title: Diet-induced hyperglycemia regulates the hepatic miRNAome in glucose-intolerant rainbow trout, Oncorhynchus mykiss

summary:
Purpose: Next-generation sequencing (NGS) of small RNAs allows transcriptome level analysis of the miRNAome. The goals of this study are to identify differentially regulated hepatic miRNAs in response to a diet with high (>20%) or low (0%) carbohydrate content following a short term fast using small RNA next generation sequencing. Specific quantitative reverse transcription polymerase chain reaction (qRT–PCR) methods were used to validate optimal high-throughput data analysis, and in silico prediction to identify bona fide target pathways regulated by differentially regulated miRNAs following these nutritional stimuli.
Method: Small RNA sequencing was performed using four randomly selected samples from each treatment group were used for sequencing. The quality of total RNA was confirmed by RNA Integrity Numbers greater than 9.0 using an Agilent Technologies 2100 Bioanalyzer. Nucleotide fractions (15-50) of small RNA were isolated from the total RNA using polyacrylamide gel electrophoresis and were ligated to a 30 adapter followed by a 50 adapter (Illumina, San Diego, CA, USA). The small RNA ligated to the adaptors was reverse transcribed to cDNA, PCR amplified and gel purified. The gel-extracted cDNA was used for library preparation, which was further used for cluster generation on Illumina's Cluster Station before sequencing using Illumina GAIIx. Raw sequence data was obtained from image data using Illumina's Sequencing Control Studio software version 2.8 (SCS v2.8) following real-time sequencing image analysis and base-calling by Illumina's Real-Time Analysis version 1.8.70 (RTA v1.8.70). The ACGT101-miR v4.2 pipeline (LC Sciences) was used to analyze sequenced data. Sequences with low Q scores, reads mapped to mRNA, RFam, Repbase and piRNA database were deleted and unique families were generated from identical sequences. These filtered unique sequences were then mapped to fish pre-miRNA and miRNA using miRBase or to the rainbow trout genome (Berthelot et al., 2014) to identify conserved miRNA following ACGT-101 User's Manual. Unique or novel miRNAs were identified after the BLAST performed against fish miRNAs from miRbase database (release 21) and published miRNAs from rainbow trout (Mennigen et al., 2016). The miRNAs that do not match any sequences in the specified databases and have the propensity of forming hairpin structure with the extended sequences at the mapped positions were classified as unique miRNAs. These unique miRNAs were further classified into different groups depending on the mappable reads to selected miRNAs in miRbase. Differentially expressed miRNAs were identified using the statistical software R (Version 3.2.2) package DESeq2 for one-way ANOVA comparison of all three treatment groups. Real-time RT–PCR validation was performed using SYBR Green assays
Results: Refeeding differentially regulates hepatic miRNA abundance in a diet dependent manner: In trout refed with a  low (0%) CHO diet, refeeding resulted in the 23 differntiaally regulated miRNA, of which 17 miRNAs were upregulated and 6 miRNAs were downregulated. In trout refed with a high (>20%) CHO diet, refeeding resulted in 49 differentially regulated miRNA, of which 20 miRNAs were upregulated and 29 miRNA were downregulated. Irrespecitve of the diet, 5 miRNAs were commonly regulated by refeeding alone, 4 miRNAs were upregulated and 1 was downregulated. When comparing hepatic miRNA profiles in trout following refeeding, there were 30 miRNAs that were differntially regulated after refeeding with different diets, 16 miRNAs were upregulated with the high CHO diet while 14 miRNAs were downregulated with the high CHO diet.
Conclusions: Our study represents the first detailed analysis of the rainbow trout miRNAome in response to nutritional stimuli. Specifically, we here identify common hepatic miRNAs differentially regulated by refeeding following a short-term fast (2d), as well as miRNAs differentially regulated in the postprandial state depending on the dieatary stimulus (no carbohydrates compared to >20% dietary carbohydrates)

overall_design: Hepatic miRNA profiles of 4-day fasted and two acutely refed groups (no carbohydrates, high carbohydrates) were generated by deep sequencing, in four replicates per experimental group, using Illumina GAIIx.

last: Kostnyiuk

first: M

last: Jubonui

last: Martnyiuk

first: S

last: Pansneat

first: L

last: Marannel

last: Mennnien


title: Senescent beta-cells exhibit a unique secretory phenotype that promotes inflammation and remodeling of the extracellular environment

status: Public on Apr 06 2021

city: Boston

first: Hui

last: Pan

address: 1 Joslin Place

department: Bioinformatics and Biostatistics Core

country: USA

institute: Joslin Diabetes Center

type: Expression profiling by high throughput sequencing

summary: Type 2 Diabetes (T2D) patients have higher proportions of senescent beta-cells than their non-diabetic counterparts (Aguayo-Mazzucato et al., 2019). Senescent beta-cells may propagate dysfunction in neighboring cells through the paracrine effects of the senescence-associated secretory phenotype (SASP). To address the heterogeneity in beta-cell SASP expression and its role in T2D, we measured expression levels of beta-cell SASP signature genes in a mouse model of acute insulin resistance using the insulin receptor antagonist, S961. We have previously shown that this model induces hyperglycemia and accelerates beta-cell senescence (Aguayo-Mazzucato et al., 2019). Pancreatic islets were isolated from 3 groups of mice: a control group, a treated group of mice with surgically installed osmotic pumps secreting S961 for 2 weeks, and a third group in which mice recovered from S961 treatment for two weeks. During treatment, mice developed marked hyperglycemia and hyperinsulinemia  which was completely reversed during the two-week recovery period. Islets were dispersed into single cells and scRNASeq was performed using the 10x Genomics Chromium Single Cell Gene Expression Assay.

overall_design: C57Bl6/J male mice (7- to 8-month-old retired breeders) from the Jackson Laboratory were used.  Pumps with the insulin receptor antagonist S961 were surgically installed in mice for two weeks as described previously (Aguayo-Mazzucato et al., 2019). Three groups of mice were used: control (pumps with no S961), S961, and recovered (mice with pumps removed). Treated mice received 20nM of S961 per week. Islets were isolated from mice belonging to each group for single-cell RNA-seq and cultured overnight. Islets were treated with 1 mL of TrypLE Express for 10 minutes at 37C,with vortexing every 3 minutes. Digestion was stopped with media with serum. After filtering, transcriptomic analysis was performed using the 10x Genomics Chromium Single Cell Gene Expression Assay core at BWH.

first: Ayush

last: Midha

first: Cristina

last: Aguayo-Mazzucato


title: SASP signature of beta-cells includes pro-inflammatory and stress response proteins

summary: To define the senescence-associated secretory phenotype (SASP) of beta-cells, we used conditioned media (CM) generated from bleomycin-treated MIN6 cells and from senescent (beta-Gal-positive) primary beta-cells. In order to culture senescent beta-cells, we isolated islet, FACS-sorted them into beta-Gal-positive and negative populations, excluding immune cells through negative selection of CD45-positive and CD11beta-positive cells. For both the MIN6 and primary beta-cell models, we cultured cells in serum-free media to generate CM for proteomic analysis using the aptamer-based SomaScan platform.

sample_organism:
Homo sapiens
Mus musculus

overall_design: After FACS-sorting, beta-Gal-positive and negative primary beta-cells were plated separately in chamber slides or 96-well plates and incubated overnight at 37C and 7% CO2 in serum-free islet media to generate CM. Bleomycin-treated and control MIN6 cells were cultured for 24 hours in serum-free MIN6 media to generate CM. CM was analyzed using SomaScan at the Genomics Proteomics Core at Beth Israel Deaconess Medical Center.


title: The effects of a novel oral nutritional supplement as compared to standard care on body composition, physical function and skeletal muscle mRNA expression in Dutch older adults with (or at risk of) undernutrition

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

country: Netherlands

institute: Wageningen University

summary: In a randomized controlled trial, 82 older adults (>65y) with (or at risk of) undernutrition (n=82) were randomly allocated to 12 weeks of supplementation with a novel supplement (586 kcal, 22 g protein of which 50% whey and 50% casein, 206 mg ursolic acid, 7 g free BCAAs, 11 µg vitamin D) or standard care (600 kcal, 24g protein of which 100% casein, 4 µg vitamin D). Body weight increased significantly in the 12 weeks, both in the intervention group (+1.6 ± 0.2 kg, p<.0001) and in the standard care group (+1.8 ± 0.2 kg, p<.0001). Gait speed during 4m and 400m tests improved over time in the intervention group, whereas the standard care showed no improvements (time*treatment effects 400m: p=0.038 and 4m: p=0.048). Gene sets related to mitochondrial functioning were strongly upregulated in the participants receiving the intervention product. We showed that a novel oral nutritional supplement improves gait speed in older adults via improvements in mitochondrial functioning.

overall_design: Microarray analysis was performed on pre- and post-treatment skeletal muscle biopsies (m. vastus lateralis) from undernourished older adults.

first: Pol

last: Grootswagers

first: Ellen

last: Smeets

first: Antwi-Boasiako

last: Oteng

first: Lisette

last: de Groot


title: The RNA Structurome in the Asexual Blood Stages of Malaria Pathogen Plasmodium falciparum

status: Public on Apr 05 2021

city: Cambridge

first: Sabah

last: Kadri

state: Massachusetts

address: 7 Cambridge Center

department: Computational Biology

institute: Broad institute

type: Non-coding RNA profiling by high throughput sequencing

summary: Plasmodium falciparum is a deadly human pathogen responsible for the devastating disease called malaria. In this study, we measured the differential accumulation of RNA secondary structures in coding and noncoding transcripts from the asexual developmental cycle in P. falciparum in human red blood cells. Our comprehensive analysis that combined high-throughput nuclease mapping of RNA structures by duplex RNA-seq, SHAPE-directed RNA structure validation, immunoaffinity purification and characterization of antisense RNAs collectively measured differentially base-paired RNA regions throughout the parasite’s development. Our mapping data not only aligned to a diverse pool of RNAs with known structures but also enabled us to identify new structural RNA regions in the malaria genome.  On average, approximately 71% of the genes with secondary structures are found to be protein coding mRNAs. The mapping pattern of these base-paired RNAs corresponded to all regions of protein-coding mRNAs, including the 5’ UTR, CDS and 3’ UTR as well as the start and stop codons. Histone family genes which are known to form secondary structures in their mRNAs and transcripts from genes which are important for transcriptional and post-transcriptional control, such as the unique plant-like transcription factor family, ApiAP2, DNA/RNA binding protein, Alba3 and proteins important for RBC invasion and malaria cytoadherence also showed strong accumulation of duplex RNA reads in various asexual stages in P. falciparum. Intriguingly, our study determined a positive relationship between mRNA structural contents and translation efficiency in P. falciparum asexual blood stages, suggesting an essential role of RNA structural changes in malaria gene expression programs.

