and extraembryonic lineages, how their transcriptional regulatory mechanism differs from that of embryonic stem cells (ESCs) remains unclear. Here, we discovered that YY1 binds to specific open chromatin regions in EPSCs. Yy1 depletion in EPSCs leads to a gene expression pattern more similar to that of ESCs than control EPSCs. Moreover, Yy1 depletion triggers a series of epigenetic crosstalk activities, including changes in DNA methylation, histone modifications and high-order chromatin structures. Yy1 depletion in EPSCs disrupts the enhancer-promoter (EP) interactions of EPSC-specific genes, including Dnmt3l. Yy1 loss results in DNA hypomethylation and dramatically reduces the enrichment of H3K4me3 and H3K27ac on the promoters of EPSC-specific genes by upregulating the expression of Kdm5c and Hdac6 through facilitating the formation of CCCTC-binding factor (CTCF)-mediated EP interactions surrounding their loci. Furthermore, single-cell RNA sequencing (scRNA-seq) experiments revealed that YY1 is required for the derivation of extraembryonic endoderm (XEN)-like cells from EPSCs in vitro. Together, this study reveals that YY1 functions as a key regulator of multidimensional epigenetic crosstalk associated with extended pluripotency.

overall_design: Gene expression profiling of ES cells, EPS cells, Ctrl shRNA- and Yy1-depleted EPS cells. Examination of YY1, H3K27ac, H3K4me1 binding sites in both ES cells and EPS cells.  Examination of CTCF, H3K4me3 binding sites in Ctrl shRNA- and Yy1-depleted EPS cells. Chromatin interactions in Ctrl shRNA- and Yy1-depleted EPS cells. Examination of the chromatin open states in ES cells and EPS cells. DNA methylation profiling in Ctrl shRNA- and Yy1-depleted EPS cells.

first: Hongjie

last: Yao

first: Xiaotao

first: Rong

last: Guo


title: Retrograde movements determine effective stem cell numbers in intestinal crypts

city: Amsterdam

first: Jacco

last: van Rheenen

state: Noord holland

address: Plesmanlaan 121

department: Molecular Pathology

country: Netherlands

institute: Netherlands Cancer Institute

summary: Our goal was to compare the transcriptomic profile of lgr5+ high, medium and low cells (corresponding to centre, border and >3 row in crypts) between small and large intestine in mice. We observed differences in the transcriptional profile between centre and border Lgr5+ cells within small and large intestinal crypts and more profound differences between the two intestinal sites.

overall_design: mRNA profiles of lgr5+ high, medium and low cells (corresponding to centre, border and >3 row in crypts) between small and large intestine in mice.

first: Maria

last: Azkanaz


title: Clonal structure and dynamics of human memory B cells and circulating plasmablasts

city: Bellinzona

first: Ganesh

last: Phad

address: Via Francesco Chiesa 5

country: Switzerland

institute: Institute for Research in Biomedicine

summary: Using single cell sequencing of multi-year serial blood samples from two healthy donors we found that the memory B cell repertoire was dominated by large clonal families producing IgM, IgG2 and IgA, of which 0.2% share nearly identical VH/VL sequences in different individuals. In contrast, IgG1 families, including those encoding antibodies to T-dependent antigens, were of small size. Longitudinal samples collected over a period of several years showed the overall stability of the memory B cell pool in all its subsets. Surprisingly, clonotype stability was also found in serial samples of circulating plasma cells that largely overlapped with long-term memory B cells and contained recurrent clonotypes of IgA and IgG2 and, at lower frequency, IgG1 specific for recall antigens. Collectively, this study provides a global view of the structure, stability and dynamics of the human memory B cell pool and reveal that a large fraction of the clonal families is active at any time point in the generation of plasma cells involving both T independent and T dependent responses.

