title: IgE-binding monocytes upregulate the coagulation cascade in allergic horses

city: Ithaca

first: Bettina

last: Wagner

state: NY

address: 240 Farrier Drive

department: Population Medicine and Diagnostic Sciences

institute: Cornell University

summary: IgE-binding monocytes are a rare peripheral immune cell type involved in the allergic response through binding of IgE on their surface. IgE-binding monocytes are present in both healthy and allergic individuals. We performed RNA sequencing to ask how the function of IgE-binding monocytes differs in the context of allergy. Using a large animal model of allergy, equine Culicoides hypersensitivity, we compared the transcriptome of IgE-binding monocytes in allergic and non-allergic horses at two seasonal timepoints: (i) when allergic animals were clinical healthy, in the winter “Remission Phase”, and (ii) during chronic disease, in the summer “Clinical Phase”. Most transcriptional differences between allergic and non-allergic horses occurred only during the “Remission Phase”, suggesting principal differences in monocyte function even in the absence of allergen exposure. F13A1, a subunit of fibrinoligase, was significantly upregulated at both timepoints in allergic horses. This suggested a role for increased fibrin deposition in the coagulation cascade to promote allergic inflammation. IgE-binding monocytes also downregulated CCR10 expression in allergic horses during the “Clinical Phase”, suggesting a defect in maintenance of skin homeostasis, which further promotes allergic inflammation. Together, this transcriptional analysis provides valuable clues into the mechanisms used by IgE-binding monocytes in allergic individuals.

overall_design: Flow cytometry isolated IgE-binding monocytes were purified from equine peripheral blood. Monocytes were purified from allergic (n=7) and healthy (n=5) horses during allergen exposure (Clinical Phase in August-September) and when there was no allergen exposure (Remission Phase in January-February).

first: Elisabeth

last: Simonin


title: RNA-seq transcriptomic analyses of group B Streptococcus Cas9 variants and CRISPRi knockdown strains

status: Public on May 15 2023

city: Pittsburgh

first: Thomas

middle: Alexander

last: Hooven

state: PA

address: 4401 Penn Ave., Rangos Research Bldg #8128

department: Pediatrics

country: USA

institute: University of Pittsburgh

type: Expression profiling by high throughput sequencing

summary: Group B Streptococcus (GBS) strain CNCTC 10/84 was modified to have specific mutations to the cas9 gene: allelic exchange replacement with a chloramphenicol resistance marker (Cas9 knockout), D10A and H845A (catalytically inactive dCas9), or R1339A and R1441A (sCas9 unable to scan for protospacer adjacent motifs). Wild type (WT) and mutant strains were grown in biological triplicate samples and used for RNA purification at two growth timepoints: OD600=0.6 and OD600=1.2. Additionally, CNCTC 10/84 dCas9 and A909 dCas9 were transformed with p3015b expression plasmids expressing sgRNA cassettes designed to downregulate the cyl operon in CNCTC 10/84 dCas9 and the covR/covS two-component system in A909 dCas9. Triplicate samples were grown alongside control transformants bearing "sham" sgRNA plasmid. RNA was purified at OD600=1.2. All RNA samples were used for RNA-seq and subsequent bioinformatic analyses.

overall_design: Comparative transcriptomics of CNCTC 10/84 Cas9 variants (KO, dCas9, sCas9, and WT) between and across two growth phases (OD 0.6 and OD 1.2). Further comparison of CNCTC 10/84 dCas9 and A909 dCas9 at OD 1.2 in which specific genes (cyl and CovR, respectively) have been downregulated; these comparisons are made to "sham" controls in which the sgRNA does not target any chromosomal genes.

first: Kathyayini

middle: P

last: Gopalakrishna

first: Gideon

middle: H

last: Hillebrand

first: Venkata

last: Bhavana

first: Jordan

middle: L

last: Elder

first: Adonis

last: D'Mello

first: Hérve

last: Tettelin

middle: A


title: Transcription effect of LY2835219 on the U2OS cell line expressing EWS-FLI1

city: Beijing

first: Chunyu

last: Yu

address: Xueyuan Road 38#

country: China

institute: Peking University

summary: Cancer-associated chromosomal rearrangements can result in the expression of numerous pathogenic fusion proteins. The mechanisms via which the fusion proteins contribute to oncogenesis are largely unknown and effective therapies for fusion-associated cancers are lacking. Here, we comprehensively scrutinized fusion proteins found in various cancers. We found that many fusion proteins are composed of phase separation-prone domains (PS) and DNA-binding domains (DBD), and these fusions have strong correlations with aberrant gene expression patterns. Furthermore, we established a high-throughput screening method, named DropScan, to screen drugs capable of modulating aberrant condensates. One of the drugs identified via DropScan, LY2835219, effectively dissolved the condensates in reporter cell lines expressing Ewing sarcoma fusions and partially rescued the abnormal expression of target genes. Our results indicate that aberrant phase separation is likely a common mechanism for these PS-DBD fusion-related cancers, and suggest that modulating aberrant phase separation is a potential route to treat these diseases.

overall_design: Wild type U2OS cell lines, U2OS integrated with mCherry, mCherry-EWS-FLI1, mCherry-EWS-FLI1mut, and mCherry-EWS(37YS)-FLI1 supplemented with 10% FBS (Gibco) and 100 units/mL penicillin and 100 μg/mL streptomycin (HyClone) at 37℃ with 5% CO2 in a humidified incubator (Above 10^7 cells in 10cm dish). Replicates of mCherry-EWS-FLI1 were treated with 7.5μM LY2835219 for 6 or 24 hours.

first: Pilong

last: Li


title: Transcriptome profile analysis reveals putative molecular mechanisms of 5-aminolevulinic acid toxicity

city: São Paulo

first: Janice

last: Onuki

state: SP

address: Av. Vital Brasil, 1500 - Butantã

country: Brazil

institute: Butantan Institute

summary: Acute hepatic porphyrias (AHPs) are inherited metabolic diseases characterized by decreased activity or reduction in synthesis of the heme biosynthetic pathway enzymes, leading to acute attacks of abdominal pain, hypertension, neuropsychiatric alterations, and a long-term increase in the incidence of hepatocellular carcinoma (HCC). 5-aminolevulinic acid (ALA) seems to be the major pathogenic factor of AHPs. Studies only reported biological effects on specific macromolecules, thus we investigated the global transcriptional changes and perturbed molecular pathways in HepG2 cells following exposure to ALA 2.5 mM or 25 mM for 2 h and 24 h using DNA microarray. The identification and analysis of differentially expressed genes (DEGs) showed distinct transcriptome profiles from each ALA treatment. Molecular pathways and biological processes enrichment analysis of DEGs by KEGG, Reactome, Metacore, and Gene Ontology, showed that ALA ‘2.5 mM-2h’ induced perturbation in the mitochondrial respiratory chain, ion homeostasis, cell proliferation/apoptosis, and UPR/ER stress, which were connected by the network hubs AP1, C/EBP, ATF-6. ALA ‘25 mM-24h’ elicited changes in oxidative stress, cell death/survival, immune response, drug detoxification, and carcinogenesis with AP-1, C/EBP, EGR1, STAT5, Cyclin D1, JAK2, ERK1/2, and RAC1 as the main network hubs and SQSTM1, HMOX1, SPP1, and LONP-1 genes as putative effectors. This study contributes to revealing molecular mechanisms of ALA potentially associated with the pathogenesis of the clinical manifestations of AHPs.

overall_design: Two-condition experiment, ALA 2.5 mM - 2 h vs. Control cells, ALA 2.5 mM - 24 h vs. Control cells, ALA 25 mM - 2 h vs. Control cells, ALA 25 mM - 24 h vs. Control cells. Biological replicates: 4 '2.5 mM 2 h' replicates, 4 '2.5 mM 24 h' replicates, 4 '25 mM 2 h' replicates, 4 '25 mM 24 h' replicates.

first: Patricia

middle: R

last: Menezes

first: Carlos

middle: E

last: Trufen

first: Flavio

last: Lichtenstein

first: Diogo

middle: V

last: Pellegrina

first: Eduardo

middle: M

last: Reis


title: Transcriptomic analysis reveals the gene regulatory networks involved in leaf and root response to osmotic stress in tomato

city: Milano

first: Raul

last: Pirona

address: Via Alfonso Corti 12

country: Italy

institute: IBBA-CNR

summary: We performed a transcriptomic analysis on two tomato genotypes, M82 and Tondo, in response to a PEG-mediated osmotic treatment, mimicking water stress. The analysis was conducted separately on leaves and roots to characterize the specific response of these two organs. A total of 6,267 differentially expressed transcripts related to osmotic stress response was detected. The construction of gene co-expression networks defined the metabolic and signaling pathways of the common and specific responses of leaf and root. The common response was characterized by ABA-dependent and ABA-independent signaling pathways, and by the interconnection between ABA and JA signaling. The root specific response concerned genes involved in cell wall metabolism and remodeling, whereas the leaf specific response was principally related to leaf senescence and ethylene signaling. The transcription factors representing the hubs of these regulatory networks were identified. Some of them have not yet characterized and can represent novel candidates for tolerance. Finally, several genes showing a genotype-specific expression regulation in response to the treatment were detected. These genes may be involved in the different sensitivity to the osmotic treatment of the two tomato genotypes. In conclusion, this work shed new light on the regulatory networks occurring in tomato leaf and root under osmotic stress and set the base for an in-depth characterization of novel stress-related genes, that may represent potential candidates for improving tolerance to water stress in tomato.

overall_design: The study used 24 arrrays with RNA extracted from two tomato genotypes (Tondo and M82), two tissues (leaf and root), in control and under PEG conditions (2 genotypes x2 conditions x2 tissues x3 biological replicates= 24 arrays).

first: Giovanna

last: Frugis

first: Franca

last: Locatelli

first: Monica

last: Mattana

first: Annamaria

last: Genga

first: Elena

last: Baldoni


title: Control of alveolar macrophage differentiation by Siah1a/2 ubiquitin ligases limits carcinogen-induced lung adenocarcinoma [Bulk RNA-seq]

city: La Jolla

first: Ze'ev

middle: A.

last: Ronai

state: CA

address: 10901 North Torrey Pines Road

department: Tumor Initiation and Maintenance Program, NCI-Designated Cancer Center

institute: Sanford Burnham Prebys Medical Discovery Institute

summary: Tumor microenvironment components, including T and myeloid cells, play important roles in lung adenocarcinoma (LADC) progression and response to therapy. Here, we identify a role for Siah1a/2 ubiquitin ligases in controlling alveolar macrophage (AM) differentiation and urethane-induced LADC. Genetic ablation of Siah1a/2 in AMs enriched their immature state, coinciding with increased pro-tumorigenic and inflammatory gene signatures and abundance of Siah1a/2 substrates NRF2 and B-catenin. Urethane administration to mice enriched the population of monocytic and immature-like AMs, which were more prevalent in the lungs of mice carrying Siah1a/2-ablated macrophages. While resembling transitional profibrotic macrophages often seen in lung fibrosis, enrichment of immature AMs coincided with the development of more frequent and larger lung tumors in urethane-treated mice harboring Siah1a/2 ablated macrophages compared with controls. Gene expression signature of Siah1a/2 ablated immature-like AMs is associated with increased infiltration of CD14+ immune cells and worse survival of LADC patients. Our findings identify Siah1a/2 as gatekeepers of cancer development by controlling AM differentiation and profibrotic phenotypes contributing to carcinogen-induced lung cancer.

overall_design: Gene expression comparisons of alvelolar macrophages from CD11c+ F480+ SiglecF+ WT, cSiah1a/2f/f::Lyz2cre, and cSiah1a/2f/f::CD11ccre mice using RNA-seq.