overall_design: Duplex RNA-seq of Ring, Trophozoite and Schizont stages in P. falciparum

first: Kausik

last: Chakrabarti


title: RNAseq of kidneys from a mice model of chronic leptospirosis infection and induced a secondary nephrotoxic injury by adenine feeding

city: Taoyuan

first: Li-Fang

last: Chou

address: 5 Fu-Shing St., Kueishan

department: Department of Medical Research and Development

country: Taiwan

institute: Chang Gung Memorial Hospital, Linkou

summary:
Background The mysterious chronic kidney disease is multifactorial causes. One of the risk factors is leptospirosis, a re-emerging infectious disease caused by Leptospira spp., for endemic leptospirosis tend to coincide with high-incidence regions of chronic kidney disease of unknown etiology. This study aims to investigate the role of leptospirosis as an emerging culprit in which chronic subclinical kidney injury, may predispose to progressive kidney disease when superimposed on secondary nephrotoxic injury.
Methods Adenine-induced renal injury in chronic leptospira-infected C57/BL6 mice were studied for evaluating the renal function, histology and gene expression changes. We employed RNA sequencing-based renal transcriptome to investigate pathogenic pathways associated with secondary nephrotoxic injury in chronic kidney injury due to leptospirosis.
Results The severity of kidney lesions was increased and the expression of immune/inflammation/fibrosis genes were significantly up-regulated in either low-dose (0.1%) and high-dose (0.2%) adenine-induced renal injury superimposed on infected mice. Whole renal transcriptome analysis reveals that the substantial amplification of the tremendous amount of expressed genes and fibrosis-related pathways was found in adenine-induced kidney injury add-on to leptospira-infected mice which correlated with kidney dysfunction and fibrosis compared to infection or adenine itself. A total of 41 differentially expressed genes associated with fibrosis were identified in infected mice fed with adenine, suggesting that these potential genes contributed to aggravated renal progression occurred in adenine-fed chronic leptospira-infected mice.
Conclusions In summary, this study indicates that chronic subclinical kidney infection when exposed to different degree of secondary nephrotoxic injury may predispose to exacerbated and progressive chronic kidney disease.

overall_design: At 28 days after leptospira infection, infected mice received a secondary nephrotoxic stimulation by adenine in diet. The control group (n=3), without infection and fed with a standard chow; the leptospira-infected mice (n=3), with infection followed by a standard chow; the 0.1% adenine treatment group (n=3), without infection and fed with a chow containing 0.1% adenine for 14 days; the leptospira-infected mice with the second injury (n=3), with infection followed by a chow with 0.1% adenine for 14 days; the 0.2% adenine treatment group (n=3), without infection and fed with a chow containing 0.2% adenine for 14 days; and the leptospira-infected mice with the second injury (n=3), with infection followed by a chow with 0.2% adenine for 14 days.

first: Chih-Wei

last: Yang


title: Cell-free microRNAs circulating with blood in young and aged mice

city: Kodaira

first: Hirohiko

last: Hohjoh

state: Tokyo

address: 4-1-1 Ogawahigashi

country: Japan

institute: National Institute of Neuroscience, NCNP

summary: We investigated cell-free miRNAs in the blood of young and aged mice. The results indicated marked difference in the level of certain miRNAs between the mice.

overall_design: Small RNA samples prepared from the plasma of young and aged C57BL6/J mice were examined by 3D-Gene Mouse miRNA chips.

first: Masashi

last: Fukuoka


title: Compare re-expression of PAX2 in PTENshRNA cells  to PTENshRNA cells alone

city: Chicago

first: Joanna

last: Burdette

state: IL

address: 900 S. Ashland Ave

institute: University of Illinois at Chicago

type: Genome variation profiling by high throughput sequencing

summary: A comparison of the re-expression of PAX2 in PTENshRNA cells  to PTENshRNA cells alone

overall_design: Re-expression of PAX2 in PTENshRNA cells

first: Angela

last: Russo

first: Dimple

last: Modi


title: Genome-wide 5-hydroxymethylcytosine Alterations in Medulloblastoma

city: Atlanta

first: Peng

last: Jin

state: Georgia

address: 615 Michael St., Rm 325

department: Human Genetics

institute: Emory University

type: Methylation profiling by high throughput sequencing

summary: Medulloblastoma (MB) is the most common malignant brain tumor in children and most of its morbidity is caused by relapse or metastasis. Genome-wide 5-hydroxymethylcytosine (5hmC) decrease has been recognized as one of hallmarks for many malignant tumors. Here we show the dramatic decrease of 5hmC abundance in MB, but with wide variations (p<0.001). Genome-wide 5hmC profiling indicates the acquisition of 5hmC at the genomic loci involved in the Notch signaling pathway. Differentially hydroxymethylated genomic regions (DhMRs) in tumor tissues showed similar features with fetus, but not adult, suggesting that DhMRs in tumor prevent tumor from differentiation for normal brain development. By combining the 5hmC abundance and the postoperative survival of the patients, we found that the decrease of 5hmC could serve as a marker and unfavorable prognosis indicator of MB. These results indicate a potential important role of 5hmC in MB.

overall_design: To understand the alteration of 5hmC in medulloblastoma, we determined the genome-wide 5hmC distribution in both tumor and normal samples by employing a previously established chemical labeling and affinity purification method coupled with high-throughput sequencing. Using the pipeline that we have established, we identified common differentially hydroxymethylated genomic regions (DhMRs) among tumor samples.


title: Regulatory neutrophil transcriptional profiling

city: Barcelona

first: Pere

last: Santamaria

address: C/Rosselló 149-153

country: Spain

institute: IDIBAPS

summary: We report that pharmacologically-induced autoantigen-specific T-regulatory type 1 (TR1) cells and TR1 cell-induced B-regulatory (Breg) cells recruit neutrophils into the liver and program their transcriptome to generate regulatory neutrophils

overall_design: We defined the transcriptional profile of neutrophils from liver and blood of pMHC-NP-treated and untreated NOD.c3c4 mice to define the transcriptional make-up defining the disease-regulatory nature of the liver neutrophils that arise in treated mice. We also studied the transcriptional changes in these liver regulatory neutrophils upon pMHC-NP therapy in the presence/absence of cytokine-blocking antibodies.

first: Patricia

last: Solé


title: Gene expression profiles of cultured megakaryocytes

city: Tokyo

first: Keiichi

last: Tozawa

address: 35 Shinanomachi

institute: Keio University School of Medicine

summary:
Cells obtained from adipose tissue are able to differentiate into megakaryocytes.
We compared the gene expression profile of human adipose tissue derived megakaryocytes with that of megakaryocytes differentiated from human CD34 positive cord blood hematopoietic stem cells.

overall_design: Megakaryocytes derived from CD34 positive cord blood stem cells and those derived from adipose tissue were analyzed. Affymetrix U133 plus 2.0 microarrays were used.

first: Yumiko

last: Matsubara

first: Yukako

last: Uruga

first: Taisuke

last: Mori


title: DNA methylation analysis by RRBS of UV irradiated human epidermal melanocytes from neonatal foreskin, HEMn-DP2.

city: Philadelphia

first: Sarah

last: Alp

state: PA

address: 3307 N Broad Street

department: Fels Institute

institute: Temple University

summary: UVR is the greastest risk factor for melanoma develoment. While the role of UVR in DNA mutagenesis is incontrovertable, is is unclear how UVR induced mutations contribute to melanoma. For example, many of the driver mutations ( BRAF V600E or NRAS Q61R) do not bear the UVR signature C>T mutation. To better understand how UVR is contributing to melanoma development, we investigated the non-mutational effect of UVR on the epigenome, specifically DNA methylation.  Aberrant DNA methylation changes are a hallmark in melanoma and there are few reports on the effects of UVR ion DNA methylation. We exposed melanocytes to UVR and cultured them for one-month to detect hertible and stable changes in DNA methylation. We found both hyper and hypo methylated sites after UVR exposure. While many of these changes occured outside of promoters and areas of active gene expression, there were changes in promoter DNA methylation changes that correlated with changes in gene expression.  These changes also correlated with those found in melanoma and UVR sensitive sites were prognostic of patient overall survival. Our work shows for the first time UVR induced DNA methylation changes in melanocytes and may be another way in which UVR contributes to melanoma development.

overall_design: Melanocyte cells were UV irradiated with a broad spectrum wave band ( UVA & UVB) for a total dos of 175 J/m2. Cells were cultured for one month before performing reduced representation bisulfite sequencing (RRBS). Genomic DNA was digested by MspI restriction enzyme, which recgonizes the CCGG sequence, and then subjected to bisulfite converstion. Methylated cytosines within CpG's are protected from sodium bisulfite conversion and remain unchanged. Unmethylated cyotosines undergo a reaction with sodium bisulfite resulting in a base change from cytosine to uracil. This base change allows for the detection of methylated versus unmethylated CpG's through high throughput sequencing.

last: Preston-Alp

first: Jaroslav

last: Jelinek

first: Jean-Pierre

last: Issa

first: M

middle: R

last: Zaidi


title: Neonatal diabetes mutations disrupt a chromatin pioneering function that activates the human insulin gene

status: Public on Apr 03 2021

city: birmingham

first: ildem

last: akerman

address: BR Tower, 3rd Floor, Room 333, Edgbaston,

department: IMSR

country: United Kingdom

institute: University of Birmingham

type: Genome binding/occupancy profiling by high throughput sequencing

summary:
Despite the central role of chromosomal context in gene transcription, human noncoding DNA variants are generally studied outside of their endogenous genomic location. This limits our understanding of disease-causing regulatory variants. INS promoter mutations cause recessive neonatal diabetes. We studied 60 patients with such mutations, and show that all single base mutations disrupt a CC dinucleotide, while none affect elements important for INS promoter function in episomal assays. To model CC mutations, we humanized a ~3.1 kb region of the orthologous mouse Ins2 gene. This drove cell-specific transcription and recapitulated developmental chromatin states. A CC mutant allele, however, abrogated active chromatin formation during pancreas development. A search for transcription factors that act through this element revealed that another neonatal diabetes gene product, GLIS3, had a unique pioneer-like ability to derepress INS chromatin, which was hampered by the CC mutation. Our in vivo analysis, therefore, connects two human genetic defects in a pioneering mechanism that underlies developmental activation of the INS gene.