overall_design: Single cell immune profiling (gene expression) of plasma cells from healthy human donor

first: Antonio

last: Lanzavecchia

first: Murodzhon

last: Akhmedov


title: Lack of VMP1 Impairs Hepatic Lipoprotein Secretion and Promotes Nonalcoholic Steatohepatitis

city: Kansas City

first: Hong-Min

last: Ni

state: KS

address: 3901 Rainbow Blvd, MS-1018

department: Pharmacology, Toxicology and Therapeutics

institute: University of Kansas Medical Center

summary: The purpose of this study is to identify the differential transcriptome profiles in WT and hepatocyte deletion of VMP1 mouse livers. Floxed VMP1 mice were intravenously injected with AAV8-TBG-null or AAV8-TBG-cre for 2 weeks. Liver mRNA were extracted and performed for Nextseq analysis in triplicates.

overall_design: Hepatic mRNA profiles of WT (floxed VMP1 injected with AAV8-TBG-null for 2 weeks) and Hepatocyte specific VMP1 KO (floxed VMP1 injected with AAV8-TBG-cre for 2 weeks).

first: Wen-Xing

last: Ding

first: Linheng

last: Li

first: Shiyuan

last: Chen


title: ΔfruB effects on the transcriptome of Streptococcus mutans probed by RNA-seq

status: Public on Apr 15 2022

city: Gainesville

first: Robert

last: Burne

state: Fl

address: 1395 Center Drive

department: Oral Biology

institute: University of Florida

summary: Streptococcus mutans, the primary etiologic agent of human dental caries, and a variety of oral Streptococcus and Actinomyces spp. synthesize high molecular mass homopolymers of fructose (fructans) with predominantly β2,1- (inulins) or β2,6-linkages (levans). The ability of S. mutans to degrade fructans contributes to the severity of dental caries. The extracellular product of fruA of S. mutans is an exo-β-D-fructofuranosidase that releases fructose from levan and inulin. Located 70 bp downstream of fruA, fruB encodes a member of the glycoside hydrolase family 32, but the function of FruB has not been established.In this study, the transcriptomic analysis of UA159 and a fruB mutant grown on 0.2% levan revealed differential expression of genes encoding ABC transporters, transcriptional regulators and genes involved in growth and stress tolerance. The results provide new insights into the mechanisms of action by which FruB utilizes homo-polymers of fructose, specifically levan. The findings significantly enhance our understanding of the genetics and physiology of this cariogenic pathogen S. mutans.

overall_design: mRNA profiling of S. mutans UA159 WT and ΔfruB inTryptone-Vitamin (TV) medium supplemented with 0.2% levan.

first: Brinta

last: Chakraborty


title: Next generation sequencing to determine DNA-binding sites of Wiskott-Aldich Syndrome Protein (WASP)  in the cell line GM 11518 (wild-type) and in the cell line ID00003/WASP-c-EOS (WASP overexpressed)

city: Thuwal

first: Gerardo

last: Ramos-Mandujano

department: Biological and Environmental Science and Engineering Division

summary: To determine the global gene occupancy by Wiskott - Aldrich syndrome Protein (WASP) we perform ChIP-seq assay in two lymphoblastoid cell lines. We identify WASP-enriched genes, including several WASP-interaction genes previously reported; in addition, our results suggest the implication of WASP in diverse cellular process

overall_design: Using ChIP-seq strategy we identify the DNA-binding sites by WASP in two lymphoblastoid cell lines

first: M

first: G

first: L

last: Cortes-Medina


title: Immunotherapy study of phagocytosis checkpoint blockade in combination with radiotherapy

city: Houston

first: Michael

last: Curran

state: TX

address: 1515 Holcombe Blvd.

department: Immunology

institute: MD Anderson Cancer Center

summary: Transcriptional profiling of tumor-infiltrating antigen-presenting myeloid subsets and CD8 T cells in mice bearing MC38-OVA tumors treated with or without radiotherapy, αSIRPα, and/or αPD-1 treatment

overall_design: CD11b-CD11c+, CD11b+CD11c+, and CD11b+CD11c- myeloid populations and CD8 T cells were isolated from the irradiated tumors 5 days after 8-Gy or sham irradiation +/- αSIRPα and αPD-1 for RNA-sequencing analyses.

first: Rodney Cheng-En

last: Hsieh

first: Wen-Hao

last: Hsu