first: Marzia

last: Scortegagna


title: Control of alveolar macrophage differentiation by Siah1a/2 ubiquitin ligases limits carcinogen-induced lung adenocarcinoma

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

overall_design: Refer to individual Series


title: Construction of ceRNA network based on an RNA‑seq shows prognostic lncRNA biomarkers in Perthes’ disease

city: Shanghai

first: hu

last: xiaolin

state: Shanghai

address: south congqing road 227#

institute: Shanghai Jiaotong University School of Medicine

type: Non-coding RNA profiling by high throughput sequencing

overall_design: Comparative gene expression profiling analysis of RNA-seq data for Perthes' disease and wild type rabbit

first: Zhang

last: Tianjiu

first: Wei

last: Chunyan

first: Hu

last: Xiaolin


title: Control of alveolar macrophage differentiation by Siah1a/2 ubiquitin ligases limits carcinogen-induced lung adenocarcinoma [scRNA-seq]

overall_design: Cd45+ cells from WT and Siah1a/2 mutant mouse lungs were sorted using flow cytometry and gene expression analyzed using 10X Genomic scRNA-seq protocol.


title: Microenvironmental Control of Hematopoietic Niche Capacity via CXCL8 and Protein Kinase C

city: Columbus

first: Bradley

middle: Wayne

last: Blaser

state: OH

address: 400 W 12th Ave

department: Comprehensive Cancer Center

institute: Ohio State University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Altered hematopoietic stem cell (HSC) fate underlies primary blood disorders but microenvironmental factors controlling HSC fate are poorly understood.  Genetically barcoded GESTALT zebrafish were used to screen for factors expressed by the sinusoidal vascular niche that alter the phylogenetic distribution of the HSC pool under native conditions.  Dysregulated expression of protein kinase C delta (PKC-d, encoded by prkcda) increased the number of HSC clones by approximately 50% and expanded polyclonal populations of immature neutrophil and erythroid precursors.  PKC agonists such as cxcl8 augmented HSC competition for residency within the niche and expanded defined niche populations.  Cxcl8 induced association of PKC-d with the focal adhesion complex, activating MEK/ERK signaling and expression of niche factors in human endothelial cells.  Our findings demonstrate the existence of reserve capacity within the niche which is controlled by PKC and has significant impact on HSC phylogenetic and phenotypic fate.

sample_organism:
Danio rerio
Homo sapiens

platform_organism:
Danio rerio
Homo sapiens

overall_design: Single Cell RNA-seq and Single Cell ATAC-seq from zebrafish embryos and zebrafish kidney marrow.  Bulk RNA-seq and bulk ATAC-seq from HUVEC cells transduced with adenovirus E4ORF1.

middle: W


title: Mechanisms of stop codon readthrough mitigation reveal principles of GCN1 mediated translational quality control

city: Martinsried/Planegg

first: Martin

last: Mueller

address: Am Klopferspitz 18

country: Germany

institute: MPI of Biochemistry

summary: Readthrough into the 3′ untranslated region (3′UTR) of the mRNA results in the production of aberrant proteins. Metazoans efficiently clear readthrough proteins, but the underlying mechanisms remain unknown. Here, we show in C. elegans and mammalian cells that readthrough proteins are targeted by a coupled, two-level quality control pathway involving the BAG6 chaperone complex and the ribosome collision-sensing protein GCN1. Readthrough proteins with hydrophobic C-terminal extensions are recognized by SGTA-BAG6 and ubiquitylated by RNF126 for proteasomal degradation. Additionally, cotranslational mRNA decay initiated by GCN1 and CCR4/NOT limits the accumulation of readthrough products. Unexpectedly, selective ribosome profiling uncovered a general role of GCN1 in regulating translation dynamics when ribosomes encounter nonoptimal codons, a feature of 3′UTR sequences. Dysfunction of GCN1 results in mRNA and proteome imbalance, increasingly perturbing transmembrane proteins and collagens during aging. These results define GCN1 as a key factor acting during translation in maintaining protein homeostasis.

sample_organism:
Caenorhabditis elegans
Homo sapiens

platform_organism:
Caenorhabditis elegans
Homo sapiens

overall_design: Selective Ribo-seq of GCN1 in C.elegans and HEK293 with respective input controls and additional mRNAseq

middle: B

first: Gopal

middle: G

last: Jayaraj

first: F.

middle: U

last: Hartl


title: pre-mRNA splicing order is defined and maintains splicing fidelity across multi-intronic transcripts

status: Public on May 14 2023

city: Sherbrooke

first: Karine

last: Choquet

state: Québec

address: 3201 rue Jean-Mignault

department: Biochemistry and Functional Genomics

country: Canada

institute: Université de Sherbrooke

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct


title: Effect of sphongosine 1-phosphate (S1P) treatment on gene expression in human myometrial cells.

city: St. Louis

first: Antonina

middle: I.

last: Frolova

state: MO

address: 660 S. Euclid Ave, CB 8064

department: OBGYN

institute: Washington University in St. Louis

summary: To gain further insight into the possible mechanism(s) by which S1P regulates cell signaling pathways in human myometrial cells, RNAseq was performed using two immortalized myometrial cell lines following treatment with exogenous S1P or vehicle control for 3 h.

overall_design: Two immortalized human myometrial cell lines were used in the experiments. hTERT-HM (hHM) is an hTERT-immortalized myometrial cell line obtained from a non-pregnant human uterus at the time of hysterectomy and were a kind gift from Dr. Jennifer Condon. The second cell line, HM6ERM2 (HM6), was previously derived from term-pregnant human uterine tissue and immortalized using hTERT as previously described. Cells were treated with either S1P or vehicle control ( bovine serum albumin).  Each treatment was performed in triplicate for each of the two cell lines.

middle: I

first: Kumar

last: Saurabh


title: The Single-cell and spatial transcriptional landscape of human development

first: Zeyuan

last: Liu

address: 15 Datun Road, Chaoyang District, Beijing, 100101, China

institute: Institute of Biophysics,Chinese Academy of Sciences

summary: The understanding of human early embryo developoment is very import for us to resovle the mysteries of how human organs especially the brain works and how human disease come up.Therefor, we carried out single cell RNA-seq and spatial transcriptional RNA-seq of human early embryo and analysed gene expression dynamics during the embryo development.We found some import genes and pathways that may control the ongoing of development.These findings help us make a step in understanding how we human being grow from a simple embryo to a complex one.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: single-cell RNA-seq and spatial RNA-seq for early human embryo

first: Bo

last: Zeng

first: Suijuan

last: Zhong

first: Xiaoqun

last: Wang

first: Qian

last: Wu


title: The MLL – Menin interaction is a therapeutic vulnerability in NUP98-rearranged AML

city: UTRECHT

first: Olaf

last: Heidenreich

state: UTRECHT

address: Prinses Máxima Centrum voor kinderoncologie | Postbus 113| 3720 AC BILTHOVEN | Heidelberglaan 25 |  3584CS UTRECHT | Nederland

country: Netherlands

summary: To investigate the effect of Menin inhibitor on patient-derived NUP98-r AML. We treated primary AML cells with Menin inhibitor in a Mesenchymal stem cell co-Culture platform. Then we charachterized global gene expression in treated AML cells upon treatment in two different time points, day7 and day12.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for patient-derived NUP98::NSD1 AML cells after treatment with Menin inhibitor (SNDX-5613/Revumenib).

first: Milad

last: Rasouli

first: Helen

last: Blair

first: Selina

last: Troester

first: Katarzyna

last: Szoltysek

first: Rachel

last: Cameron

first: Minoo

last: Ashtiani

first: Anja

last: Krippner-Heidenreich

first: Florian

last: Grebien

first: Gerard

last: McGeehan

first: C

last: Zwaan


title: Liver lipophagy ameliorates nonalcoholic steatohepatitis through lysosomal lipid exocytosis

city: Kamigyo-ku, Kyoto-shi

first: Atsushi

last: Hoshino

state: Kyoto-hu

address: 465, Kajii-chou

country: Japan

institute: Kyoto Prefectural University of Medicine

summary: Nonalcoholic steatohepatitis (NASH) is a progressive disorder with aberrant lipid accumulation and subsequent inflammatory and profibrotic response. Lipid reduction through cytoplasmic lipolysis might adversely worsen steatohepatitis, however, the effect of autophagic lipolysis, lipophagy, remains obscure. We engineered the adaptor protein to induce lipophagy with lipid droplet targeting signal and modified LC3 interacting region. Activating hepatocyte lipophagy obviously mitigated both steatosis and NASH pathology. Mechanistically, lipophagy promoted the excretion of lipid from liver via lysosomal exocytosis and attenuated harmful accumulation of nonesterified fatty acid. This exocytosis was dependent on Ca2+ signal unlike the lysosomal dysfunction-related exocytosis. High content compound screening identified alpelisib and digoxin, clinically-approved compounds, as effective activators of lipophagy. Administration of alpelisib or digoxin inhibited the transition to steatohepatitis in mice fed high fat with low methionine low choline diet. Given all these data, activating lipophagy may be a promising therapeutic approach to prevent NASH progression.

overall_design: Liver mRNA profiles of control and lipophagy(LIR) induced NASH model mice


title: Transcriptomic signatures reveal a shift towards an antiinflammatory gene expression profile but also the induction of type-I and type-II interferon signaling networks through aryl hydrocarbon receptor activation in murine macrophages

city: Leipzig

first: Jörg

last: Lehmann

address: Perlickstr. 1

department: Preclinical Development and Validation

institute: Fraunhofer Institute for Cell Therapy and Immunology

summary:
The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that regulates a broad range of target genes involved in the xenobiotic response, cell cycle control and circadian rhythm. AhR is constitutively expressed in macrophages, acting as key regulator of cytokine production. While proinflammatory cytokines, i.e., IL-1β, IL-6, IL-12, are suppressed through AhR activation, anti-inflammatory IL-10 is induced. However, the underlying mechanisms of those effects and the importance of the specific ligand structure are not yet completely understood.
Therefore, we have compared the global gene expression pattern in activated murine bone marrow-derived macrophages (BMMs) subsequently to exposure with either benzo[a]pyrene (BaP) or indole-3-carbinol (I3C), representing xenobiotic vs. nutritional AhR ligands, respectively, by means of mRNA sequencing. AhR dependency of observed effects was proved using BMMs from AhR-knockout mice. In total, more than 1,000 differentially expressed genes (DEGs) could be mapped, covering a plethora of AhR-modulated effects on basal cellular processes, i.e., transcription and translation, but also immune functions, i.e., antigen presentation, cytokine production, and phagocytosis. Among DEGs were genes that are already known to be regulated by AhR, i.e., Irf1, Ido2, and Cd84. However, we identified DEGs not yet described to be AhR-regulated in macrophages so far, i.e., Slpi, Il12rb1, and Il21r. All six genes likely contribute to shifting the macrophage phenotype from proinflammatory to anti-inflammatory. The majority of DEGs were specific for BaP but not affected through I3C exposure, probably due to higher AhR affinity of BaP in comparison to I3C.
Mapping of known aryl hydrocarbon response element (AHRE) sequence motifs in identified DEGs revealed more than 200 genes not possessing any AHRE, and therefore being not eligible for canonical regulation. Bioinformatic approaches modeled a central role of type I and type II interferons in the regulation of those genes. Additionally, RT-qPCR and ELISA confirmed a BaP- and AhR-dependent expressional induction and AhR-dependent secretion of IFN-γ, suggesting a potentially novel auto- or paracrine activation pathway of macrophages.