This record contains the GLIS3 ChIP-seq and input control in human pancreatic islet cells revealing GLIS3 targets in human islet tissue.

overall_design:
In vivo analysis of GLIS3 and neonatal diabetes mutations reveal a pioneering essential step in the activation of the insulin gene:
Chromatin immunoprecipitation was performed on donor-derived human islets (3000 IEQ). DNA was sequenced  using illumina HiSeq 2500. Raw reads were aligned to hg19 (human genome) and peaks were called using MACS2 against the input control file with q < 0.05.

first: Ildem

last: Akerman

first: Jorge

middle: M

last: Ferrer


title: PRMT5 circular RNA promotes the metastasis of urothelial carcinoma of the bladder cells through sponging miR-30c to induce epithelial-mesenchymal transition

status: Public on Apr 04 2021

city: Guangzhou

first: Xin

last: Chen

address: 651 Dongfeng dong Road

country: China

institute: Sun Yat-sen University Cancer Center

summary: We are using circRNA expression profiling to identify novel circular RNA regulating EMT progression in urothelial carcinoma of the bladder.

overall_design: Three tumor tissues and three matched adjacent non-tumor tissues

first: Dan

last: Xie


title: Transcripts identified by microarray analysis that were deregulated in E10.5 Pax2-cre;Tbx2(fl/fl);Tbx3(fl/fl) (Tbx2/3cDKO) otocysts.

city: Hannover

first: Mark-Oliver

last: Trowe

address: Carl-Neuberg-Str. 1

department: Molekularbiologie

country: Germany

institute: Medizinische Hochschule Hannover

summary: Evaluation of the transcriptional changes in E10.5 otocysts upon loss of Tbx2 and Tbx3 in the otic epithelium.

overall_design: Total RNA from dissected otocysts of E10.5 Tbx2/3cDKO and control embryos was analyzed by microarray analysis.

first: Andreas

last: Kispert


title: A tissue-specific self-interacting chromatin domain forms independently of enhancer-promoter interactions

city: Oxford

first: Jelena

last: Telenius

address: MRC Weatherall Institute of Molecular Medicine University of Oxford John Radcliffe Hospital Headington

department: Weatherall Institute of Molecular Medicine (WIMM)

institute: Oxford University

summary: A variety of self-interacting domains, defined at different levels of resolution, have been described in mammalian genomes. These include Chromatin Compartments (A and B), Topologically Associated Domains (TADs), contact domains, sub-TADs, insulated neighborhoods and frequently interacting regions (FIREs).  Whereas many studies have found the organisation of self-interacting domains to be conserved across cell types, some do form in a lineage-specific manner. However, it is not clear to what degree such tissue-specific structures result from processes related to gene activity such as enhancer-promoter interactions or whether they form earlier during lineage commitment and are therefore likely to be prerequisite for promoting gene expression. To examine these models of genome organisation in detail, we used a combination of high-resolution chromosome conformation capture, a newly-developed form of quantitative fluorescence in-situ hybridisation and super-resolution imaging to study a 70 kb self-interacting domain containing the mouse alpha-globin locus. To understand how this self-interacting domain is established, we studied this structure when the genes are inactive and during erythroid differentiation when the genes are progressively switched on. In contrast to many current models of long-range gene regulation, we show that an erythroid-specific, decompacted self-interacting domain, delimited by convergent CTCF/cohesin binding sites, forms prior to the onset of robust gene expression. Using previously established mouse models we also show that formation of the self-interacting domain does not rely on interactions between the alpha-globin genes and their enhancers or on tissue-specific changes in CTCF binding. Rather, formation of the domain simply depends on the presence of activated lineage-specific cis-elements driving a transcription-independent mechanism for opening chromatin throughout the 70 kb region to create a permissive environment for gene expression. These findings are consistent with a model of loop-extrusion in which all segments of chromatin within a region delimited by CTCF boundary elements contact each other. Our findings suggest that activation of tissue-specific element(s) within such a self-interacting region is sufficient to influence all chromatin within such a domain.

overall_design: NG Capture-C combines 3C library preparation with oligonucleotide capture for the desired viewpoint restriction fragments. Presented in this manuscript are chromosome11 viewpoints from promoters Hba1 and Hba2, enhancers R1 and R2 and CTCF binding site 39.5 in the alpha-globin subTAD, together with the central locations of plasmid FISH probes EX, E, A, C, CX designed at intervals across the locus. The method was applied to mES cells and to wild type mouse erythroblasts derived from in vitro culture of mouse foetal liver cells after 0h and 30h differentiation. We also derived erythroblasts from the foetal livers of a mouse model (AMKO) in which a 16kb region encompassing the entire Hba1 and Hba2 genes is deleted.

first: Jill

last: Brown

first: Nigel

last: Roberts

first: Marieke

last: Oudelaar

first: Veronica

last: Buckle


title: Single-Cell Resolution of Lineage Trajectories in the Arabidopsis Stomatal Lineage and Developing Leaf

city: Stanford

first: Dominique

last: Bergmann

state: CA

address: 371 Jane Stanford Way

institute: Stanford University

summary: Dynamic cell identities underlie flexible developmental programs. The stomatal lineages in the Arabidopsis leaf epidermis feature asynchronous and indeterminate divisions that can be modulated by environmental cues. The products of these lineages, stomatal guard cells and pavement cells, regulate plant-atmosphere exchanges, and the epidermis as a whole influences overall leaf growth. How flexibility is encoded in development of the stomatal lineage, and how cell fates are coordinated in the leaf are open questions. Here, we offer single-cell transcriptomes to uncover models of cell differentiation within Arabidopsis leaf tissue.

overall_design: Included are raw and processed single-cell (sc)RNA-seq datasets that profile cells of the Arabidopsis stomatal lineage and developing leaf. The overall design and methods used are described in the associated manuscript.

first: Camila

middle: B

last: Lopez-Anido

middle: C


title: HP1 drives de novo 3D genome reorganization in early Drosophila embryos (RNA-seq)

city: Milano

first: Yinxiu

last: Zhan

address: Via Adamello 16

country: Italy

institute: European Institute for Oncology

summary: The genome of the fertilized egg becomes structurally and functionally reorganized during early embryogenesis. This includes the segregation of active and inactive chromosome regions into A and B compartments, which are a fundamental feature of genome organization in both vertebrates and invertebrates. Mutually exclusive and attractive interactions within each compartment are thought to contribute to compartment formation1,2. However, the molecular nature of compartmental forces, and thus how compartments form, remain unknown. Here we show that HP1a is a major driver of compartmental segregation in Drosophila early development. Depletion of HP1a leads to an overall decrease of compartmentalization and increased intra-chromosomal interactions. Polymer modeling analysis of Hi-C data and ChIP-seq suggest that establishment of the B compartment is driven by HP1a-mediated attractive interactions. Thus, HP1 controls the establishment of higher order 3D structure during early embryogenesis.

overall_design: Role of HP1 on transcriptional level at zygotic genome activation. HP1 was depleted using RNAi and embryos at zygotic genome activation of knockdown and wildtype embryos were collected and analyzed.

first: Fides

last: Zenk

first: Luca

last: Giorgetti

first: Nicola

last: Iovino


title: HP1 drives de novo 3D genome reorganization in early Drosophila embryos (HiC)

overall_design: We study the role of HP1 and H3K9me3 in establishing 3D genome organization at zygotic genome activation (stage 5). HP1 was depleted using RNAi and embryos at zygotic genome activation of knockdown and wildtype embryos were collected and analyzed.


title: MondoA Drives Malignancy in B-ALL through Enhanced Adaptation to Metabolic Stress

city: Munich

first: Maxim

last: Barenboim

address: Koelner Platz 1

department: Department of Pediatrics

institute: TECHNICAL UNIVERSITY OF MUNICH

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

overall_design: Refer to individual Series


title: Histone mark  ChIP-seq in Mutz3 cells

first: Kathrin

last: Bernt

address: 3501 Civic Center Boulevard,

institute: Children's Hospital of philadelphia

summary: We show that translocations place regulatory regions in close proximity to MN1 and enhance gene expression

overall_design: ChIP-seq of H3K27ac and H3K4me1 in a human AML cell line with a t(12;22) translocation that does not generate an MN1-ETV6 fusion protein

first: Simone

middle: S

last: Riedel


title: HP1 drives de novo 3D genome reorganization in early Drosophila embryos


title: Transcriptional response of Prochlorococcus strain MIT0604 to mitomycin C and UV stress

first: Elaina

last: Thomas

state: MA

address: 15 Vassar St

department: Civil and Environmental Engineering

institute: MIT

summary: Prochlorococcus genomes harbor a new type of mobile genetic elements named tycheposons. To study the effects on environmental stress on the gene expression and induction of tycheposons, we subjugated cultures of Prochlorococcus strain MIT0604 containing 7 such elements to treatments with mitomycin C and UV stress.

overall_design: 3 replicate cultures were treated with mitomycin C; 3 replicate cultures were applied a UV shock by irradiating cells. These treated samples were then compared to 3 replicate cultures that were kept as control with no treatment.

first: Raphaël

last: Laurenceau

first: Christina

last: Bliem

first: Thomas

last: Hackl


title: RNA-sequence results of knockdown ETV5 in Human CRC cell line HT29

city: Shanghai

first: Xi

last: Cheng

state: State...

address: Ruijin er lu 197

institute: Shanghai Institute of Digestive Surgery

summary: We report the results of knockdowm ETV5 in human CRC celllines HT29, we built 3 controls and 3 experimental cases

overall_design: To evaluate the genes and pathways correlated with ETV5 difference expression

first: Zhijian

last: Jin

first: Weihua

last: Qiu

first: Xin

last: Lu

first: Ren

last: Zhao


title: miR-30a target genes in the subcutaneous fat pad of diabetic mice

city: Houston

first: Sean

last: Hartig

state: TX

address: One Baylor Plaza, BCM130

department: Molecular and Cellular Biology

institute: Baylor College of Medicine

summary: In this study, we tested the hypothesis that restoration of miR-30a expression in WAT would improve peripheral insulin sensitivity. Exogenous miR-30a expression in subcutaneous WAT depots of obese mice improved insulin sensitivity, decreased ectopic liver fat deposition, and reduced WAT inflammation compared to matched controls.  We identified the target genes and pathways of miR-30a using RNA-Seq technology

overall_design: We injected adenovirus (Adv) expressing miR-30a directly into the subcutaneous fat pad of diabetic mice.  RNA was isolated for library preparation and high throughput sequencing.


title: HP1 drives de novo 3D genome reorganization in early Drosophila embryos (ChIP-seq)

first: Yinxiu

last: Zhan

overall_design: Mapping histone modifications and chromatin proteins during zygotic genome activation


title: The expression profiles of microRNA, circRNA, lncRNA, mRNA in the bladder tissue of partial bladder outlet obstruction rats before and after treatment of human urine-derieved stem cells

status: Public on Apr 02 2021

city: Shenzhen

first: Yuqi

last: Wu

state: Guangdong

address: Xueyuan Road 1098

institute: Shenzhen University


title: Transcriptome Analysis of Signaling Pathways Targeted by Ellagic Acid in Hepatocellular Carcinoma Cells

city: Harbin

first: Guangyue

last: Li

state: heilongjang

address: No.26 Hexing Road Xiangfang District

department: College of Life Science

institute: Northeast Forestry University

summary: Ellagic acid (EA) possesses prominent inhibitory activities against various cancers, including hepatocellular carcinoma (HCC). Recent study showing EA’s activities in suppressing HCC cell proliferation and tumor formation, we further demonstrate that EA reduces cell viability, and induces DNA damage and cell cycle arrest at G1 phase. Employing RNA-Seq, we identified 5765 differentially expressed genes (DEGs) encoding proteins with over 2.0-fold change in EA-treated HepG2 cells. Following Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses, the DEGs caused by EA treatment showed significant enrichment in pathways regulating DNA replication and cell cycle progression. Consistently, integrative analyses of this RNA-seq dataset with a TCGA-LIHC dataset derived from HCC patients confirmed these two pathways as EA’s primary targets. Furthermore, we identified p21 as a primary target gene of EA using the interaction network analysis of the DEGs regulating these pathways. Consistent with the results of our bioinformatic analyses, p21 knockdown desensitized four different liver cancer cell lines to EA treatment. Collectively, our study demonstrated that EA primarily targeted the G1/S phase checkpoint and DNA synthesis to inhibit HCC cell proliferation.