overall_design: RNA expression of each four replicates of murine bone marrow-derived macrophages (wild-type and AhR-knockout) under exposure of BaP, I3C or DMSO vehicle control in PAMP-induced PRR activation

first: Johannes

last: Schmidt

first: Janine

last: Haupt

first: Sina

last: Riemschneider

first: Christoph

last: Kämpf

first: Dennis

last: Löffler

first: Conny

last: Blumert

first: Kristin

last: Reiche

first: Ulrike

last: Koehl

first: Stefan

last: Kalkhof


title: Temporal transcriptional control of neural induction in human induced pluripotent stem cells.

first: Shankar

last: Subramaniam

address: 9500 Gilman Dr

institute: University of California, San Diego

summary: Neural induction of human induced pluripotent stem cells represents a critical switch in cell state during which pluripotency is lost and commitment to a neural lineage is initiated. Although many of the key transcription factors involved in neural induction are known, we know little of the temporal and causal relationships that are required for this state transition. Here, we have carried out a longitudinal analysis of the transcriptome of human iPSCs undergoing neural induction. Using the temporal relationships between the changing profile of key transcription factors and subsequent changes in their target gene expression profiles, we have identified distinct functional modules operative throughout neural induction. In addition to modules that govern loss of pluripotency and gain of neural ectoderm identity, we discover other modules governing cell cycle and metabolism Strikingly, some of these functional modules are retained throughout neural induction, even though the gene membership of the module changes. Systems analysis identifies other modules associated with cell fate commitment, genome integrity, stress response and lineage specification. We then focussed on OTX2, one of the most precociously activated transcription factors during neural induction. Our temporal analysis of OTX2 target gene expression identified several OTX2 regulated gene modules representing protein remodelling, RNA splicing and RNA processing. Further CRISPRi inhibition of OTX2 prior to neural induction promotes an accelerated loss of pluripotency and a precocious and aberrant neural induction disrupting some of the previously identified modules. We infer that OTX2 has a diverse role during neural induction and regulates many of the biological processes that are required for loss of pluripotency and gain of neural identity. This dynamical analysis of transcriptional changes provides a unique perspective of the widespread remodelling of the cell machinery that occurs during neural induction of human iPSCs.

overall_design: iPSC cell line was treated with dual SMAD inhibitors, 100 nM LDN193189 (Sigma Aldrich) and 10 µM SB431542 (Sigma Aldrich), to induce the neural induction. Samples were collected to generate 0 – 48 hours time series containing 8 time points, 0, 2, 4, 8, 18, 24, 30 and 48 hour and 0 - 8 days time series containing 7 time points, 0, 1, 2, 3, 4, 6 and 8 day. Further, we used CRISPRi to knock down the OTX2 expression and Samples were collected at time points of 0, 1, 2 and 7 days following initiation of neural induction.

first: Shakti

last: Gupta

first: Noel

middle: J

last: Buckley


title: Gene expression profile at single cell level of human Simpson Golabi Behmel Syndrome (SGBS) and murine 3T3-L1 cells prior to and during adipogenesis

city: Palo Alto

first: Christopher

middle: Ann

last: Jin

address: 3165 Porter Drive

department: Genetics

institute: Stanford Medical School

summary: We used single cell RNA sequencing (scRNA-seq) to analyze the transcriptional diversity of cells undergoing adipogenesis using two cellular models of adipogenesis in vitro, human SGBS cells and murine 3T3-L1 cells.

overall_design: SGBS and 3T3-L1 cells were isolated at two timepoints: prior to the addition of differentiation stimuli (D0) and during adipogenesis (D5 for 3T3-L1, D8 for SGBS). Live cells were isolated using Fluorescence-activated cell sorting (FACS) according to the absence of staining with propidium iodide. Next, equal numbers of SGBS and 3T3-L1 cells were mixed and analyzed using scRNA-Seq.

first: Joshua

last: Knowles

first: Ewa

last: Bielczyk-Maczynska

first: Jiehan

last: Li


title: Palmitoyl acyltransferase ZDHHC7 suppresses prostate cancer through inhibiting androgen receptor expression

city: Atlanta

first: Jindan

last: Yu

state: GA

address: E330, 1760 Haygood Dr NE

department: Urology

institute: Emory University

summary: We examined ZDHHC family gene expression in human tissue collection and identified ZDHHC7 as a PCa-relevant member. RNA-seq analyses of PCa cells with ZDHHC7 de-regulation revealed global alterations in androgen response and cell cycle pathways. Mechanistically, ZDHHC7 inhibits AR gene transcription and therefore reduces AR protein levels and abolishes AR signaling in PCa cells. Accordingly, ZDHHC7 depletion increased oncogenic properties of PCa cells, whereas restoring ZDHHC7 is sufficient to suppress PCa cell proliferation and invasion in vitro and mitigate xenograft tumor growth in vivo. Lastly, we demonstrated that ZDHHC7 is down-regulated in human PCa compared to benign adjacent tissues and its loss is associated with worse clinical outcomes. Taken together, our findings unequivocally prove ZDHHC7 as a tumor suppressor gene that inhibits AR signaling and PCa progression, identifying ZDHHC7 loss as a biomarker for aggressive PCa and a target for therapeutic intervention.

overall_design: RNA-seq of prostate cancer cells with ZDHHC7 overexpression or knockdown

first: Zhuoyuan

last: Lin


title: Transctiptome analysis of Dsup-expressing and control D. melanogaster lines in non-stressed conditions and after ionizing radiation treatment

city: Dubna

first: Mikhail

last: Zarubin

address: Russia, Moscow region, Dubna, Juliot Curie, 20

country: Russia

institute: Joint Institute for Nuclear Research

summary: We used microarrays to make transcriptome profiling to reveal mechanism of radiotolerance enhancement in D. melanogaster

overall_design: 3-day-old adult male flies were taken for RNA isolation from 4 control repeats and 4 transgenic lines of D. melanogaster male organisms. Next the same procedure was performed after treatment with ionizing radiation of 1000 Gy dose for 3 control repeats and 4 transgenic lines of D. melanogaster.

first: Elena

last: Kravchenko


title: pre-mRNA splicing order is defined and maintains splicing fidelity across multi-intronic transcripts [II]

first: Karine

last: Choquet

summary: Combinatorially, intron excision within a given nascent transcript could proceed down any of thousands of paths, each of which would expose different dynamic landscapes of cis-elements and contribute to alternative splicing. In this study, we found that post-transcriptional multi-intron splicing order in human cells is largely predetermined, with most genes spliced in one or a few predominant orders. Strikingly, these orders were conserved across cell types and stages of motor neuron differentiation. Introns flanking alternatively spliced exons were frequently excised last, after their neighboring introns. Perturbations to the spliceosomal U2 snRNA altered the preferred splicing order of many genes, and these alterations were associated with the retention of other introns in the same transcript. In one gene, early removal of specific introns was sufficient to induce delayed excision of three proximal introns, and this delay was caused by two distinct cis-regulatory mechanisms. Together, our results demonstrate that multi-intron splicing order in human cells is predetermined, is influenced by a component of the spliceosome and ensures splicing fidelity across long pre-mRNAs.

overall_design: Direct RNA and targeted cDNA-PCR nanopore sequencing in K562 cells, HeLa cells, differentiating motor neurons, myoblasts and upon U2 snRNA knockdown

first: Autum

last: Koenigs

first: Sarah-Luisa

last: Dülk

first: Brendan

last: Smalec

first: Silvi

last: Rouskin

first: L

middle: S

last: Churchman


title: pre-mRNA splicing order is defined and maintains splicing fidelity across multi-intronic transcripts [I]

overall_design: RNA-seq during human motor neuron differentiation, RNA-seq and NET-seq upon U2 snRNA knockdown; in vitro DMS-MaPseq for WT and mutated IFRD2 constructs


title: Dietary protein shapes the profile and repertoire of intestinal CD4+ T cells

status: Public on May 13 2023

city: New York

first: Tiago

last: Castro

state: New York

address: 1230 York Avenue

institute: The Rockefeller University

summary: This study shows in a polyclonal setting that routine exposure to food protein promotes accumulation of antigen-experienced CD4+ T cells in the intestinal epithelium, which acquire a cytotoxic tissue-resident transcriptional program. This process is disrupted upon either allergic sensitization or tolerance.

overall_design: Single CD4+ T cells from the small intestine epithelium or lamina propria of mice with different dietary and microbial manipulations; libraries are the antibody derived tags, gene expression and tcr sequencing.

first: Ainsley

last: Lockhart

first: Aubrey

last: Reed

first: Calvin

last: Herman

first: Maria

middle: C

last: Canesso

first: Daniel

last: Mucida


title: Aging-induced pseudouridine synthase 10 impairs hematopoietic stem cells

first: zhenzhen

last: zhang

state: Haidian Distinct

address: No.30 Shuangqing Road

department: School of Medicine

institute: Tsinghua University

summary: Aged hematopoietic stem cells (HSCs) exhibit compromised reconstitution capacity and differentiation-bias towards myeloid lineage. However, the molecular mechanism behind it remains not fully understood. In this study, we evaluated the expression of 5 pseudouridine (Ψ) synthases between young and aged HSCs, and observed that three of them are increased during aging. Functional evaluation assay reveals that enforced PUS10 and PUS7 recapitulate the phenotype aged HSCs. The increase of PUS10 in aged HSCs is due to aging-declined CRL4DCAF1-mediated ubiquitination degradation signaling. Mechanistically, the destructive role of enforced PUS10 on HSCs is not achieved by its Ψ synthases activity, but through miR139, dysfunction of which impairs the reconstitution capacity of HSCs. Consistently, we observed no difference of tRNA pseudouridylation profile between young and aged HSPCs, while enforced PUS10 alters the microRNA profile in HSCs. Targeted dysfunction of Pus10 results in compromises the self-renewal capacity of HSC.

overall_design:
Comparative gene expression profiling analysis of RNA-seq data for Vector and PUS10-OE HSC.
Comparative gene expression profiling analysis of RNA-seq data for NTC(non-target control) and miR139-KD HSC.
Comparative gene expression profiling analysis of RNA-seq data for Pus10-WT and Pus10-KO HSC.

first: Yuqian

last: Wang

first: Zhenzhen

last: Zhang

first: Jianwei


overall_design: Comparative microRNA expression profiling analysis of RNA-seq data for Vector and PUS10-OE HSCs.


title: Chemokine receptor CXCR7 activates Aurora Kinase A and promotes neuroendocrine prostate cancer growth

first: Jindan

last: Yu

summary: CXCR7, an atypical chemokine receptor (ACKR3), is up-regulated in NEPC, and its expression is associated with molecular markers of NEPC and cell proliferation. Transcriptomic analyses revealed a major role of CXCR7 in regulating downstream genes involved in the cell cycle and mitotic spindle. Membrane-bound CXCR7 is known to recruit β-arrestin (ARRB2), and the complex internalizes into clathrin-coated vesicles where they act as a scaffold for cytoplasmic kinase assembly and substrate activation. We identify Aurora Kinase A (AURKA) as a top candidate that is binding to and activated by CXCR7-ARRB2. Immunofluorescence confocal microscopy detected high-density CXCR7, ARRB2, and AURKA in the pericentrosomal area with focal staining of ARRB2 and ARUKA at centrosomes. Mass spectrometry showed that CXCR7 interacts with many proteins associated with intracellular vesicles, microtubules, and Golgi apparatus. CXCR7-ARRB2-containing vesicles traffic along the microtubules to the perinuclear Golgi apparatus, where CXCR7-ARRB2 interacts with AURKA to promote its activation and cell growth. Finally, we showed that pharmacological inhibitors of AURKA abolish CXCR7-driven tumor growth. Our study reveals CXCR7-mediated activation of AURKA and establishes CXCR7 and its downstream kinases as critical targets for therapeutic intervention in CRPC or NEPC patients.

overall_design: Gene expression profiling from LNCaP MDVr and C4-2B MDVr cells were performed by RNA-seq. There are 2 treatment groups in triplicate with pLKO as a control/reference group.

first: Galina

last: Gritsina


overall_design: Comparative tRNA pseudouridylation  profiling analysis of DM-Ψ-seq data for young and aged HSPCs.


summary: Aged hematopoietic stem cells (HSCs) exhibit compromised reconstitution capacity and differentiation-bias towards myeloid lineage. While, the molecular mechanism behind it remains not fully understood. In this study, we observed that the expression of pseudouridine (Ψ) synthase 10 is increased in aged hematopoietic stem and progenitor cells (HSPCs) and enforced PUS10 recapitulates the phenotype of aged HSCs, which is not achieved by its Ψ synthase activity. Consistently, we observed no difference of tRNA pseudouridylation profile between young and aged HSPCs. No significant alteration of hematopoietic homeostasis and HSC function is observed in young Pus10-/- mice, while aged Pus10-/-mice exhibit mild alteration of hematopoietic homeostasis and HSC function. Moreover, we observed that PUS10 is ubiquitinated by E3 ubiquitin ligase CRL4DCAF1 complex and the increase of PUS10 in aged HSPCs is due to aging-declined CRL4DCAF1-mediated ubiquitination degradation signaling. Taken together, this study for the first time evaluated the role of PUS10 in HSC aging and function, and provided novel insight for HSC rejuvenation and clinical application.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for NTC(non-target control) and miR652-KD HSC.