overall_design: RNA-seq was constructed in HepG2 cells with Ellagic acid (EA) and cells with DMSO treatment

first: Shuang

first: Chen

last: Zhong

first: Bo

first: Guang yue

first: Guang chao

last: Sui


title: Targeting glycolysis reduces infection and dampens inflammation in a host-directed therapy for dengue

city: Singapore

first: Hong Kai

last: Lee

address: 8 Biomedical Grove

department: Singapore Immunology Network

country: Singapore

institute: Agency for Science, Technology and Research

summary: Monocytes are the key mediators of inflammatory responses in virus patients, and are also the only blood cell type that supports productive infection both in vitro and in vivo. In this study, we assessed how dengue infection alters the metabolic response of monocytes and affects ensuing inflammatory mechanisms. Using a glycostress assay, we showed that in vitro infected primary human monocytes by dengue virus and monocytes isolated from thrombocytopenic dengue patients, during critical stage of the disease, had increased glycolysis. Single cell transcriptomics revealed an enhanced expression of interferon-related genes and glycolytic genes in classical and intermediate monocytes of dengue patients with more pronounced thrombocytopenia.  Targeting  glycolysis by 2-deoxy-glucose (2DG) reduced dengue infection and dampened pro-inflammatory cytokines in monocytes, both in vitro and in mouse models. Collectively, these results suggest that glycolysis could be a possible target for pharmaceutical intervention to regulate pathogenic host responses in dengue.

overall_design:
Assessment on how dengue infection alters the metabolic response of monocytes and affects ensuing inflammatory mechanisms.
>>> Submitter declares that the raw data will be deposited in the European Genome-phenome Archive (EGA) due to patient privacy concerns. <<<

last: Loy

first: Amit

last: Singhal


title: SIRT6-CBP dependent Nuclear Tau Accumulation and its Role in Protein Synthesis.

city: Beer Sheva

first: Debra

last: Toiber

state: Israel

address: Ben Gurion Road #1, Building 40, room 205

country: Israel

institute: Ben Gurion University of the Negev

summary: Several neurodegenerative diseases present Tau accumulation as the main pathological marker. Tau post-translational modifications such as phosphorylation and acetylation are increased in neurodegenerative patients. Here, we show that Tau hyper-acetylation at residue 174 increases its own nuclear presence and is the result of DNA damage signaling or the lack of SIRT6, both causative of neurodegeneration. Tau-K174ac is deacetylated in the nucleus by SIRT6. However, lack of SIRT6 or chronic DNA damage result in nuclear Tau-K174ac accumulation. Once there, it induces global changes in gene expression affecting protein translation, synthesis and energy production. Concomitantly, AD patients showed increased Nucleolin and a decrease in SIRT6 levels. AD patients present increased levels of nuclear Tau, particularly Tau-K174ac. Our results suggest that increased Tau-K174ac in AD patients is the result of DNA damage signaling and SIRT6 depletion. We propose that Tau-K174ac toxicity is due to its increased stability, nuclear accumulation and nucleolar dysfunction.

overall_design: Four conditions were tested by triplicate (Tau-WT, Tau-K174Q, Tau-S199E and Tau-K174Q-S199E). Overall the data set contains 12 samples. Tau-WT condition was used as reference.

first: Miguel

last: Portillo

first: Ekaterina

last: Eremenko

first: Shai

last: Kaluski

first: Alfredo

last: Garcia

first: Lior

last: Onn

first: Daniel

last: Stein

first: Zeev

last: Slobodnik

first: Adam

last: Zaretsky

first: Uwe

last: Ueberham

first: Monica

last: Einav

first: Martina

middle: K

last: Brückner

first: Thomas

last: Arendt


title: Nostoc punctiforme transcriptome comparison of a SigmaB2 mutant and SigmaB2 overexpression strain

city: Northridge

first: Michael

middle: L.

last: Summers

state: California

address: 18111 Nordhoff St

department: Dept. of Biology

institute: California State University Northridge

summary: Sigma factor B2 is one of 13 sigma factors in this cyanobacterium. To determin the SigB2 regulon, transcriptional differences between a sigB2 mutant and overexpression strain were determined in this study.

overall_design: Analysis used RNA isolated from exponentially grown sigB2 mutant (M) and sigB2 overexpression (OE) strains to maximize gene expression changes in the SigB2 regulon.  Comparisons were made using data from 4 arrays containing dye swaps of two biological samples.

middle: L

first: Jenevieve

last: Polin

first: Arsalan

middle: A

last: Ali


title: The expression profile of miRNA in the bladder tissue of partial bladder outlet obstruction rats before and after treatment of human urine-derived stem cells

first: Yuqi

last: Wu

summary: Obstruction in any part between the outlet of the bladder and the external urethra is called partial Bladder outlet obstruction (pBOO), which is a common disease in urology, which may cause bladder dysfunction. We have comfirmed that human urine-derived stem cells (hUSCs) could improve the bladder function of pBOO rats in our previous study. We thus explored the role of miRNAs in pBOO rats before and after treatment of hUSCs, and revealed that they could serve as effective prevention and treatment therapeutic targets. Rat bladder tissues were used for miRNA sequencing. We identified 8 differentially expressed miRNAs in the pBOO rats before and after treatment of hUSCs.

overall_design:
Bladder tissues miRNA profiles of partial bladder oublet obstruction rat before and after 12 weeks of  treatment of human urine-derived stem cells  were generated by high throughput sequencing using Illumina Hiseq 3000.
Model 1 means a pBOO rat without USCs treated, while model 2 means another rat, comparing to Ctrl rats that means normal SD rats.

first: Menjiang

last: Tu

first: Pei

last: zhu

first: Xiangwei

last: Wang


title: Transcriptional changes induces by activating the nuclear receptor NR2F1

city: New York

first: Bassem

middle: D

last: Khalil

state: NY

address: 1 Gustave L. Levy Pl

institute: Icahn School of Medicine at Mount Siani

summary:
Purpose: to study the transcriptional changes that are induced by activating NR2F1 using an agonist (C26) that we developed.
Methods: T-HEp3-GFP cells were pretreated with DMSO or 0.5 µM C26 for 6 days then inoculated on CAM and treated every 24 hours. After 7 days, tumors were collected and dissociated into single cells.  FACS was used to isolate T-Hep3 tumor cells using GFP. RNA was extracted from fresh frozen cell pellets using Qiagen RNeasy Plus Universal mini kit following manufacturer’s instructions (Qiagen, Hilden, Germany). RNA samples received were quantified using Qubit 2.0 Fluorometer (Life Technologies, Carlsbad, CA, USA) and RNA integrity was checked using Agilent TapeStation 4200 (Agilent Technologies, Palo Alto, CA, USA). RNA sequencing libraries were prepared using the NEBNext Ultra RNA Library Prep Kit for Illumina following manufacturer’s instructions (NEB, Ipswich, MA, USA). Briefly, mRNAs were first enriched with Oligo(dT) beads. Enriched mRNAs were fragmented for 15 minutes at 94°C. First strand and second strand cDNAs were subsequently synthesized. cDNA fragments were end repaired and adenylated at 3’ ends, and universal adapters were ligated to cDNA fragments, followed by index addition and library enrichment by limited-cycle PCR. The sequencing libraries were validated on the Agilent TapeStation (Agilent Technologies, Palo Alto, CA, USA), and quantified by using Qubit 2.0 Fluorometer (Invitrogen, Carlsbad, CA) as well as by quantitative PCR (KAPA Biosystems, Wilmington, MA, USA). The sequencing libraries were clustered on 1 lane of a flowcell. After clustering, the flowcell was loaded on the Illumina HiSeq instrument (4000 or equivalent) according to manufacturer’s instructions. The samples were sequenced using a 2x150bp Paired End (PE) configuration. Image analysis and base calling were conducted by the HiSeq Control Software (HCS). Raw sequence data generated from Illumina HiSeq was converted into fastq files and de-multiplexed using Illumina's bcl2fastq 2.17 software. One mismatch was allowed for index sequence identification. Three replicate samples per group were used . R (v.4.0.3) was used to perform bioinformatics analysis. Quality control was performed using FastQC (v0.11.8). Trim Galore! (version 0.6.5) was used to trim the adapter sequences with a quality threshold of 20. The human genome reference used was GRCh38.p13 and GENCODE release 36. Alignment was performed using STAR aligner (v2.7.5b). Gene level read counts were obtained by using Salmon (v1.2.1) for all libraries . All samples passed the quality control requirements with > 60% of reads uniquely mapping (>20M uniquely mapped reads for each library) using STAR. Differential expression analysis was performed using the gene level read counts and the DESeq2 (v1.28.1) R package. Heatmaps show the z-scores of gene-level read counts and were generated using heatmaply (v1.1.0). Genes with adjusted p-value less than 0.05 were considered differentially expressed.

overall_design: mRNA profiles of DMSO or C26-treated tumors

first: Julio

last: Aguirre-Ghiso


title: Next Generation Sequencing Quantitative Analysis of soybean seeds  Transcriptomes

city: Beijing

first: Wei

last: Wei

address: Chaoyang District, Beichen West Road, Campus #1, No.2

department: State Key Lab of Plant Genomics

institute: Institute of Genetics and Developmental Biology, Chinese Academy of Sciences

summary: To investigate the regulatory mechanisms of GmZF392, GmZF351 and GmNFYA in oil accumulation, seeds at H3 stage of transgenic plants and JACK plants were collected for RNA extraction and sequencing. Before sequencing, qPCR showed that expression of GmZF392, GmZF351 and GmNFYA increased to about 10, 109 and 7 times respectively compared to JACK. Clean reads of RNA-sequencing were mapped to soybean genome and genes that had a 1.5 fold increase or 50% decrease, compared to JACK, were defined as differential expressed genes (DEGs).

overall_design: mRNA profiles of soybean seeds at H3 stage. Wild type (Jack), GmZF351 overexpressing soybean,  GmZF392 overexpressing soybean and GmNFYA overexpressing soybean.


title: Immortalized mouse microglia cell (IMG) response to amyloid beta treatment

city: Boston

first: Yae-Huei

last: Liou

address: 665 Huntington Ave.

department: MET

institute: Harvard School of Public health

summary: RNA-seq analysis of IMG after 8 hours fibril or oligomeric amyloid beta treatment.

overall_design: Differential expression of mRNA from control (scramble), fibrillar mayloid beta (oAb) and oligomeric amyloid beta (fAb) treated IMG. 4 samples from each group were analyzed.

first: Chih-Hao

last: Lee


title: Next-generation sequencing of migratory innate lymphoid cells, T cells and dendritic cells (WT and CD11a KO) in mesenteric lymph

city: Glasgow

first: John

middle: Joseph

last: Cole

address: University Avenue

institute: University of Glasgow


title: The expression profile of circRNA, lncRNA and mRNA in the bladder tissue of partial bladder outlet obstruction rats before and after treatment of human urine-derived stem cells

summary: Obstruction in any part between the outlet of the bladder and the external urethra is called partial Bladder outlet obstruction (pBOO), which is a common disease in urology, which may cause bladder dysfunction. We have comfirmed that human urine-derived stem cells (hUSCs) could improve the bladder function of pBOO rats in our previous study. We thus explored the role of miRNAs in pBOO rats before and after treatment of hUSCs, and revealed that they could serve as effective prevention and treatment therapeutic targets. Rat bladder tissues were used for circRNA, lncRNA and mRNA sequencing.

overall_design:
Bladder tissues circRNA, lncRNA and mRNA profiles of partial bladder oublet obstruction rat before and after 12 weeks of treatment of human urine-derived stem cells were generated by high throughput sequencing using Illumina Hiseq 3000.
Model 1 means a pBOO rat without USCs treated, while model 2 means another rat, comparing to Ctrl rats that means normal SD rats.


title: Trx knockdown in intestinal stem cells (ISCs) of Drosophila melanogaster

city: Warsaw

first: Michal

last: Pawlak

address: Domaniewska 39 B

country: Poland

institute: Roche Polska Sp. z o.o.

summary: Male flies were crossed to virgins of genotype yw;esg-Gal4,UAS-GFP;tub-Gal80ts/Tm3,Sb. Progeny was collected and aged 3-4 days before shifting for 7 days to 29 degrees to induce RNAi expression. Guts were dissected for 2 biological replicates/condition, each containing 80-100 guts, dissociated and GFP-positive cells were FACS-sorted. RNA was isolated using the PicoPure RNA-isolation kit and libraries were generated. Sequencing was done by distributing the 4 libraries over 5 lanes on the Illumina HiSeq2000 platform.

overall_design: Two biological replicate samples for each of two groups before and after siRNA knockdown of Trx.

first: Helen

last: Tauc

first: Imilce

last: Rodriguez-Fernandez

first: Jason

last: Hackney

first: Tal

last: Ronnen Oron

first: Jerome

last: Korzelius

first: Hagar

middle: F

last: Moussa

first: Subhra

last: Chaudhuri

first: Zora

last: Modrusan

first: Bruce

last: Edgar

first: Heinrich

last: Jasper


title: ATACseq of Polycomb KD ISCs [NGS2975]

summary: Tissue homeostasis requires long-term lineage fidelity of somatic stem cells. Whether and how age-related changes in somatic stem cells impact the faithful execution of lineage decisions remains largely unknown. Here, we address this question using genome-wide chromatin accessibility and transcriptome analysis as well as single cell RNA-seq to explore stem cell intrinsic changes in the aging Drosophila intestine. These studies indicate that in stem cells of old flies, promoters of Polycomb (Pc) target genes become differentially accessible, resulting in the increased expression of enteroendocrine (EE) cell specification genes. Consistently, we find age related changes in the composition of the EE progenitor cell population in aging intestines, as well as a significant increase in the proportion of EE-specified ISCs and progenitors in aging flies. We further confirm that Pc-mediated chromatin regulation is a critical determinant of EE cell specification in the Drosophila intestine. Pc is required to maintain expression of stem cell genes while ensuring repression of differentiation and specification genes. Our results identify Pc group proteins as central regulators of lineage identity in the intestinal epithelium and highlight the impact of age-related decline in chromatin regulation on tissue homeostasis.

overall_design: For Pc-RNAi experiments, virgins of the ISC ts fly line were crossed with cherry-RNAi or Pc-RNAi (BL36070) males and maintained at 18 °C. Progeny was collected and shifted to 29°C 4-7 days after eclosion. Flies were kept at 29 °C for 8-11 days at which point they were dissected and ISCs were sorted by Fluorescent Activated Cell Sorting (FACS) as we have previously described (Tauc et al., 2014). In short, midguts from flies of WDah; esg-Gal4, UAS-2xEYFP; Su(H)GBE-Gal80, were dissected in 1xPBS, 1% Bovine Serum Albumin (BSA) and dissociated in 0.5% Trypsin-EDTA solution for less than 2h at room temperature (RT), during which dissociated cells were collected periodically every 20-30min, resuspended in 1xPBS, 1%BSA and 2%FBS and kept on ice until sorting. A BD Biosciences FACSAria II flow cytometer cell sorter was used for cell sorting.

last: Oron


title: Small RNA sequencing reveals distinct miRNA signatures and networks during fetal and adult erythroid differentiation

city: Nicosia

first: Carsten

middle: Werner

last: Lederer

address: 6 Iroon Avenue, Ayios Dometios

department: Molecular Genetics Thalassaemia Department

country: Cyprus

institute: The Cyprus Institute of Neurology and Genetics

summary: MicroRNAs (miRNAs) are small non-coding RNAs (sncRNAs) crucial for post-transcriptional and translational regulation of cellular and developmental pathways. Study of miRNAs in erythropoiesis thus elucidates the process and facilitates diagnosis and therapy development for related disorders. Here, we used high-throughput DNA Nanoball small RNA sequencing to characterise sncRNAs, and in particular miRNAs, comprehensively in human erythroid cell cultures. Based on primary human peripheral-blood-derived CD34+ (hCD34+) cells and on two influential erythroid cell lines with adult and fetal expression patterns, HUDEP-2 and HUDEP-1, respectively, our study links differential expression (DE) to erythroid differentiation, cell type, and fetal and adult hemoglobin expression patterns. Sequencing results were validated by reverse-transcription quantitative PCR (RT-qPCR) of selected DE miRNAs (miR-451a, miR-182-5p, miR-146a-5p, miR-221-3p, miR-222-3p) and indicated overall common erythroid differentiation signatures in primary cells and cell lines, in terminal stages of erythropoiesis characterized by decreased complexity of global miRNA expression and reciprocally increased expression of individual lineage-specific miRNAs. Interestingly, against a backdrop of evident clustering and thus similarity of same-stage hCD34+ and HUDEP-2 cells, differential expression of several miRNAs highlighted readily interpretable discrepancies between both cell types. Moreover, differential expression analysis between the closely related HUDEP-2 and HUDEP-1 cells provided pointers to miRNAs, transcription factors (TFs), target genes, and pathways mechanistically linked to globin switching. In resulting TF-miRNA co-regulatory networks, TFs known to be implicated in hemoglobin switching, including the major γ-globin repressors BCL11A and ZBTB7A, were targeted by many co-regulated miRNAs, outlining intricate combinatorial miRNA regulation of globin expression in erythroid cells.

overall_design: Small RNA sequencing profile of hCD34+, HUDEP-1 and HUDEP-2 cells before (early, _E) and after (late,_L) erythroid differentiation. Three biological replicates in each stage were analyzed, labelled A, B and C (total 18 samples)

middle: W

first: Panayiota

last: Papasavva

first: Nikoletta

middle: Y

last: Papaioannou

first: Petros

last: Patsali

first: Maria

last: Sitarou

first: Soteroulla

last: Christou

first: Marina

last: Kleanthous

first: Ryo

last: Kurita

first: Yukio

last: Nakamura


title: aCGH analysis of liver tumors generated in FAH-/- Trp53-/- mice, a mouse model of metabolic liver injury

city: Ulm

first: André

last: Lechel

address: Albert-Einstein-Allee 23

department: Department of Internal Medicine I

institute: University hospital Ulm

type: Genome variation profiling by genome tiling array

summary:
Background and aims: A coordinated stress and regenerative response is important following hepatocyte damage. Here, we investigate the phenotypes that result from genetic abrogation of individual components of the CHK2/ p53/ p21 pathway in a murine model of metabolic liver injury.
Methods: NTBC was reduced or withdrawn in Fah /  mice lacking Chk2, p53 or p21, and survival, tumor development, liver injury and regeneration were analyzed. Partial hepatectomies were performed and mice were challenged with the Fas-antibody Jo2.
Results: In a model of metabolic liver injury, loss of p53, but not of Chk2, impairs the oxidative stress re-sponse and aggravates liver damage, indicative of a direct p53-dependent protective effect on hepatocytes. Cell cycle control during chronic liver injury critically depends on the presence of both p53 and its downstream effector p21. In p53-deficient hepatocytes, unchecked proliferation occurs despite a strong induction of p21, revealing a complex interdependency between p21 and p53. The increased regenerative potential in the absence of p53 cannot fully compensate the surplus injury and is not sufficient to promote survival. Despite the different phenotypes as-sociated with the loss of individual components of the DNA damage response, gene expression patterns are dominated by the severity of liver injury, but reflect distinct effects of p53 on prolif-eration and the anti-oxidative stress response.
Conclusion: Characteristic phenotypes result from the genetic abrogation of individual components of the DNA damage response cascade in a liver injury model. The extent to which loss of gene function can be compensated, or affects injury and proliferation, depends on the level at which the cas-cade is interrupted.

overall_design: Liver tumors from 4 invidual FAH-/- Trp53-/- mice were analyzed, pooled liver DNA from 3 sex matched FAH+/+ Trp53+/+ littermates were used as controls

first: Laura

middle: E

last: Buitrago-Molina

first: Silke

last: Marhenke

first: Arndt

last: Vogel


title: aCGH analysis of liver tumors generated in FAH-/- mice, a mouse model of metabolic liver injury

summary:
Background and aims: A coordinated stress and regenerative response is important following hepatocyte damage. Here, we investigate the phenotypes that result from genetic abrogation of individual components of the CHK2/ p53/ p21 pathway in a murine model of metabolic liver injury.
Methods: NTBC was reduced or withdrawn in FAH-/-  mice lacking Chk2, p53 or p21, and survival, tumor development, liver injury and regeneration were analyzed. Partial hepatectomies were performed and mice were challenged with the Fas-antibody Jo2.
Results: In a model of metabolic liver injury, loss of p53, but not of Chk2, impairs the oxidative stress re-sponse and aggravates liver damage, indicative of a direct p53-dependent protective effect on hepatocytes. Cell cycle control during chronic liver injury critically depends on the presence of both p53 and its downstream effector p21. In p53-deficient hepatocytes, unchecked proliferation occurs despite a strong induction of p21, revealing a complex interdependency between p21 and p53. The increased regenerative potential in the absence of p53 cannot fully compensate the surplus injury and is not sufficient to promote survival. Despite the different phenotypes as-sociated with the loss of individual components of the DNA damage response, gene expression patterns are dominated by the severity of liver injury, but reflect distinct effects of p53 on prolif-eration and the anti-oxidative stress response.
Conclusion: Characteristic phenotypes result from the genetic abrogation of individual components of the DNA damage response cascade in a liver injury model. The extent to which loss of gene function can be compensated, or affects injury and proliferation, depends on the level at which the cas-cade is interrupted.

overall_design: Liver tumors from 8 invidual FAH-/- mice were analyzed, pooled liver DNA from 6 sex matched FAH+/+ littermates were used as controls

last: Elisa Buitrago-Molina

first: Robert

last: Geffers


title: RNAseq of Aging Drosophila ISCs

first: Michal

last: Pawlak

overall_design: Whole midguts were dissociated into single cells across flies lifespan, covering early adulthood (7 days of age), middle age (30 days of age), old age (60 days) and geriatric age/very old (>70 days) and ISCs were sorted by Fluorescent Activated Cell Sorting (FACS) as we have previously described (Tauc et al., 2014). In short, midguts from flies of WDah; esg-Gal4, UAS-2xEYFP; Su(H)GBE-Gal80, were dissected in 1xPBS, 1% Bovine Serum Albumin (BSA) and dissociated in 0.5% Trypsin-EDTA solution for less than 2h at room temperature (RT), during which dissociated cells were collected periodically every 20-30min, resuspended in 1xPBS, 1%BSA and 2%FBS and kept on ice until sorting. A BD Biosciences FACSAria II flow cytometer cell sorter was used for cell sorting.