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

overall_design: Refer to individual Series


title: S1PR2 Signaling in Chronic Glucocorticoid Exposure-Induced Metabolic Disorders

city: Berkeley

first: Jen-Chywan

address: 315 Morgan Hall, UC Berkeley

department: Nutritional Sciences & Toxicology

institute: UC Berkeley

summary: Glucocorticoids play a key role in metabolic adaptation during stress, such as fasting and starvation. Excess and/or chronic glucocorticoid exposure, however, cause metabolic disorders that include insulin resistance dyslipidemia and hepatic steatosis. We previously showed that chronic glucocorticoid treatment elevates hepatic production of ceramides and sphingosine-1-phosphate (S1P). Ceramides are converted to sphingosine that is further converted to S1P. We showed that ceramide-initiated signaling in turn inhibits insulin signaling and increases lipogenic program in hepatocytes. However, the role of S1P, a signaling molecule that modulates physiological responses through membrane S1P receptors (S1PRs), in glucocorticoid actions is unclear. Here we found that inhibiting S1PR2 activity, but not S1PR1 and S1PR3, resulted in improved glucocorticoid-induced insulin resistance. Similarly, reducing hepatic S1PR2 expression showed similar results. Interestingly, reducing S1PR2 expression did not affect the ability of glucocorticoids to modify insulin signaling. Instead, glucocorticoids enhanced gluconeogenesis was reduced. Moreover, glucocorticoid-induced dyslipidemia and fatty liver was also compromised in mice with reduced hepatic S1PR2 expression. Indeed, RNA sequencing analysis showed that lipogenic, gluconeogenic and glycolytic genes are significantly lower in glucocorticoid-treated hepatic S1PR2 knockdown mice than those of glucocorticoid-treated hepatic scramble small hairpin RNA (shRNA) expressing mice (Control). Overall, our studies highlight the importance role of S1PR2 signaling in the mediating glucocorticoid regulation on gluconeogenesis and hepatic lipid homeostasis.

overall_design: WT mice were provided by the Andras Nagy laboratory (Samuel Lunenfeld Research Institute, Mount Sinai Hospital) and the Jeff Gordon laboratory (Washington University) Mice received 3x1011GC per mouse of AAV8 expressing small hairpin RNA for scramble or S1PR2 (Vector Biolabs, Melvern, PA) diluted with PBS. AAV was injected intravenously though the tail vein. Male 7-12 week old mice were treated with approximately 0.84mg/kg of dexamethasone for 7 days in their drinking water which was prepared by dissolving dexamethasone sodium phosphate (Sigma, Cleveland, OH, PHR1768) in drinking water

first: Rebecca

middle: A

last: Lee

first: Ariel

last: Tsay

first: Danielle

last: Li

first: Maggie

last: Chang

first: Nicholas

last: Yiv

first: Jenna

last: Tan

first: Sonia

last: Scallon

first: Justin


title: Unique Transcriptional Profiles Underlie Osteosarcomagenesis Driven by Different p53 Mutants (ATAC-Seq)

status: Public on May 12 2023

city: Houston

first: Xiaoping

last: Su

state: Texas

address: 7300 Brompton St, Apt 5633

institute: MD Anderson Cancer Center

summary: Open chromatin regions in the murine osteosarcoma cell lines expressing different mutants of p53 were identified using ATAC-seq

overall_design: Paired end ATAC Seq

first: Dhruv

last: Chachad


title: Unique Transcriptional Profiles Underlie Osteosarcomagenesis Driven by Different p53 Mutants (ChIP-Seq)

summary: To identify mutant p53 GOF and analyze mutant p53 ChIP peaks ChIP-seq was performed on hESC-derived human osteoblasts expressing homozygous WT p53 or heterozygous expression of p53R248W. Processing for ChIP was performed on day 24 after confirmation of osteoblast differentiation

overall_design: ChIP-Seq

first: Dung

middle: f

first: An

last: Xu

first: Lalit

last: Patel


title: Unique Transcriptional Profiles Underlie Osteosarcomagenesis Driven by Different p53 Mutants (RNA-Seq)

summary: To identify mutant p53 GOF, murine primary osteosarcomas expressing p53R172H or p53R245W over null and p53-null osteosarcomas were processed for bulk sequencing; DEGs were identified in p53R172H and p53R245W expressing tumors by comparing to p53-null tumors; DEGs were used to identify dysregulated pathways and mutant p53 GOF

overall_design: Paired-end RNA-seq of primary murine osteosarcomas expressing different mutants of p53


title: RNAseq analysis of Vibrio cholerae A1552 WT and ∆rvvA (∆VCA0257)

city: Santa Cruz

first: Giordan

last: Kitts

address: 1156 High St.

department: METX

institute: UCSC

summary: RNA-sequencing of Vibrio cholerae A1552 wild-type and ∆VCA0257 (∆rvvA) strains grown under virulence-inducing conditions (AKI).

overall_design: Three biological replicates of wild-type V. cholerae A1552 smooth wild-type and ∆rvvA strains were grown to exponential phase in virulence-inducing conditions (AKI), and samples were collected for RNA isolation

first: Fitnat

middle: H

last: Yildiz


title: TREM2 inhibition triggers anti-tumor cell activity of myeloid cells in glioblastoma

first: Albert

last: Kim

address: 660 S. Euclid Avenue

department: Department of Neurological Surgery

institute: Washington University School of Medicine


title: Differential effects of glutamine inhibition strategies on anti-tumor CD8 T cell function

city: Nashville

first: Jeff

last: Rathmell

state: TN

address: 1211 Medical Center Drive

department: PMI

institute: Vanderbilt University Medical Center

summary: Activated T cells undergo metabolic reprogramming to meet anabolic, differentiation, and functional demands. Glutamine supports many processes in activated T cells, and inhibition of glutamine metabolism alters T cell function in autoimmune disease and cancer. Multiple glutamine-targeting molecules are under investigation, yet the precise mechanisms of glutamine-dependent CD8 T cell differentiation remain unclear. We show that distinct strategies of glutamine inhibition by glutaminase-specific inhibition with small molecule CB-839, pan-glutamine inhibition with 6-diazo-5-oxo-L-norleucine (DON), or by glutamine deprivation (No Q) produce distinct metabolic differentiation trajectories in CD8 T cells. T cell activation with CB-839 treatment had a milder effect than DON or No Q treatment. A key difference was that CB-839-treated cells compensated with increased glycolytic metabolism, while DON and No Q-treated cells increased oxidative metabolism. However, all glutamine treatment strategies elevated CD8 T cell dependence on glucose metabolism, and No Q treatment caused adaptation toward reduced glutamine dependence. DON treatment reduced histone modifications and numbers of memory-like cells in adoptive transfer studies, but those T cells that persisted could expand normally upon secondary antigen encounter. In contrast, No Q-treated cells persisted well yet demonstrated decreased secondary expansion. Consistent with impaired function, CD8 T cells activated in the presence of DON had reduced ability to control tumor growth and reduced tumor infiltration in adoptive cell therapy. Overall, each approach to inhibit glutamine metabolism confers distinct effects on CD8 T cells and highlights that targeting the same pathway in different ways can elicit opposing metabolic and functional outcomes.

overall_design: Uinsg RNAseq and CUT&RUN to profile how glutamine metabolism reprogram CD8+ T cell function.

first: Madden

last: Matthew Z

first: Xiang

last: Ye

first: Channing

last: Chi

first: Jeffrey


title: Single cell RNA sequencing (scRNAseq) of stromal cells from mouse colon assembloids and epithelial cells from colon assembloids, organoids, and tissue

city: Berlin

first: Michael

last: Sigal

address: Augustenburger Platz 1

department: Dept. Hepatology and Gastroenterology

institute: Charité Universitätsmedizin

summary: We established a colon assembloid system comprising epithelium and diverse subtypes of stromal cells. To explore how closely assembloids resemble the native tissue, we performed the transcriptional profiling of stromal cells from assembloids and epithelial cells from assembloids, organoids, and colon tissue at the single-cell level.

overall_design: Live stromal cells from assembloids and epithelial cells from assembloids, organoids, and colon tissue were isolated by Fluorescence-activated cell sorting (FACS) and analyzed using scRNAseq.

first: Hilmar

last: Berger

first: Manqiang

last: Lin

first: Thomas

last: Conrad

first: Hao


title: M1BP RNAi during Drosophila flight myogenesis: Analyses at 2-day adult stage

city: Marseille

first: Andrew

last: Saurin

address: Campus de Luminy

country: France

institute: IBDM / CNRS

summary: Oxidative metabolism is the predominant energy source for muscle contraction. How this is transcriptionally regulated during muscle development to accommodate different metabolic requirements and muscle types is largely unknown. We show that the formation of mitochondria cristae harbouring the respiratory chain is concomitant with a large-scale transcriptional upregulation of genes linked with oxidative phosphorylation (OXPHOS) during the middle of Drosophila flight muscle development. We further demonstrate using high-resolution imaging, transcriptomic and biochemical analyses that Motif-1-binding protein (M1BP) transcriptionally regulates the expression of genes encoding critical components for OXPHOS complex assembly and integrity. In the absence of M1BP, mitochondrial respiratory complexes are improperly assembled and aggregate in the mitochondrial matrix, triggering a strong protein quality control response. Together, this study provides the first insight into the transcriptional regulation of oxidative metabolism during Drosophila myogenesis and identifies M1BP as a novel player in this process.