first: Helen

last: Tauc

first: Imilce

last: Rodriguez-Fernandez

first: Jason

last: Hackney

first: Tal

last: Ronnen Oron

first: Jerome

last: Korzelius

first: Hagar

last: F. Moussa

first: Subhra

last: Chaudhuri

first: Zora

last: Modrusan

last: A. Edgar

last: Jasper


title: aCGH analysis of liver tumors generated in FAH-/- Trp53-/- and FAH-/- mice, a mouse model of metabolic liver injury

last: Lechel

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

overall_design: Refer to individual Series


title: ATACseq of Aging Drosophila ISCs [NGS2046]

last: Oron

last: Moussa

first: Bruce

last: Edgar

first: Heinrich


title: RNAseq of Polycomb KD ISCs

overall_design: For polycomb (Pc)-RNAi experiments, virgins of the ISCts fly line were crossed with cherry-RNAi or Pc-RNAi (BL36070) males and maintained at 18 °C. Progeny was collected and shifted to 29°C 4-7 days after eclosion. Flies were kept at 29 °C for 8-11 days at which point they were dissected and ISCs were isolated. In short, midguts were dissected in 1xPBS, 1% Bovine Serum Albumin (BSA) and dissociated in 0.5% Trypsin-EDTA solution for less than 2h at room temperature (RT), during which dissociated cells were collected periodically every 20-30min, re537 suspended in 1xPBS, 1%BSA and 2%FBS and kept on ice until sorting. A BD Biosciences FACSAria II flow cytometer cell sorter was used for cell sorting.


title: RNA-seq of P14 CD8 T cells during acute LCMV infection

city: Chicago

first: Weiguo

last: Cui

address: 303 E Chicago Ave

department: Pathology

institute: Northwestern University

summary: We performed RNA-seq, ATAC-seq, and H3K27Ac ChIP-seq on P14 CD8 T cells during acute infection with LCMV Armstrong on days 0 (naïve cells), 9 (memory precursor effector cells (MPEC) and short-lived effector cells (SLECs)), and 40+ (memory cells) post infection.

overall_design: 4 samples (Naive, MPEC, SLEC, Memory), 2-3 biological replicates per sample

first: David

last: Schauder


title: H3K27Ac ChIP-seq of P14 CD8 T cells during acute LCMV infection

overall_design: 4 samples (Naive, MPEC, SLEC, Memory), 2 biological replicates per sample, plus one input control


title: ATAC-seq of P14 CD8 T cells during acute LCMV infection

overall_design: 4 samples (Naive, MPEC, SLEC, Memory), 2 biological replicates per sample


title: ATAC-seq of MIG, MIG-E47, or MIG-ID2 transduced CD8 T cells in vitro

summary: We performed RNA-seq and ATAC-seq on in vitro activated P14 CD8 T cells transduced with MIG (empty), MIG-E47, or MIG-ID2.

overall_design: 3 samples (MIG, MIG-E47, MIG-ID2), 2 biological replicates per sample


title: Identification of human glucocorticoid response markers using integrated multi-omic analysis

city: Manchester

first: Adam

last: Stevens

address: Oxford Road

department: Medicine

institute: University of Manchester


title: Identification of human glucocorticoid response markers using integrated multi-omic analysis [Adipose Tissue]

summary:
A multi-omic approach in a clinical experimental study identified circulating biomarkers reflecting glucocorticoid exposure.
Background: Endogenous glucocorticoids (GC) are mechanistically linked to common diseases and are important as drugs in the treatment of many disorders. There is no marker that can measure and quantify GC action. Our aim was to identify circulating biomarkers of GC action using a clinical experimental study.  Methods: In a randomized, crossover, single-blind trial, subjects with primary adrenal insufficiency received intravenous hydrocortisone infusion in a circadian pattern (physiological GC exposure) or isotonic saline (GC withdrawal) over 22 hours. Samples were collected at 7 AM (end of infusion). Integrated multi-omic analysis was used because of the complexity in GC action and the low number of subjects. The transcriptome in peripheral blood mononuclear cells (PBMCs) and adipose tissue, plasma miRNAomic, and serum metabolomics were compared between the interventions. Replication of the plasma miRNA findings was performed in three independent studies.  Results: During GC withdrawal, overnight urinary cortisol and cortisone excretion were undetectable. Correlation and hypernetwork analyses identified a transcriptomic profile derived from PBMCs and adipose tissue predictive of GC exposure, and a multi-omic cluster predictive of GC exposure. From the circulating ‘omic data, decreased expression of plasma miR-122-5p was associated with increased GC exposure. This finding was reproduced in three independent studies.  Conclusion: We developed a human experimental model for physiological GC exposure and withdrawal. The integrated multi-omic data identified circulating miRNAs and metabolites associated with GC-responsive genes. In independent studies, miR-122-5p was shown to be associated with GC exposure.  Background: Endogenous glucocorticoids (GC) are mechanistically linked to common diseases and are important as drugs in the treatment of many disorders. There is no marker that can measure and quantify GC action. Our aim was to identify circulating biomarkers of GC action using a clinical experimental study.

overall_design: Ten subjects (4 women with 3 of them post-menopausal) with a median age of 50 years (range, 25 to 57) and a median disease duration of 23.5 years (range, 1 to 33) completed all aspects of the study. The median daily replacement dose of hydrocortisone (HC) prior to the study was 30 mg (range, 20 to 30) and 9 out of 10 subjects had treatment with fludrocortisone (mineralocorticoid) at a median daily dose of 0.1 mg (range, 0.1 to 0.2). The main time points for sample collection in each intervention were at 9 AM on the first intervention day (“before start”) and at 7 AM on the second intervention day (“morning”) (Fig. 1B). The subjects’ last ordinary oral HC dose was administered the day before admission to the study unit.   Infusion of HC mixed with isotonic saline (“GC exposure”) had no effect on systolic and diastolic blood pressure, body weight, serum sodium and potassium, or plasma glucose concentrations compared to the same amount of isotonic saline infusion alone (“GC withdrawal”).Differential expression was conducted on the ‘omic data from the samples collected at 7 AM comparing physiological GC exposure to GC withdrawal

first: Gudmundur

last: Johannsson

first: Dimitrios

last: Chantzichristos


title: Lipocalin 2 modulates dendritic cell activity and shapes immunity to influenza in a microbiome dependent manner

city: Vienna

first: Sylvia

last: Knapp

address: Währinger Gürtel 18-20

country: Austria

institute: Medical University of Vienna

summary: Lipocalin 2 (LCN2) is a secreted glycoprotein with roles in multiple biological processes. It contributes to host defense by interference with bacterial iron uptake and exerts immunomodulatory functions in various diseases. Here, we aimed to characterize the function of LCN2 in lung macrophages and dendritic cells (DCs) using Lcn2-deficient mice. Transcriptome analysis revealed strong LCN2-related effects in CD103+ DCs during homeostasis, with differential regulation of antigen processing and presentation and antiviral immunity pathways. We next validated the relevance of LCN2 in a mouse model of influenza infection, wherein LCN2 protected from excessive weight loss and improved survival. LCN2-deficiency was associated with enlarged mediastinal lymph nodes and increased lung T cell numbers, indicating a dysregulated immune response to influenza infection. In vivo T cell chimerism and in vitro T cell proliferation assays indicated that improved antigen processing by CD103+ DCs, rather than T cell intrinsic effects of LCN2, contribute to the exacerbated T cell response. Considering the antibacterial potential of LCN2 and that commensal microbes can modulate antiviral immune responses, we speculated that LCN2 might cause the observed influenza phenotype via the microbiome. Comparing the lung and gut microbiome of WT and Lcn2-/- mice by 16S rRNA gene sequencing, we observed profound effects of LCN2 on gut microbial composition. Interestingly, antibiotic treatment or co-housing of WT and Lcn2-/- mice prior to influenza infection equalized lung CD8+ T cell counts, suggesting that the LCN2-related effects are mediated by the microbiome. In summary, our results highlight a novel regulatory function of LCN2 in the modulation of antiviral immunity.

overall_design: 21 RNA Seq Samples of CD103+ dendritic cells (CD103) or alveolar macrophages (AMs) sorted from lungs WT and Lcn2KO mice (homeostasis); and 5 RNA Seq samples of CD103+ dendritic cells isolated from mediastinal lymph nodes (mLN) of mice infected with 10,000 PFU of OT1-PR/8 influenza virus.

first: Martin

last: Watzenböck

first: Barbara

last: Drobits

first: Sophie

last: Zahalka

first: Anna-Dorothea

last: Gorki

first: Asma

last: Farhat

first: Federica

last: Quattrone

first: Anastasiya

last: Hladik

first: Karin

last: Lakovits

first: Gabriel

last: Richard

first: Therese

last: Lederer

first: Birgit

last: Strobl

first: Gijs

last: Versteeg

first: Philipp

last: Starkl

first: Louis

last: Boon


title: RNA-seq of MIG, MIG-E47, or MIG-ID2 transduced CD8 T cells in vitro

first: Weiguo

last: Cui

first: David

last: Schauder


title: Identification of human glucocorticoid response markers using integrated multi-omic analysis [Peripheral blood mononuclear cells]

first: Adam

last: Stevens

first: Gudmundur

last: Johannsson

first: Dimitrios

last: Chantzichristos


title: Next-generation sequencing of migratory innate lymphoid cells, T cells and dendritic cells in mesenteric lymph [CD11a KO DC]

status: Public on Apr 01 2021

first: John

last: Cole

summary: This study was designed to identify NGS-derived transcriptomes (RNA-sequencing) of different cell populations (ILCs, T cells and DCs) isolated from the lamina propria of the intestine and from the thoracic duct. For the DC subset, CD11a WT and KO samples were used.

overall_design:
Transcriptomic profile of migratory cells isolated from the thoracic duct and comparison to their tissue resident counterparts.
This dataset includes CD11a KO dendritic cells from the small intestinal lamina propria and intestinal lymph nodes. These samples were prepared and sequenced alongside the samples in GSE160156.

first: Vicky

last: Morrison

first: Claire

last: McIntyre

first: Simon

last: Milling


title: Quantitative analysis of phosphoproteome in necroptosis reveals a role of TRIM28 phosphorylation in promoting necroptosis-induced cytokine production

city: Shanghai

first: Dean

last: Li

address: Haike Road 100

institute: Chinese Academy of Science

summary: Necroptosis is a form of regulated necrotic cell death which promotes inflammation. In cells undergoing necroptosis, the activated RIPK1 kinase mediates the formation of RIPK1/RIPK3/MLKL complex to promote MLKL oligomerization and execution of necroptosis. RIPK1 kinase activity also promotes cell-autonomous activation of proinflammatory cytokine production in necroptosis. However, the signaling pathways downstream of RIPK1 kinase in necroptosis and how RIPK1 kinase activation controls inflammatory response induced by necroptosis are still largely unknown. Here we quantitatively measured the temporal dynamics of over 7000 confident phosphosites during necroptosis using mass spectrometry. Our study defined a RIPK1-dependent phosphorylation pattern in late necroptosis that is associated with a proinflammatory component marked by p-S473 TRIM28. We show that the activation of p38 MAPK mediated by oligomerized MLKL promotes the phosphorylation of S473 TRIM28, which in turn mediates inflammation during late necroptosis. Taken together, our study illustrates a mechanism by which p38 MAPK may be activated by oligomerized MLKL to promote inflammation in necroptosis.

overall_design: mRNA profiles of Trim28 -/-, Trim28 +/-, Trim28 S473A/- and Trim28 S473E/- human HT-29 cell treated with either TNFα and Smac and zVAD or vehicle were generated by deep sequencing, in quadruplicate, using Illumina Nextseq.