overall_design: Total RNA samples from 30x 2-day adult flight muscles (hand dissected) from  Mef2-Gal4 drosophila crosses to WT (w1888) or M1BP RNAi lines. All samples are performed in triplicate with approximate concentration of 50ng/ul (49ul per sample)

first: Gabreila

last: Poliacikova

first: Marine

last: Barthez

last: Rival

first: Aïcha

last: Aouane

first: Nuno

last: Luis

first: Fabrice

last: Richard

first: Nicolas

last: Brouilly

first: Frank

last: Schnorrer

first: Corine

last: Maurel-Zaffran

first: Yacine

last: Graba


title: TREM2 inhibition triggers anti-tumor cell activity of myeloid cells in glioblastoma [RNAseq]

first: Albert

last: Kim

summary: The prevailing view is that myeloid cells in the tumor microenvironment (TME) are immunosuppressive and promote glioblastoma (GBM) progression. However, myeloid cells have the functional plasticity to restrict or support tumor cell growth. TREM2 plays important roles in brain microglial function in neurodegenerative diseases, but the role of TREM2 in the GBM TME has not been examined. We found TREM2 is highly expressed in myeloid subsets, including macrophages and microglia in human and mouse GBM tumors and that high TREM2 expression correlates with poor prognosis in GBM patients. TREM2 loss of function in human macrophages and mouse myeloid cells increased tumoricidal capacity. TREM2 in myeloid cells restricts IFNγ-induced immunoactivation and proinflammatory polarization. In orthotopic mouse GBM models, Trem2-/- mice and mice with acute brain Trem2 reduction demonstrate survival benefit. Trem2 inhibition reprograms myeloid phenotypes and increases PD-1+CD8+ T cells in the TME. Trem2 deficiency enhances the effectiveness of anti-PD-1 treatment and may represent a therapeutic strategy for GBM patients.

overall_design: Bone marrow-derived macrophages (BMDMs) were isolated and differentiated from trem2 transgenic mice, Trem2+/+ (n=2), Trem2-/- (n=2). Cells were treated with PBS (n=2), 10 ng/mL IFNγ (n=2), 100 ng/mL LPS (n=2), or dead SB28 tumor cells (n=2) for 6 hours. Total RNA from each sample/genotype was isolated and sequenced. Primary microglia were enriched from brain cortex of postnatal day 2 Trem2+/+ mouse pups (n=12). Primary microglia were treated with Trem2-lowering antisense oligonucleotide (ASO) or inactive ASO for 72 hours. Trem2 ASO (n=2), inactive (n=2). Cells were then treated with PBS (n=2),10 ng/mL IFNγ (n=2) or dead SB28 tumor cells (n=2) for 6 hours. Total RNA from each sample/treatment was isolated and sequenced. Trem2+/+ mice were intracerebroventricularly injected with 400 μg (10ul) Trem2 ASO (n=5) or inactive ASO (n=5). 7 days later, all mice were intracerebrally injected with SB28 cells. 30 days later, SB28 glioblastoma were dissected from ASO-treated mice brain and dissociated into single-cell suspensions for RNA-seq. Trem2 ASO (n=2), inactive ASO (n=2).

first: Jason

middle: D

last: Ulrich

first: Rui

last: Sun


title: PolyA RNA-seq from liposarcoma cells treated with Nutlin3R and/or Carfilzomib

city: Aurora

first: Matthew

last: Galbraith

state: CO

department: Department of Pharmacology; Linda Crnic Institute for Down Syndrome

institute: University of Colorado Anschutz Medical Campus

summary: Analysis of steady-state mRNA levels in liposarcoma cells treated for 28 hr with 0.05% DMSO or 10 uM Nutlin3R and/or 12.5 nM Carfilzomib

overall_design: Two independent biological replicates each of 94T778 and Lipo-246 cells, treated for 28 hr with 0.05% DMSO or 10 uM Nutlin3R and/or 12.5 nM Carfilzomib

first: Michael

middle: P

last: Ludwig

first: Kelly

last: Sullivan

first: Joaquin

middle: M

last: Espinosa


title: Single cell RNA sequencing of mouse indced pluripotent stem cell (iPSC)-derived and mouse primary embryonic lung mesenchyme

city: Boston

first: Pushpinder

last: Bawa

state: Massachusetts

address: 670 Albany St

department: Boston University Medical Campus

institute: Center for Regenerative Medicine

summary: The lung mesenchyme plays important roles in lung development and is affected in many respiratory diseases, yet relatively little is known about the biology of lung mesenchymal progenitors. We sought to establish an induced pluripotent stem cell (iPSC)-based model to study lung mesenchyme development and epithelial-mesenchymal interactions. We generated a mouse iPSC line carrying a lung mesenchyme-specific reporter/tracer to establish a protocol for differentiation into lung mesenchymal progenitors. We derived lung mesenchyme from iPSCs both by directed differentiation via a lateral plate mesodermal progenitor state (induced lung mesenchyme, iLM), and by co-development during lung epithelial differentiation (co-developed lung mesenchyme, cLM). We found that directed differentiation via a lateral plate mesoderm progenitor was not only more efficient, but also yielded engineered lung mesenchymal cells that were more similar to primary lung mesenchyme from day 12.5 mouse embryos, as determined by single cell RNAseq. Our iPSC-derived population will provide an inexhaustible source of cells for studying lung development, modeling diseases, and developing therapeutics.

overall_design: Global transcriptomic profiling at single cell resolution of iPSC-derived cells and primary embryonic lung cells. 3 samples contain iPSC-derived cells : 1) "iLM": Tbx4-LERGFP+ cells (i.e. lung mesenchyme) generated using directed lung mesenchymal differentiation; 2) "iLMneg": Tbx4-LERGFP- cells (i.e. non-lung mesenchyme) generated using directed lung mesenchymal differentiation; 3) "cLM": Tbx4-LERGFP+ cells (i.e. lung mesenchyme) and Epcam+/Tbx4-LERGFP- (i.e. epithelial) cells "co-developed" by directed lung epithelial differentiation to simultaneously produce both endoderm/epithelia and mesenchyme. A 4th sample contains primary E12.5 lung mesenchyme  and epithelium.

first: Andrea

last: Alber

first: Jonathan

last: Lindstrom-Vautrin

first: Carlos

last: Villacorta-Martin

first: Darrell

middle: N

last: Kotton


title: Human IRF1 governs macrophagic IFN-γ immunity to mycobacteria

city: Paris

first: Jeremy

last: Manry

address: 24 boulevard du Montparnasse

institute: INSERM, Institut Imagine

summary: Inborn errors of human IFN-γ-dependent macrophagic immunity underlie mycobacterial diseases, whereas inborn errors of IFN-α/β-dependent intrinsic immunity underlie viral diseases. Both types of IFNs induce the transcription factor IRF1. We describe unrelated children with inherited complete IRF1 deficiency and early-onset, multiple, life-threatening diseases caused by weakly virulent mycobacteria and related intramacrophagic pathogens. These children have no history of severe viral disease, despite exposure to many viruses, including SARS-CoV-2, which is life-threatening in individuals with impaired IFN-α/β immunity. In leukocytes or fibroblasts stimulated in vitro, IRF1-dependent responses to IFN-γ are, both quantitatively and qualitatively, much stronger than those to IFN-α/β. Moreover, IRF1-deficient mononuclear phagocytes do not control mycobacteria and related pathogens normally when stimulated with IFN-γ. By contrast, IFN-α/β-dependent intrinsic immunity to nine viruses, including SARS-CoV-2, is almost normal in IRF1-deficient fibroblasts. Human IRF1 is essential for IFN-γ-dependent macrophagic immunity to mycobacteria, but largely redundant for IFN-α/β-dependent antiviral immunity.

overall_design: Total RNA profiles from primary fibroblasts of the two IRF1 patients, a previously validated IFNGR1 patient, a previously validated IFNGR2 patient, a previously validated IFNAR1 patient, a previously validated IRF9 patient, a previously validated STAT1 patient and three healthy controls. Total RNA profiles from iPSC-derived macrophages of one IRF1 patient, a previously validated IFNGR1 patient, a previously validated STAT1 patient and two healthy controls. Total RNA profiles from monocyte-derived macrophages of one IRF1 patient, a brother of the IRF1 patient, and three healthy controls.

first: Jérémie

last: Rosain

first: Jacinta

last: Bustamante


title: M1BP is an essential transcriptional activator of oxidative metabolism during Drosophila development

first: Andrew

last: Saurin


title: M1BP RNAi during Drosophila flight myogenesis: Analyses at 48h APF stage

overall_design: Differential gene expression analyses of M1BP RNAi in dorsolongitudinal flight muscle in pupae at 48h APF

first: Gabreila

last: Poliacikova

first: Marine

last: Barthez

first: Thomas

last: Rival

first: Aïcha

last: Aouane

first: Nuno

last: Luis

first: Fabrice

last: Richard

first: Nicolas

last: Brouilly

first: Frank

last: Schnorrer

first: Corine

last: Maurel-Zaffran

first: Yacine

last: Graba


title: Unique Transcriptional Profiles Underlie Osteosarcomagenesis Driven by Different p53 Mutants

first: Xiaoping

last: Su


title: DNA methylation dynamics during esophageal epithelial regeneration following repair with acellular silk fibroin grafts in rat

city: Irvine

first: Timothy

last: Downing

address: 419 S. Circle View Drive

department: Department of Biomedical Engineering

institute: UCI

type: Methylation profiling by high throughput sequencing

summary: Bi-layer silk fibroin (BLSF) scaffolds represent an emerging technology in the development of acellular biomaterials for esophageal repair. Their therapeutic potential is attributed to their robust mechanical properties, elasticity, and low immunogenicity. However, the underlying molecular mechanisms of scaffold-mediated wound healing processes in the esophagus remain unclear. Previous research has identified signaling cascades involved in neoepithelial regeneration in a rat model of onlay esophagoplasty with BLSF grafts, including c-MET, TrkA, and PI3K-Akt signaling. Interestingly, these pro-survival signaling cascades are largely governed by DNA methylation (DNAme). Such findings motivate us to apply reduced-representation bisulfite sequencing (RRBS) to characterize the temporal dynamics of DNAme in host and regenerated tissues up to 1 week post-operation. Globally, we observe hypermethylation at post-surgical repair timepoints and an inverse correlation between DNAme and the expression levels of differentially expressed proteins during regeneration. Site-specific hypomethylation targets genes associated with immune activation, which may be indicative of immune cell infiltration at the surgical implant site, while site-specific hypermethylation appears to target PI3K-Akt signaling. Our work provides mechanistic insight into the molecular processes governing esophageal regeneration, which can motivate therapeutic innovations toward temporal modulation of tissue regeneration during scaffold-mediated esophageal repair via epigenetic regulators that target key pathways.

overall_design: Reduced-representation bisulfite sequencing (RRBS) with rat esophageal tissues in nonsurgical controls (NSC, n=4), and 1 day (n=5) or 1 week (n=5) after surgical repair with  Bi-layer silk fibroin (BLSF) scaffolds.

first: Gokhan

last: Gundogdu

first: Hanjuan

last: Shao

first: Joshua

last: Mauney

middle: L

first: Lauren

last: Urban

first: Annie

last: Trinh

first: Jiachun

last: Li

first: Travis

last: Nguyen


title: TREM2 inhibition triggers anti-tumor cell activity of myeloid cells in glioblastoma [scRNA]

city: St. Louis

first: Albert

last: Kim

first: Jason

last: Ulrich

first: Rui

last: Sun


title: Nr2f1a maintains atrial nkx2.5 expression to repress pacemaker identity within venous atrial cardiomyocytes of zebrafish

city: Cincinnati

last: Waxman

address: 240 Albert Sabin Way

department: Molecular Cardiovascular Biology Division

institute: Cincinnati Children's Hospital Medical Center

summary: Maintenance of cardiomyocyte identity is vital for normal heart development and function. However, our understanding of cardiomyocyte plasticity remains incomplete. Here, we show that sustained expression of the zebrafish transcription factor Nr2f1a prevents the progressive acquisition of ventricular cardiomyocyte (VC) and pacemaker cardiomyocyte (PC) identities within distinct regions of the atrium. Transcriptomic analysis of flow-sorted atrial cardiomyocytes (ACs) from nr2f1a mutant zebrafish embryos showed increased VC marker gene expression and altered expression of core PC regulatory genes, including decreased expression of nkx2.5, a critical repressor of PC differentiation. At the arterial (outflow) pole of the atrium in nr2f1a mutants, cardiomyocytes resolve to VC identity within the expanded atrioventricular canal. However, at the venous (inflow) pole of the atrium, there is a progressive wave of AC transdifferentiation into PCs across the atrium toward the arterial pole. Restoring Nkx2.5 is sufficient to repress PC marker identity in nr2f1a mutant atria and analysis of chromatin accessibility identified a Nr2f1a-dependent nkx2.5 enhancer expressed in the atrial myocardium directly adjacent to PCs. CRISPR/Cas9-mediated deletion of the putative nkx2.5 enhancer leads to a loss of Nkx2.5-expressing ACs and expansion of a PC reporter, supporting that Nr2f1a limits PC differentiation within venous ACs via maintaining nkx2.5 expression. The Nr2f-dependent maintenance of AC identity within discrete atrial compartments may provide insights into the molecular etiology of concurrent structural congenital heart defects and associated arrhythmias.

overall_design: RNA-seq and ATAC-seq of flow sorted amhc:EGFP+ cells

first: Kendall

middle: E

last: Martin

first: Padmapriyadarshini

last: Ravisankar

first: Manu

last: Beerens

first: Calum

last: MacRae


title: M1BP RNAi during Drosophila flight myogenesis: Analyses at 64h APF stage

first: Andrew

last: Saurin

overall_design: Differential gene expression analyses of M1BP RNAi in dorsolongitudinal flight muscle in pupae at 64h APF

first: Gabreila

last: Poliacikova

first: Marine

last: Barthez

first: Thomas

last: Rival

first: Aïcha

last: Aouane

first: Nuno

last: Luis

first: Fabrice

last: Richard

first: Nicolas

last: Brouilly

first: Frank

last: Schnorrer

first: Corine

last: Maurel-Zaffran

first: Yacine

last: Graba


title: Mutation of Vsx genes in zebrafish highlights the robustness of the retinal specification network [RNA-Seq]

city: Napoli

first: Lorena

last: Buono

address: via Emanuele Gianturco 32

institute: IRCSS-SDN

summary: Genetic studies in human and mice have established a dual role for Vsx genes in retina development: an early function in progenitors’ specification, and a later requirement for bipolar-cells fate determination. Despite their conserved expression patterns, it is currently unclear to which extent Vsx functions are also conserved across vertebrates, as mutant models are available only in mammals. To gain insight into vsx function in teleosts, we have generated vsx1 and vsx2 CRISPR-Cas9 double knockouts (vsxKO) in zebrafish. Our electrophysiological and histological analyses indicate severe visual impairment and bipolar cells depletion in vsxKO larvae, with retinal precursors being rerouted towards photoreceptors or Müller glia fates. Surprisingly, neural retina is properly specified and maintained in mutant embryos, which do not display microphthalmia. We show that although important cis-regulatory remodelling occurs in vsxKO retinas during early specification, this has little impact at a transcriptomic level. Our observations point to genetic redundancy as an important mechanism sustaining the integrity of the retinal specification network, and to Vsx genes regulatory weight varying substantially among vertebrate species.

overall_design: Head of zebrafish embryos at 18 hpf were dissected and used for RNA extraction. Tails were used for genotyping.

first: Joaquin

last: Letelier


title: Mutation of Vsx genes in zebrafish highlights the robustness of the retinal specification network [ATAC-Seq]

overall_design: Head of zebrafish embryos at 18 hpf were dissected and used for Tn5 tagmentation. Tails were used for genotyping.


title: Analysis of circRNA expression in human neuronal differentiation.

status: Public on May 11 2023

city: Stockholm

first: Michelle

last: Watts

address: Solnavägen 30

country: Sweden

institute: Karolinska Institutet

summary:
Expression of circular RNA (circRNA) was profiled during neuronal differentiation of human neuroepithelial stem (NES) cells.
From the same data, analysis of linear RNA expression was also performed.

overall_design:
RNA expression profiling analysis of total RNA-seq data for NES cells and NES cells differentiated for five days (D5) or 28 days (D28).  ***The submitters are not legally permitted to submit the raw data to GEO or a restricted-access database such as dbGaP or EGA. Therefore, raw data is not provided.
---------------------------
Authors state:
As this submission relates to sequencing data of human samples, raw data may contain potentially identifiable information. We therefore can not make raw data available either through the GEO or a restricted-access database such as dbGaP or EGA. The raw data is therefore excluded from this submission.

first: Marika

last: Oksanen

first: Sanna

last: Lejerkrans

first: Francesca

last: Mastropasqua

first: Myriam

last: Gorospe

first: Kristiina

last: Tammimies


title: Effect of ROS (H2O2) on undifferentiated NSC

city: Solna

first: susanne

last: neumann

address: Solnavägen 1

institute: Karolinska Instituet

summary: Release of reactive oxygen species, such as H2O2, has been observed during inflammatory processes but is also a physiological side product of metabolism. We have observed that differentiation of neural progenitor cells (NPC) is affected by H2O2 exposure. We here assess the transcriptional drivers of the phenotypical changes NPCs undergo during differentiation in response to H2O2. Here we publish only the untreated controls.

overall_design: 2 timepoints (1h or 8h on PDL coated plates) with biological triplicates per timepoint, except 1h where biological duplicates were used.

first: Susanne

last: Neumann

first: Caroline

last: Lindblad

first: Ruxandra

last: Covacu

first: Lou

last: Brundin


title: RNA secondary structome of different retrotransposons with DNA/protein binding

city: Qingdao

first: Lei

last: Sun

state: Shandong

address: Shandong street

institute: Shandong University

summary: We determined the RNA secondary structure of different retrotransposon by icSHAPE. The retrotransposon has differnet states including protein bound, DNA and protein bound.

overall_design: RNA was prepared as in the cryo-EM studies in three conditions: apo RNA, RNA-protein complex, and RNA-DNA-protein ternary complex. In each condition, the RNA sequence has three types: 5’ORF, 3UTR, and 5’ORF-linker-3’UTR chimeric. We design reverse transcription primers for the nine RNA samples with specific barcodes. Then, all samples were performed by icSHAPE.

first: Qiangfeng

last: Zhang


title: A transcriptomic study reveals beneficial effects of HsTX1[R14A] on the cognition of a mouse model of sporadic Alzheimer’s disease

city: Melbourne

first: Deanna

last: Deveson Lucas

state: Victoria

address: 15 Innovation walk, Monash University, Clayton

country: Australia

institute: Monash University

summary: Increased expression of the voltage-gated potassium channel Kv1.3 in activated microglia and the subsequent release of pro-inflammatory mediators are closely associated with the progression of Alzheimer’s disease (AD). Studies have demonstrated that blockade of microglial Kv1.3 has the potential to improve cognitive function in mouse model of familial AD. Our laboratory has developed a potent and highly selective Kv1.3 peptide blocker, HsTX1[R14A]. This study aims to demonstrate the efficacy of HsTX1[R14A] in a mouse model of sporadic AD, the senescence accelerated mouse (SAMP8). SAMP8 mice were subcutaneously dosed with either PBS or HsTX1[R14A] (1 mg/kg) every other day from 4 months old for 8 weeks. We then performed gene expression profiling analysis using data obtained from RNA-seq of the brains collected from these mice at the conclusion of treatment.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for HsTX1[R14A]- or PBS-treated SAMP8 mouse brains

first: Yijun

last: Pan

first: Jiaqi

first: Liang

last: Jin

first: Joseph

last: Nicolazzo


title: Mutation of Vsx genes in zebrafish highlights the robustness of the retinal specification network

first: Lorena

last: Buono

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

overall_design: Refer to individual Series


title: RNA-Sequencing of mouse melanoma cell line WT31 and its subline WT31_Passage

city: Mannheim

first: Carsten

last: Sticht

address: Theodor-Kutzer-Ufer

department: ZMF

institute: University Heidelberg

summary: RNA-Sequencing analysis of in vitro cultured, sub-confluent (50-75%) mouse melanoma cell line WT31 and its subline WT31_Passage. The subline was generated by repeated in vivo hepatic passaging (5 times).

overall_design: The melanoma cell line WT31 was derived from Tyr∷NrasQ61K/°; INK4a−/− mice and originally generated by O. Sansom (Beatson Institute for Cancer Research, Scotland, UK). After thawing the cells were not passaged more than three times in cell culture. WT31_P5IV subline was generated by hepatic passaging of WT31 five times over the liver after intravenous injection. RNA was extracted from cultured, sub-confluented cells (50% - 75%) for both cell lines and prepared for RNA-sequencing. The library preparation and the sequencing with an Illumina HiSeq 4000 sequencing system (Illumina, CA, USA) were then performed by BGI (Hongkong, China).

first: Sebastian

last: Wohlfeil

first: Bianca

last: Dietsch

first: Carolina

last: Delatorre


title: Early differentially expressed immunological genes   in mild and severe COVID-19

city: Dublin 4

first: Gabriel

last: Gonzalez

state: Co. Dublin

address: Belfield campus

country: Ireland

institute: University College Dublin

summary: We retrospectively analysed the expression of 579 immunological genes in 60 COVID-19 subjects (SARS +ve) and 59 COVID-negative (SARS -ve) subjects using the NanoString nCounter (Immunology panel), a technology based on multiplexed single-molecule counting. Biobanked Human peripheral blood mononuclear cells (PBMCs) samples underwent Nucleic Acid extraction and digital detection of mRNA to evaluate changes in antiviral gene expression between SARS -ve controls and patients with mild (SARS +ve Mild) and moderate/severe (SARS +ve Mod/Sev) disease.

overall_design: 119 samples (60 SARS-CoV-2 positive / 59 SARS-CoV-2 negative)

first: Rachel

last: MacCann

first: Alejandro

last: Garcia


title: Investigating the Cellular Transcriptomic Response Induced by THP-1 macrophages  and T.rubrum- co-culture

city: Ribeirão Preto

first: Ana

middle: Lucia

last: Fachin

state: SP

address: Av. Costabile Romano, 2201

department: Biotechonology  Unit

institute: Unaerp

summary: To evaluate the cellular and molecular response of live and inactive conidia from the co-culture of T. rubrum and THP-1 cell line, by interleukins, LDH quantification and the transcriptional profile of RNA-seq involved in deep infection.

sample_organism:
Trichophyton rubrum
Homo sapiens

platform_organism:
Trichophyton rubrum
Homo sapiens

overall_design: The control (THP-1) and co-culture were carried out in triplicate

first: Bruna

last: Cantelli

first: Gabriela

middle: G

last: Segura

first: Kamila

last: Peronni

first: Pablo

middle: R

last: Sanches

first: Tamires

last: Birencourt

first: Rene

middle: O

last: Beleboni

first: Wilson

last: da Silva Junior

first: Nilce

last: Martinez-Rossi

first: Mozart

last: Marins


title: Gene expression profile at single cell level of non-malignant cells in KRAS syngeneic mouse tumor models

status: Public on May 10 2023

city: Houston

first: John

last: Heymach

address: 1515 Holcombe blvd

institute: UT MD Anderson Cancer Center

summary: STK11/LKB1 mutation is a primary driver for immunotherapy resistance. We employed KRAS/LKB1 syngeneic mouse models by injecting tumor cells with Kras mutation, Kras/Stk11 mutation and MCT4 knockout. We used single-cell RNA-seq to analyze the impact of LKB1 deficiency on the immune microenvironment.

overall_design: LKR13K, LKR13KL and LKR13KL MCT4-KO tumor cells were injected into 129SV mice. The tumors were collected and performed scRNAseq at the Univerisity of Texas MD Anderson Cancer Center Sequencing Core.