first: Rui

last: Zu

first: Zhen

last: Yu

first: Junying

last: Yuan

first: Bing

last: Shan


title: small RNA-seq analysis of transcriptome profiling in Apis cerana

city: Seoul

first: Igojo

last: Kang

state: Seoul

address: Gwanak_1 Gwanak-ro, Gwanak-gu

department: Applied Biology and Chemistry

country: South Korea

institute: Seoul National University

summary: High-throughput RNA sequencing was carried out for understanding of caste and organs of Asian honeybees.

overall_design: Examination of 18 development stage and 11 different organs in Apis cerana

first: Woojin

last: Kim

first: Jae

last: Lim

first: Chanseok

last: Shin


title: Protein neddylation as a therapeutic target in pulomonary and extrapulonary small cell carcinomas

city: Seattle

first: Justin

last: Norton

state: WA

address: 1100 Fairview Ave N

department: Human Biology

institute: Fred Hutchinson Cancer Research Center


title: Protein neddylation as a therapeutic target in pulomonary and extrapulonary small cell carcinomas [MSK-LX227C]

summary: Gene expression in human small cell carinoma PDX models treated with MLN4924

overall_design: Identification of differentially expressed genes in the extrapulmonary small cell carcinoma PDX model MSK-LX227C upon 15 days of treatment with MLN4924 or vehicle

first: David

last: MacPherson

middle: P


title: Protein neddylation as a therapeutic target in pulomonary and extrapulonary small cell carcinomas [FHSC39]

overall_design: Identification of differentially expressed genes in the SCLC PDX model FHSC39 upon 15 days of treatment with MLN4924 or vehicle


title: Gene expression changes induced by Dendrocalamus latiflorus leaf extract (DLE) and white tea extract (WTE) and pioglitazone (PGZ) in db/db mice liver

first: kun

last: luo

state: Beijing

address: Haidian district

institute: Tsinghua university

summary: Eight-week-old C57BL/6J and T2DM db/db mice randomly divided into 4 groups of 8 based on equal bodyweight and fasting blood glucose (FBG) levels (mice were fasted for 6 h). The C57BL/6J mice (normal group) were treated with saline. The T2DM db/db mice were treated with saline, DLE (200 mg/kg), WTE (180 mg/kg), or pioglitazone (PGZ, 20 mg/kg) for 12 weeks, and were called the model group, DLE group, WTE group and PGZ group, respectively. All mice received oral administration of the indicated treatments once a day.  After 12 weeks drug administration, liver aliquots from four randomly selected mice from the normal, model, PGZ and DLE groups were ground into powder in liquid nitrogen, followed by RNAseq for differentially expressed gene determination and pathway enrichment analysis.

overall_design: Liver mRNA profiles of wild type mice, and db/db mice treated with saline, DLE or PGZ.

first: Kun

last: Luo

first: Xiaoling

last: Zhang

first: Wenting

last: Huang

first: Lan

last: Xie

first: Jing

last: Cheng


title: Protein neddylation as a therapeutic target in pulomonary and extrapulonary small cell carcinomas [JHU-LX108]

overall_design: Identification of differentially expressed genes in the SCLC PDX model JHU-LX108 upon 15 days of treatment with MLN4924 or vehicle


title: Sulfopin, a covalent inhibitor of Pin1, blocks Myc-driven tumors in vivo

city: Rehovot

first: Efrat

last: Resnick

address: Herzel 234

department: Chemical and structural biology

institute: Weizmann institute of science

summary:
The peptidyl-prolyl isomerase, Pin1, is exploited in cancer to activate oncogenes and inactivate tumor suppressors. However, despite considerable efforts, Pin1 has remained an elusive drug target. Here, we screened an electrophilic fragment library to identify covalent inhibitors targeting Pin1’s active site Cys113, leading to the development of Sulfopin, a nanomolar Pin1 inhibitor. Sulfopin is highly selective, as validated by two independent chemoproteomics methods, achieves potent cellular and in vivo target engagement, and phenocopies Pin1 genetic knockout.
Pin1 inhibition had only a modest effect on cancer cell line viability. Still, Sulfopin induced downregulation of c-Myc target genes, reduced tumor progression and conferred survival benefit in murine and zebrafish models of MYCN-driven neuroblastoma, and in a murine model of pancreatic cancer. Our results demonstrate that Sulfopin is a suitable chemical probe for assessing Pin1-dependent pharmacology in cells and in vivo, and that Pin1 warrants  further investigation as a potential cancer drug target.

overall_design: 6 samples, 3 treated with sulfopin, 3 control

first: Nir

last: London


title: STARTRAC analyses of scRNAseq data from a mouse tumor model reveal T cell dynamics and therapeutic targets in cancer

city: South San Francisco

first: Dev

last: Bhatt

address: 1120 Veterans Blvd

institute: Amgen

summary: To understand anti-tumor immune response, we performed single cell RNA-Seq and paired single cell TCR sequencing on T cells derived from multiple tissues of MC38-tumor-bearing mice

overall_design: Paired single cell RNA and TCR-sequencing of T cells

first: Xin


title: Olfactory expression of trace amine-associated receptors requires cooperative cis-acting enhancers

city: Evanston

first: Thomas

last: Bozza

address: 2200 Campus Drive

summary: Our study shows reduced expression of Trace-Amine Associated Receptors (TAARs) in the olfactory epithelium of mice in which Taar elements 1 and 2 (TE1+2) are deleted in cis.

overall_design: Olfactory epithelium RNAseq profiles of p30 T element deletion and control mice

first: Shah

last: Ami

first: Ratkowski

last: Madison

first: Rosa

last: Alessandro

first: Feinstein

last: Paul

first: Bozza

last: Thomas


title: Selective expansion of regulatory T cells using an orthogonal IL-2/IL-2 receptor system facilitates transplantation tolerance

city: Bethesda

first: Peng

last: Li

state: MD

address: 9000 Rockville Pike

department: NHLBI

institute: NIH

summary: Adoptive transfer of regulatory T cells (Tregs) has been shown to improve alloengraftment in animal models. However, it is technically challenging to expand Tregs ex vivo for the purpose of infusing large numbers of cells in the clinic. We demonstrated an innovative approach to engineer an orthogonal IL-2-IL-2 receptor (IL-2R) pair that selectively interacts with each other and transmits native IL-2 signals, but does not interact with the natural IL-2 or IL-2R counterparts, thereby enabling selective stimulation of target cells in vivo. Here, we introduced this orthogonal IL-2R into Tregs. Upon adoptive transfer in a murine mixed hematopoietic chimerism model, orthogonal IL-2 injection significantly promoted orthogonal IL-2R+Foxp3GFP+CD4+ cell proliferation without increasing other T cell subsets, and facilitated donor hematopoietic cell engraftment followed by acceptance of heart allografts. Our data indicate that selective IL-2 stimulation enabled by cytokine receptor engineering improves Treg potential for the induction of organ transplantation tolerance.

overall_design: RNA-Seq analyses using regulatory T cells (Tregs) that were stimulated with either no cytokine, WT IL-2, or ortho IL-2.

first: Toshihito

last: Hirai

first: Teresa

last: Ramos

first: Po-Yu

last: Lin

first: Federico

last: Simonetta

first: Leon

last: Su

first: Lora

last: Picton

first: Jeanette

last: Baker

first: Jian-Xin

first: Kinya

last: Seo

first: Juliane

last: Lohmeyer

first: Sara

last: Wagers

first: Melissa

last: Mavers

first: Warren

middle: J

last: Leonard

first: Bruce

middle: R

last: Blazar

last: Garica

first: Robert

last: Negrin


title: Cell cycle heterogeneity directs spontaneous 2C state entry and exit in mouse embryonic stem cells (ChIP-Seq)

city: Hangzhou

first: Jin

last: Zhang

state: Zhejiang

address: yuhangtang road

institute: Zhejiang University

summary: Mouse embryonic stem cells (ESCs) show cell-to-cell heterogeneity. A small fraction of 2-cell-like cells (2CLCs) marked by endogenous retrovirus activation emerge spontaneously. The 2CLCs are instable and they transit back to the pluripotent state without extrinsic stimulus. To understand how this bidirectional transition takes place, we performed single-cell RNA-seq on isolated 2CLCs which underwent 2C-like state exit and re-entry, and revealed a novel circular transitional process between 2C-like and pluripotent states. Mechanistically, we found that the cell cycle played an important role in mediating these transitions by regulating assembly of nucleolus and peri-nucleolar heterochromatin to influence 2C gene Dux expression. Collectively, our findings provide a roadmap of the 2C-like state entry and exit in ESCs and also a causal role of the cell cycle in promoting these transitions.

overall_design: Examination the binding chhrmatin of 2 combined different histone modifications in mESCs.

first: Yuqing

last: Zhu

first: Chen

last: Cheng

first: Lang

last: Chen


title: Cardiac cell type-specific gene regulatory programs and disease risk association [RNA-seq]

city: La Jolla

first: Sebastian

last: Preissl

address: 9500 Gilman Drive

institute: UCSD

summary: Misregulated gene expression in human hearts can result in cardiovascular diseases that are leading causes of morbidity and mortality worldwide. However, the limited information on the genomic location of candidate cis-regulatory elements (cCRE) such as enhancers and promoters in distinct cardiac cell types has restricted the understanding of these diseases. Here, we defined >287,000 cCREs in the four chambers of the human heart at single-cell resolution, which revealed cCREs and candidate transcription factors associated with cardiac cell types in a region-dependent manner and during heart failure. We further discovered cardiovascular disease-associated genetic variants enriched within these cCREs including 38 candidate causal atrial fibrillation variants localized to cardiomyocyte cCREs. Additional functional studies revealed that two of these variants affect a cCRE controlling KCNH2/HERG expression and action potential repolarization. Overall, this comprehensive atlas of human cardiac cCREs provides the foundation for illuminating cell type-specific gene regulation in human hearts during health and disease.

overall_design:
We performed single nucleus ATAC-seq to define a comprehensive catalogue of candidate cis-regulatory elements (cCREs) for the cell types in four regions of non-failing human hearts and generate in parallel snRNA-seq datasets for a subset to delineate gene expression patterns. We used the cCRE catalogue to computationally deconvolute dynamic enhancers in failing hearts and to assign cardiovascular disease risk variants to cCREs in individual cardiac cell types. Finally, we applied reporter assays, genome editing and electrophysiogical measurements in in vitro differentiated human cardiomyocytes to validate the molecular mechanisms of cardiovascular disease risk variants.