first: Yu

last: Qian

first: Minghao

last: Dang

first: Linghua

last: Wang


title: DNA Methylation Variations and Epigenetic Aging in Telomere Biology Disorders

city: Umeå

first: Magnus

last: Hultdin

state: Vasterbotten

address: 6M-building, NUS, 2d floor

department: Department of Medical Biosciences, Pathology

institute: Umeå University

type: Methylation profiling by genome tiling array

summary: DNA methylation profiling of 35 Telomere Biology Disorder (TBD) cases and 20 age-matched controls using the Infinium MethylationEPIC BeadChip arrays (Illumina). The cutoff for methylation differences between the cases and controls was set to |Δβ≥|0.2.

overall_design: The molecular analysis was conducted in leukocytes from whole blood. The DNA was bisulfite converted using the EZ DNA Methylation Kit (Zymo Research, Irvine, CA) and genome-wide assessment of DNA methylation profiles was performed using the Infinium MethylationEPIC BeadChip arrays (Illumina, San Diego, CA) with coverage of >850 000 CpG sites. The beta values for cases and controls were adjusted for leukocyte cell composition using the GSE103541 data set from the Gene Expression Omnibus database repository.

first: Olivia

last: Carlund

first: Anna

last: Norberg

first: Pia

last: Osterman

first: Mattias

last: Landfors

first: Sofie

last: Degerman


title: Goat & cow milk-derived extracellular vesicle miRNomes

city: Saint Genès-Champanelle

first: christine

last: Leroux

address: Route de Theix

institute: INRAE

summary: Comparison of goat and cow milk-derived extracellular vesicle miRNomes (microRNA expression profiles) determined usiing RNA-sequencing (NGS).

sample_organism:
Bos taurus
Capra hircus

platform_organism:
Bos taurus
Capra hircus

overall_design:
Milk from 4 goats and 5 cows were used to obtain RNA from  milk extracellular vesicles
Days postpartum cows: 30 +/- 7
Days postpartum goats: 26 +/- 9

first: Patrice

last: Martin

first: Zuzana

last: Krupova

first: Christine


title: NAT10-mediated RNA acetylation enhances HNRNPUL1 mRNA stability to contribute cervical cancer progression

city: Chongqing

first: yingfei

last: Long

address: Shuanghulu Street

department: The Third Clinical College

institute: Chongqing Medical University

summary: To investigate the principal mechanism of NAT10 in cervical cancer, RNA-seq was accomplished in SiHa and Hela cells with NAT10 knockdown and in control cells

overall_design: The initial letter H represents Hela cells and the initial letter S represents Siha cells. shNC represents the control group and shNAT10 represents the grouping of knocking down NAT10 gene

first: Yingfei


title: Complementary Alu sequences mediate enhancer-promoter selectivity

city: Beijing

first: Yuanchao

last: Xue

address: Datun Road #15

department: Institute of Biophysics

institute: Chinese Academy of Sciences

summary: Enhancers determine spatiotemporal gene expression programs by engaging with long-range promoters. However, it remains unknown how enhancers find their cognate promoters. We recently developed a RIC-seq technology to identify enhancer-promoter connectivity using pairwise interacting enhancer RNAs and promoter-derived noncoding RNAs in HeLa cells. Here, we apply this technology to generate high-confidence enhancer-promoter RNA interaction (EPRI) maps in six additional cell lines. Using these maps, we discover that 37.9% of the enhancer-promoter RNA interaction sites are overlapped with Alu sequences. These pairwise interacting Alu and non-Alu RNA sequences tend to be complementary and potentially form duplexes. Knockout of Alu elements compromises enhancer-promoter looping, whereas Alu insertion or CRISPR-dCasRx-mediated Alu tethering to unregulated promoter RNAs can create new loops to homologous enhancers. Importantly, mapping 535,404 noncoding risk variants back to the EPRI maps enabled us to construct variant-to-function maps for interpreting their molecular functions, including 15,318 deletions or insertions in 11,677 Alu elements that affect 6,497 protein-coding genes. We further demonstrate that polymorphic Alu insertion at PTK2 enhancer can promote tumorigenesis. Our study uncovers a principle for determining enhancer-promoter pairing specificity and provides a framework to link noncoding risk variants to their molecular functions.

overall_design: We applied RIC-seq technology to construct high-resolution enhancer-promoter RNA interaction maps in HeLa and additional six cell lines representing three distinct germ layers, and Chromatin-associated nascent RNA sequencing (ChromRNA-seq) to confirm the decreased transcription of SE351-linked genes after knocking down the corresponding eRNA.

first: Liang

last: Liang

first: Changchang

last: Cao

first: Lei

last: Ji

first: Zhaokui

last: Cai


title: Thioredoxin1 binding metastasis-associated lung adenocarcinoma transcript 1 attenuates inflammation and apoptosis after intracerebral hemorrhage

city: Wuhan

first: Dong

last: Chen

state: Hubei

address: 388 GaoXin 2nd Road, East Lake Hi-Tech Development Zone

department: Center for Genome Analysis

institute: ABLife, Inc.

summary: Post-transcriptional regulation and RNA-binding proteins (RBPs) play vital roles in the occurrence and secondary injury after intracerebral hemorrhage (ICH) and post-transcriptional regulation, respectively.  Therefore, we screened out the differentially expressed RBPs after ICH and found thioredoxin1 (Txn1) as one of the most significantly differentially expressed RBPs. We also used an ICH model and in vitro experiments to investigate the role of Txn1 in clarifying the mechanism of Txn1 in ICH. First, we found that Txn1 was majorly expressed in microglia and neurons in the central nervous system, and its protein level was significantly reduced in perihematoma tissues. Furthermore, we established a Txn1-overexpressing ICH model by stereotaxic injection of adeno-associated virus (AAV) and discovered that the overexpression of Txn1 reduced secondary injury and improved prognosis after ICH.  Moreover, to understand the mechanism of Txn1 after ICH, RNA immunoprecipitation combined with high-throughput sequencing (RIP-seq) showed that Txn1 binds to inflammation-and apoptosis-related mRNAs and affects RNA expression through RNA splicing and translational initiation. Finally, RNA pull-down assays and in vitro experiments confirmed that Txn1 binds to metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) to reduce inflammation and apoptosis. Collectively, our results show that Txn1 reduces ICH-induced brain injury by affecting the post-transcriptional regulation of MALAT1. Consequently, Txn1 may represent a potential therapeutic target for alleviating ICH-induced brain injury.

overall_design: RIP-seq

first: Ru

first: Guoqiang

last: Yang

first: Qingwu


title: N6-Methyladenosine modification of mRNA regulates the transition from 2D to 3D growth in the moss Physcomitrium patens [RNA-seq]

city: Coyoacan

first: V Miguel

last: Palomar

state: Mexico City

address: Circuito Exterior SN

department: Biochemistry

country: Mexico

institute: School of Chemistry, UNAM

summary: In this study, we identified the main genes of the m6A methyltransferase complex (MTC) in P. patens and we demonstrate that their inactivation by genome editing leads to loss of m6A presence in mRNA and results in defects in the moss life cycle. We detected a delay in the formation of gametophore buds, affecting the transition from 2D to 3D growth and subsequent gametophore formation in all MTC mutant plants. We also observed a defect in spore development, with mutant lines producing reduced-size spores that completely failed to germinate. We performed a genome-wide analysis that revealed several transcripts being affected in the Ppmta background. In addition, we demonstrate that the PpAPB1-4 transcripts, known to be central factors regulating the 2D to 3D transition, are m6A-modified, while in the Ppmta mutant the lack of the m6A mark is associated with a corresponding decrease in their transcript accumulation.

overall_design: Total RNA was isolated from Ppmta mutant and wild type lines and subjected to high throughput sequencing. Two replicates are included.

first: David

last: Garcias-Morales

first: V

first: Florence

last: Charlot

first: Fabien

last: Nogué

first: Alejandra

last: Covarrubias

first: Jose

last: Reyes


title: Sites of transcription initiation drive mRNA isoform selection [RNA-seq]

city: Freiburg

first: Valérie

last: Hilgers

address: Stübeweg 51

institute: MPI of Immunobiology and Epigenetics

summary: The generation of distinct messenger RNA isoforms through alternative splicing and alternative 3' end formation influences the expression and function of genes, often in a cell-type specific manner. Here, we quantitatively assess the regulatory relationships between transcription initiation and co-transcriptional processing steps, particularly 3' end formation. Applying multiple long-read-sequencing approaches to obtain an assembly accurately representing even the longest mRNA isoforms from end-to-end, we quantify mRNA isoform choice in Drosophila and human tissues, including the transcriptionally complex nervous system. We find that in Drosophila brains as well as in human cerebral organoids, 3' end site choice is globally influenced by the site of transcription start. We define a subset of TSSs, “dominant promoters” that impose a transcriptional constraint to predetermine splice and polyadenylation variants, which are characterized by specific epigenetic signatures. In vivo deletion or overexpression of dominant promoters disrupted the 3' end expression landscape. Our study demonstrates the crucial impact of transcription initiation site choice on the regulation of transcript diversity and tissue identity.

overall_design: Short-read sequencing of wild type (w1118) Drosophila melanogster heads (2 replicates), and ovaries (2 replicates)

first: Carlos

last: Alfonso-Gonzalez

first: Ivano

last: Legnini

first: Sarah

last: Holec

first: Laura

last: Arrigoni

first: Ulrike

last: Bönisch

first: Agnieszka

last: Rybak-Wolf

first: Nikolaus

last: Rajewsky


title: Sites of transcription initiation drive mRNA isoform selection [PacBio]

overall_design: Pacbio sequencing of wild type (w1118) Drosophila melanogaster heads, in 3 replicates.


title: Identification of the RNA-protein interactions of LRPPRC in A549

first: yuxin

last: Liang

address: No.2 North Street, Zhongguancun, Haidian District, Beijing

institute: Institute of  chemistry，Chinese Academy of Science

summary: To map target transcripts bound with LRPPRC.

overall_design: We performed RNA immunoprecipitation in A549 cell lysis with two technical replications.

first: Wei

last: Zhou

first: Wenxi

last: Wang

first: Yuxin

first: Ruibing

last: Jiang

first: Fensheng

last: Qiu

first: Juan

last: Ni

first: Yang

last: Liu

first: Lina

first: Maowei

first: Weiwei

last: Li

first: Qiqi

last: Xue

first: Lubin

last: Qi

first: Michael

middle: J

last: Donovan

first: Weijia

last: Huang

first: Chenhuan

last: Yu

first: Yue

last: Zhao

first: Jiong

first: Ting

last: Fu

first: Jianguo

last: Feng

first: Weihong

last: Tan

first: Xiaohong

last: Fang


title: Sites of transcription initiation drive mRNA isoform selection [RNADirect]

first: Valérie

last: Hilgers

overall_design: Direct RNA sequencing of wild type (w1118) Drosophila melanogaster heads (1 replicates ), embryos 14-16 hours (3 replicates) , and ovaries (3 replicates)

first: Carlos

last: Alfonso-Gonzalez

first: Ivano

last: Legnini

first: Sarah

last: Holec

first: Laura

last: Arrigoni

first: Ulrike

last: Bönisch

first: Agnieszka

last: Rybak-Wolf

first: Nikolaus

last: Rajewsky


title: Intracellular pH dynamics regulates intestinal stem cell lineage specification

city: San Francisco

first: Todd

last: Nystul

state: CA

address: 513 Parnassus Ave, Box 0412

institute: UCSF

summary: Emerging evidence is revealing critical roles of intracellular pH (pHi) in development (), but it remains unclear whether pHi regulates adult stem cell lineage specification. We find that pHi dynamics is a key regulator of cell fate in the mouse intestinal stem cell (ISC) lineage. We identify a pHi gradient along the crypt axis in intestinal organoids and find that dissipating this gradient by inhibiting Na-H exchanger 1 (NHE1) activity genetically or pharmacologically abolishes crypt budding. Using lineage tracing and single cell RNA sequencing we demonstrate that pHi dynamics acts downstream of Atoh1, with increased pHi promoting differentiation toward the secretory lineage, while reduced pHi biases differentiation into absorptive lineage. Consistent with this, disrupting the pHi gradient blocks new Paneth cell differentiation. Paneth cells provide an essential Wnt signal to ISCs in organoids, and we find that the loss of crypt budding upon NHE1 inhibition can be rescued with exogenous WNTs. Altogether, our findings indicate that pHi is tightly regulated in the ISC lineage and that an increase in pHi is required for Paneth cell specification and thus tissue maintenance. These observations reveal a previously unrecognized role for pHi dynamics in the cell fate specification within an adult stem cell lineage.

overall_design: Both wild-type (WT) and inducible-CRISPR NHE1 knockout (KO) small intestinal organoids were derived from adult mice. Upon NHE1 inhibition, WT organoids were treated with 5-(N-Ethyl-N-isopropyl)-amiloride (EIPA) and KO organoids were treated with doxycycline (Dox) post-plating. All WT, WT+EIPA, KO, KO+Dox organoids were dissociated into single cell suspension and sorted by Fluorescence-activated cell sorting (FACS) prior to scRNAseq.

first: Yi

last: Liu


title: Cardiac pericytes mediate the fibrotic remodeling response to myocardial infarction

city: Los Angeles

first: Reza

last: Ardehali

address: 675 Charles E Young Dr S

department: Internal Medicine, Division of Cardiology

institute: David Geffen School of Medicine at UCLA

summary: Despite the prevalence of pericytes amongst the microvasculature of the heart, the role of pericytes during ischemia-induced remodeling remains unclear. Using chondroitin sulfate proteoglycan 4 (Cspg4) lineage mouse reporters we observed pericytes migrating to the site of injury, expressing pro-fibrotic genes, and causing increased vessel leakage after myocardial infarction. Single cell RNA-sequencing of injured cardiac pericytes exhibited increased expression of genes related to vascular permeability, extracellular matrix production, basement membrane degradation and transforming growth factor-b (TGFb) signaling. Deletion of TGFb receptor 1 in Cspg4-expressing cells improved cardiac ejection fraction and reduced fibrosis post-MI. Whereas  genetic ablation of Cspg4-expressing cells resulted in a rapid decline in cardiac function and increased vascular permeability following MI. Collectively, we show that cardiac pericytes participate in the post-fibrotic response after acute ischemic injury, information that will help guide the development of novel strategies to preserve vascular integrity and attenuate cardiac fibrosis.

overall_design: Hearts from Cspg4-CreER/+;Rosa26-tdT/+ mice were extracted and single-cell RNA sequencing was performed. Mice received either a Sham operation or an MI via LAD ligation.

first: Pearl

last: Quijada

first: Shuin

last: Park

first: Peng

first: Kamal

last: Kolluri

first: David

last: Wong

first: Kevin

last: Shih

first: Kai

first: Arash

last: Pezhouman

first: Lingjun

last: Wang

first: Ali

last: Daraei

first: Matthew

last: Tran

first: Elle

last: Rathbun

first: Kimberly

last: Burgos Villar

first: Maria

last: Garcia-Hernandez

first: Thanh

last: Pham

first: Charles

last: Lowenstein

first: Luisa

last: Iruela-Arispe

first: Stanley

middle: T

last: Carmichael

first: Eric

last: Small


title: Sites of transcription initiation drive mRNA isoform selection [HumanOrganoids]

first: Valérie

last: Hilgers

overall_design: Oxford Nanopore cDNA-PCR sequencing human cerebral organoids, in 3 replicates.

first: Carlos

last: Alfonso-Gonzalez

first: Ivano

last: Legnini

first: Sarah

last: Holec

first: Laura

last: Arrigoni

first: Ulrike

last: Bönisch

first: Agnieszka

last: Rybak-Wolf

first: Nikolaus

last: Rajewsky


title: Sites of transcription initiation drive mRNA isoform selection [FLAMseq]

overall_design: FLAM-sequencing of wild type (w1118) Drosophila melanogaster heads, 3 replicates


title: Sites of transcription initiation drive mRNA isoform selection [cDNANanopore]

overall_design:
Oxford nanopore sequencing of wild type (w1118) Drosophila melanogaster heads (6 replicates), 14-16 hours embryos (3 replicates), 18-20 hours embryos (3 replicates) , and ovaries (3 replicates).

The processed data for GSE203582 is included in the GSE203583_CIA.assembly.allTissues59K.gtf file, which is linked to the SuperSeries GSE203583 record.


title: Sites of transcription initiation drive mRNA isoform selection

sample_organism:
Drosophila melanogaster
Homo sapiens

platform_organism:
Drosophila melanogaster
Homo sapiens

overall_design: Refer to individual Series


title: Plakoglobin is a mechanoresponsive regulator of naïve pluripotency

city: Cambridge

first: Florian

last: Hollfelder

address: 80 tennis court road

country: United Kingdom

institute: University of Cambridge

summary: Biomechanical cues are instrumental in guiding embryonic development and cell differentiation. Understanding how these physical stimuli translate into transcriptional programs could provide insight into mechanisms underlying mammalian pre-implantation development. Here, we explore this by exerting microenvironmental control over mouse embryonic stem cells (ESCs). Microfluidic encapsulation of ESCs in agarose microgels stabilized the naïve pluripotency network and specifically induced expression of Plakoglobin (Jup), a vertebrate homologue of β-catenin. Indeed, overexpression of Plakoglobin was sufficient to fully re-establish the naïve pluripotency gene regulatory network under metastable pluripotency conditions, as confirmed by single-cell transcriptome profiling. Finally, we found that in the epiblast, Plakoglobin was exclusively expressed at the blastocyst stage in human and mouse embryos – further strengthening the link between Plakoglobin and naïve pluripotency in vivo. Our work reveals Plakoglobin as a mechanosensitive regulator of naïve pluripotency and provides a paradigm to interrogate the effects of volumetric confinement on cell-fate transitions.

overall_design: Mouse embryonic stem cells were encapsulated in soft agarose microgels, cultured and RNA-seq profiles were obtained and compared with cells cultured on tissue culture plastic. The study revealed the up-regulation of plakoglobin (Jup) in microgels. Single-cell RNA sequencing using inDrop of cells overexpressing plakoglobin demonstrates a re-establishment of pluripotency  under metastable culture.

first: Joachim

last: De Jonghe

first: Timo

last: Kohler

first: Anna

last: Ellerman


title: Dysregulation of amino acid metabolism upon rapid depletion of cap-binding protein eIF4E [RNA-Seq]

status: Public on May 09 2023

city: Berkeley

first: Paige

last: Diamond

address: Barker Hall

department: MCB

institute: University of California - Berkeley

summary: Protein synthesis is metabolically costly, and the level of translation must match nutrient availability and cellular needs. Overall protein synthesis levels are modulated by regulating translation initiation. The cap-binding protein eIF4E—the earliest contact between mRNAs and the translation machinery—serves as one point of control, but its contributions to mRNA-specific translation regulation remain poorly understood. We acutely depleted eIF4E, which is essential in budding yeast, and observed surprisingly modest effects on cell growth and protein synthesis. Long-lived transcripts were downregulated, likely reflecting accelerated turnover, and the strongest gene-specific effects arose as secondary effects of reduced protein biosynthesis on amino acid pools. Futile cycles of amino acid synthesis and degradation were accompanied by translational activation of GCN4, which is typically induced by amino acid starvation. We further identified translational tuning of PCL5, a negative regulator of Gcn4, that provides a consistent protein-to-mRNA ratio under varying translation environments. This translational control depended in part on a uniquely long poly-(A) tract in the PCL5 5’ UTR and on poly-(A) binding protein. These results highlight the intricate interplay between translation, amino acid homeostasis, and gene regulation and uncover new layers of feedback control in cellular response to stress and nutrient availability.

overall_design: To determine the impact of reduced eIF4E levels on ribosome occupancy transcriptome-wide, we established a eIF4E-AID (auxin inducible degron) yeast strain. We then depleted eIF4E by adding IAA for either 1 or 8 hours. We determined changes in translation and RNA abundance by ribosome profiling and RNA-seq, respectively. A DMSO treated sample was used as a control and both conditions (IAA and DMSO) were performed in duplicate.

first: Nicholas

last: Ingolia

last: McGlincy


title: Dysregulation of amino acid metabolism upon rapid depletion of cap-binding protein eIF4E [aro10_screen]


title: Dysregulation of amino acid metabolism upon rapid depletion of cap-binding protein eIF4E [pcl5_screen]


title: Class 3 PI3K participates in nuclear gene transcription and co-activates the circadian clock to promote de novo purine synthesis [ChIP-seq]

city: Paris

first: Ganna

last: Panasyuk

address: 156-160 Rue Vaugirard, Faculté de Médecine Paris Descartes,, 4th Floor - Lab-408/Office-404,, 4th Floor - Lab-408/Office-404,

institute: Inserm U1151/CNRS UMR 8253, Necker Enfants Malades Institute (INEM)

summary: Metabolic demands fluctuate rhythmically and rely on coordination between the circadian clock and nutrient sensing signaling pathways, yet mechanisms of their interaction remain not fully understood. Here, surprisingly, we find that class 3 Phosphatidylinositol-3-kinase (PI3K), known best for its essential role as a lipid kinase in endocytosis and lysosomal degradation by autophagy, has an overlooked nuclear function in participating to gene transcription as a co-activator of heterodimeric transcription factor and circadian driver BMAL1-CLOCK. Canonical pro-catabolic functions of class 3 PI3K in trafficking rely on the indispensable complex between lipid kinase Vps34 and regulatory subunit Vps15. We find that although both subunits of class 3 PI3K interact with RNA Pol2 and co-localize with active transcription sites, exclusive loss of Vps15 blunted BMAL1-CLOCK transcriptional activity. Thus, we established non-redundancy between nuclear Vps34 and Vps15 reflected in a persistent nuclear pool of Vps15 in Vps34-depleted cells and ability of Vps15 to co-activate BMAL1-CLOCK independently of its complex with Vps34. In physiology, we find Vps15 is required for metabolic rhythmicity in liver and, unexpectedly, it promoted pro-anabolic de novo purine nucleotide synthesis. We show Vps15 activated transcription of Ppat, a key enzyme for production of inosine monophosphate, a central metabolic intermediate for purine synthesis. Finally, we demonstrate that in fasting, which represses clock transcriptional activity, BMAL1 recruitment to chromatin is unmodified but it coincides with depletion of Vps15 on the promoters of its targets, Nr1d1 and Ppat. Our findings suggest that nuclear pool of class 3 PI3K Vps15 subunit could couple transcriptional activity of the circadian clock for temporal regulation of energy homeostasis and it opens novel avenues for establishing the complexity for nuclear class 3 PI3K signaling.

overall_design: Chromatin immunoprecipitation of liver tissue with Bmal1 and H3K27ac in WT and Vps15 liver KO at ZT6.

first: Nathaniel

middle: F

last: Henneman

first: Chantal

last: Alkhoury

first: Volodymyr

last: Petrenko

first: Yui

last: Shibayama

first: Arianna

last: Segaloni

first: Alexis

last: Gadault

first: Ivan

last: Nemazanyy

first: Edouard

last: Le Guillou

first: Amare

last: Wolide

first: Konstantina

last: Antoniadou

first: Xin

last: Tong

first: Teruya

last: Tamaru

first: Takeaki

last: Ozawa

first: Muriel

last: Girard

first: Karim

last: Hnia

first: Dominic

last: Lutter

first: Charna

last: Dibner