The raw sequencing data are available through dbGaP (controlled access): phs002204.v1.p1

first: James

last: Hocker

first: Olivier

middle: B

last: Poirion

first: Fugui

first: Justin

last: Buchanan

first: Kai

first: Joshua

last: Chiou

first: Tsui-Min

last: Wang

first: Xiaomeng

last: Hou

first: Qingquan

first: Yang

last: Li

first: Yanxiao

first: Elie

middle: N

last: Farah

first: Allen

first: Andrew

last: McCulloch

first: Kyle

last: Gaulton

first: Bing

last: Ren

first: Neil

last: Chi


title: Cell cycle heterogeneity directs spontaneous 2C state entry and exit in mouse embryonic stem cells (RNA-Seq)

first: Jin

last: Zhang

overall_design: Examine of control and cytarabine treatment gene expression

first: Yuqing

first: Chen

first: Lang


title: c-Myc facilitates prediction of "7+3" induction failure in de novo acute myeloid leukaemia with high-risk cytogenetics

city: Taichung City

first: thh

last: lab

address: 1650 Taiwan Boulevard Sect. 4, Taichung, Taiwan 40705, ROC

institute: TCVGH

summary: This study aimed to identify whether c-Myc could facilitate prediction of "7+3" induction failure in de novo acute myeloid leukaemia (AML) with high-risk cytogenetics.  Seventy-five untreated de novo AML patients who completed the "7+3" induction therapy were enrolled (24 patients from a prospective cohort; 51 patients from a retrospective cohort) and stratified into complete remission (CR) and non-CR groups.  Myc-associated molecular signature between the CR and non-CR groups in the prospective cohort was compared after gene set enrichment analysis.  c-Myc protein expression was assessed via immunohistochemical staining.  A high c-Myc-immunopositivity was defined when ≥ 40% of bone marrow myeloblasts were c-Myc (+).  Our results revealed that AML patients who did not achieve CR by the "7+3" induction therapy had a higher Myc gene expression than those that achieved CR (p=0.047).  Myc gene expression was positively correlated with c-Myc protein expression (p=0.014).   Although the non-CR group did not have more c-Myc protein expression compared to that in the CR group (p=0.151), combination of high c-Myc-immunopositivity and high-risk cytogenetics increased the probability of "7+3" induction failure compared to that in high-risk cytogenetics alone.  Induction strategies other than “7+3” might be a solution for de novo patients with high c-Myc-immunopositivity and high-risk cytogenetics.

overall_design: The 24 patients with de novo AML were stratified into CR (n = 15) and non-CR (n = 9) groups according to their treatment response to the first cycle of induction therapy.

first: Chieh-Lin

last: Teng

first: Tzu-Hung

last: Hsiao

first: Ren-Ching

first: Tsai-Jung

last: Lu


title: Genome-wide maps of Cap signal in control and Tet3 inactivated lung smooth muscle cells

city: Bad Nauheim

first: Mario

last: Looso

address: Ludwigstrasse 43

institute: Max Planck Institute for Heart and Lung Research

summary: To examine the initiation cap signal after Tet3 inactivation, we performed the Cap analysis gene expression (CAGE) sequencing with total RNA from control and Tet3 inactivated lung smooth muscle cells.

overall_design: Examination of Cap signal from control and Tet3 inactivated mice.

first: Wu

last: Fan

first: Liu

last: Hang


title: A Thrombin-PAR1/2 Feedback Loop Amplifies Thrombo-Inflammatory Endothelial Responses To The Viral RNA Analogue Poly(I:C)

city: Milwaukee

first: Robert

last: Burns

state: Wisconsin

address: 8727 W Watertown Plank Rd

institute: Blood Research Institute, Versiti

summary: This study describes a positive feedback mechanism by which thrombin sustains and amplifies the prothrombotic and proinflammatory function of endothelial cells exposed to the viral RNA analogue, poly(I:C) via activation of PAR1/2.

overall_design: Triplicate cultures of HUVEC were treated for 3h with poly(I:C) and/or thrombin and subjected to transcriptional profiling

first: Saravanan

last: Subramaniam

first: Hartmut

last: Weiler


title: Transcriptional profiling of brain tissues from a Q130 Long Evans Knock-in rat model of Huntington’s disease at 10-, 19- and 24-months of age

city: Princeton

first: Jeff

last: Aaronson

state: New Jersey

address: 155 Village Boulevard, Suite 200

department: Discovery & Systems Biology

institute: CHDI

summary: Huntington's disease (HD) is an autosomal dominant neurodegenerative disorder that is characterized by motor, cognitive, and psychiatric alterations. The mutation responsible for this disease is an abnormally expanded and unstable CAG repeat within the coding region of the gene encoding huntingtin (Htt). A collaboration between the CHDI Foundation and Dr. H.P. Nguyen (huu.nguyen-r7w@ruhr-uni-bochum.de) at Universitat Bochum focused on generating a knock-in rat model of HD that contains expanded CAG repeats inserted within the rat huntingtin gene (Hdh) in order to provide a genetic reconstruction of the human causative mutation within the rat model. The goal of this study is RNA expression profiling by RNA sequencing (RNA-seq) in brain tissues of 10-, 19- and 24-month-old heterozygous knock-in rats with uninterrupted CAG length approaching 130 along with littermate control wild-type animals.

overall_design: Transcriptomic analysis (RNASeq) was performed on samples from striatum, cortex and cerebellum collected from wild type and heterozygous Q130KI rats at 10, 19, and 24 months; at 10 and 19 months, the number of replicates is 5 males/5 females for age/genotype/tissue combinations and at 24 months, the number of replicates is 2-4 males/2-3 females. Twelve samples from GSE152443 (striatum samples from 2 and 6 months animals, male only, 3 animals for each timepoint/genotype) were added as pipeline controls to monitor for reproducibility and batch effect.

first: Jim

last: Rosinski


title: Transcriptomes of splenocytes from 16-week-old lupus-prone female NZB/NZW F1 mice and non-lupus-prone male NZB/NZW F1 and female BALB/c mice

city: Louisville

first: Michele

middle: M.

last: Kosiewicz

state: Kentucky

address: 505 S. Hancock St., Room 609

department: Microbiology & Immunology

institute: University of Louisville

summary: Transcriptomic analysis of gene expression of splenocytes from 16-week-old female NZB/NZW F1 (BWF1) mice, which are prone to lupus, and comparison to gene expression of splenocytes from male BWF1 and female BALB/c mice, which are not prone to lupus. Results provide insight into the differences in immunological activities that make female BWF1 mice more susceptible to lupus.

overall_design: RNA-seq data of RNA extracted from whole splenocytes.  Spleens were taken from 16-week-old  female and male BWF1 and female BALB/c mice (n=3)

middle: M

first: James

last: Harder


title: Microarrays of 4-month-old sugarcane collected in winter

city: São Paulo

first: Maira

last: Marins Dourado

state: São Paulo

address: Avenida Professor Lineu Prestes, 748, bloco 9, sala 951

department: Biochemistry

country: Brazil

institute: Institute of Chemistry, University of São Paulo

summary: Microarrays using as samples leaves +1 of 4-month-old sugarcanes collected in winter. The 12 samples are in duplicates, collected every 2 h for 22h, starting 1.5 h before dawn. The microarray experiments are a time point against an equimolar pool of all samples, using as crossreference a dye-swap of the biological replicate collected at the same time point.

overall_design: The leaves +1 of 4-month-old sugarcanes were collected in winter every 2h for 22h, starting 1,5h before dawn. Each sample is a mix of three individuals. Hybridizations were done using a custom 4x44 k oligoarray (Agilent Technologies) and samples were labeled using Low Input Quick Amp Labelling protocol of the Two-Color Microarray-Based Gene Expression Analysis system (Agilent Technologies). Hybridizations were done in duplicates for each time point against an equimolar pool of all samples. The duplicates are a dye-swap of a biological replicate of the same time point.

first: Dourado

last: Maira

first: Dantas

last: LuÃza

first: Hotta

middle: T

last: Carlos


title: The gut microbiota modulates environmental risk for cognitive impairment

city: Los Angeles

first: Christine

last: Olson

state: CA

address: 609 Charles E Young Dr

department: Integrative Biology and Physiology

institute: UCLA

summary: Cognitive impairment (CI) is a prevalent neurological condition characterized deficient attention, causal reasoning, learning and/or memory. Many genetic and environmental factors increase risk for CI, and the gut microbiome is increasingly implicated. However, the identity of gut microbes associated with CI risk, their effects on CI, and their mechanisms of action remain unclear. Here we examine the gut microbiome in response to restricted diet and intermittent hypoxia, known environmental risk factors for CI. Modeling the environmental factors together in mice potentiates CI and alters the gut microbiota. Depleting the microbiome by antibiotic treatment or germ-free rearing prevents the adverse effects of environmental risk on CI, whereas transplantation of the risk-associated microbiome into naïve mice confers CI. Parallel sequencing and gnotobiotic approaches identify the pathobiont Bilophila wadsworthia as enriched by the environmental risk factors for CI and as sufficient to induce CI. Consistent with CI-related behavioral abnormalities, B. wadsworthia and the risk-associated microbiome disrupt hippocampal activity, neurogenesis and gene expression. The CI induced by B. wadsworthia and by environmental risk factors is associated with microbiome-dependent increases in intestinal IFNy-producing Th1 cells. Inhibiting Th1 cells abrogates the adverse effects of both B. wadsworthia and environmental risk factors on CI. Together, these findings identify select gut bacteria that contribute to environmental risk for CI in mice by promoting inflammation and hippocampal dysfunction.

overall_design:
For each group, n=6 was dissected and analyzed as biological replicates. Hippocampi were microdissected and RNA extracted using the Qiagen RNEasy Mini Kit. RNA quality was assessed to be RIN>8.9 using the 4200 Tapestation (Agilent). RNA libraries were prepared using the QuantSeq FWD’ mRNA-Seq Library Prep Kit (Lexogen) and sequenced via the Illumina HiSeq platform by the UCLA Neuroscience Genomics Core. Sequences were filtered using FastQC v. 0.11.9 (Andrews, 2010) for quality control, followed by Trimmomatic (Bolger et al.)to remove barcodes and reads with an average phred score of 33 (parameters: illuminaclip:2:30:6, slidingwindow:5:30, leading:30, trailing:30, crop:65, minlen:20). Parsed reads were then aligned to the mouse genome mm10 using HISAT2 (Kim et al.; Kim et al.). Read counts were obtained using HTSeq-count (Anders et al.). Differential gene expression was determined using DESeq2 (Love et al.). Heatmaps were constructed using the R package (Team, 2013) pheatmap (Kolde, 2015), GO term enrichment analysis was conducted using DAVID (Huang da et al., 2009a, b) and Protein-Protein network analysis using STRING (Szklarczyk et al., 2019).

Definitions:
KD = KD diet is a ketogenic 6:1 fat to carbohydrate ratio diet from Harlan Teklad (TD.1150300).
"SPF, then Abx" = mice are raised specific pathogen free (SPF), and then treated with a heavy dosage of antibiotics (Abx) to deplete the gut microbiota.

first: Alonso

last: Iñiguez

first: Elaine


title: Single cell RNAseq of mouse CD3+ Adipose Tissue T cells

city: Ann Arbor

first: Carey

last: Lumeng

state: MI

address: 109 Zina Pitcher Place

department: Pediatrics

institute: University of Michigan

summary: Despite studies implicating adipose tissue T cells (ATT) in the initiation and persistence of adipose tissue inflammation, fundamental gaps in knowledge regarding ATT function impedes progress towards understanding how obesity influences adaptive immunity. We hypothesized ATT activation and function would have tissue-resident specific properties and that obesity would potentiate their inflammatory properties. We assessed ATT activation and inflammatory potential within mouse  stromal vascular fraction (SVF).To assess intracellular signaling mechanisms responsible for ATT inflammation impairments, single-cell RNA sequencing of ATTs from epididymal adipose tissue was performed from male chow and high fat diet (60% for 12 weeks) fed mice.

overall_design: FACS sorted CD3+ cells from epididymal adipose tissue from male lean (n=2) and HFD fed (n=1) C57Bl/6 mice were analyzed by single cell RNA-seq (10X genomics)

first: Cara

last: Porsche