title: Single Nucleus Sequencing Reveals Artifactual Gene Expression and Variance in Post-Mortem Brain

summary: Single nucleus sequencing of fresh frozen post-mortem brain

overall_design: 3 Donors, 2 replicates per donor


title: Single Cell Sequencing Reveals Artifactual Gene Expression in Microglia in Response to Tissue Dissociation

status: Public on Dec 01 2020

city: Boston

first: Beth

last: Stevens

state: Massachusetts

address: 3 Blackfan Circle, 12220 Center for Life Sciences

country: USA

institute: Boston Children's Hospital

type: Expression profiling by high throughput sequencing

summary: Single Cell Sequencing of FACS purified microglia/myeloid cells isolated with different dissociation protocols

overall_design: 12 animals, 3 replicates per isolation protocol, P90

first: Samuel

middle: E

last: Marsh


title: Single Cell Sequencing Reveals Artifactual Gene Expression Following Dissociation in All CNS Cells

summary: Single Cell Sequencing of all CNS cell types via enzymatic digestion with or without inhibitor cocktail present.

overall_design: 4 animals, 2 replicates per isolation protocol, P90


title: Arabidopsis Response to Abscisic Acid Regulate RNA NAD+-Capping

status: Public on Nov 23 2020

city: Shanghai

first: Xiang

last: Yu

state: Shanghai

address: 800 DongChuan

country: China

institute: Shanghai Jiao Tong University

type: Non-coding RNA profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Transcriptomic profiling of neonatal mouse granulosa cells reveals new insights into primordial follicle activation

status: Public on Dec 10 2020

city: Callaghan

first: Jessie

middle: M

last: Sutherland

state: New South Wales

address: Life Sciences Building, Ring Road

department: School of Biomedical Science and Pharmacy

country: Australia

institute: University of Newcastle

summary: The dormant population of ovarian primordial follicles is determined at birth and serves as the reservoir for future female fertility. Of equal importance to fertility is the rate that primordial follicles activate and enter folliculogenesis, yet our understanding of the molecular, biochemical, and cellular processes underpinning primordial follicle activation remains limited. The survival of primordial follicles relies on the correct complement and morphology of granulosa cells, which provide signalling factors essential for oocyte and follicular survival. To investigate the contribution of granulosa cells in the primordial-to-primary follicle transition, ovaries from C57Bl/6 mice were enzymatically dissociated at two time points spanning the initial wave of primordial follicle activation. Post-natal day (PND) 1 ovaries yielded primordial granulosa cells, and PND4 ovaries yielded a mixed population of both primordial and primary granulosa cells. The comparative transcriptome of granulosa cells at these time points was generated via the Illumina NextSeq 500 system which identified 132 significantly differentially expressed transcripts. This transcriptomic dataset confirmed the expression of factors known to be involved in primordial follicle activation belonging to TGF-B and EIF4E signalling pathways, as well as novel factors linked to pathways involved in primordial follicle activation (ZFX, POD1, FRZB). This study highlights the dynamic changes in gene expression of granulosa cells during primordial follicle activation and provides evidence for a renewed focus into the Wnt signalling pathway’s role in primordial follicle activation.

overall_design: mRNA profiles of granulosa cells dissociated from postnatal day 1 and postnatal day 4 mouse ovaries

first: Emmalee

middle: A

last: Ford

first: Emily

middle: R

last: Frost

first: Emma

middle: L

last: Beckett

first: Shaun

middle: D

last: Roman

first: Eileen

last: McLaughlin


title: Inhibiting collagen I production and tumor cell colonization in the lung via miR-29a-3p loading of exosome-/liposome-based nanovesicles

status: Public on Nov 20 2020

city: Zhengzhou

first: Yan

last: Yan

address: Jianshe Road

institute: The First Affiliated Hospital of Zhengzhou University

summary: The lung is one of the most common sites for cancer metastasis. Collagens in the lung provide a permissive microenvironment that supports the colonization and outgrowth of disseminated tumor cells. Therefore, down-regulating the production of collagens may contribute to the inhibition of lung metastasis. It has been suggested that miR-29 exhibits effective anti-fibrotic activity by negatively regulating the expression of collagens. Indeed, our clinical lung tumor data shows that miR-29a-3p expression negatively correlates with collagen I expression in lung tumors and positively correlates with patients’ outcomes. However, suitable carriers need to be selected to deliver this therapeutic miRNA to the lungs. In this study, we found that the chemotherapy drug cisplatin facilitated miR-29a-3p accumulation in the exosomes of lung tumor cells, and this type of exosomes exhibited a specific lung-targeting effect and promising collagen down-regulation. To scale up the preparation and simplify the delivery system, we designed a lung-targeting liposomal nanovesicle (by adjusting the molar ratio of DOTAP/cholesterol–miRNAs to 4:1) to carry miR-29a-3p and mimic the exosomes. This liposomal nanovesicle delivery system significantly down-regulated collagen I secretion by lung fibroblasts in vivo, thus alleviating the establishment of a pro- metastatic environment for circulating lung tumor cells.

overall_design: Cisplatin elicited LLC exosomes (CPT-exo) were isolated from culture supernatants of LLC treated with 5uM cisplatin. Microarray analysis was performed to generate miRNA profiles of CPT-exo. Two independent experiments were performed.

first: Ming

last: Wang

first: Zhihai

last: Qin


title: TRACE-seq: “TRanslatomic” Analysis Captured in Extracellular vesicles using sequencing

status: Public on Dec 02 2020

city: BOSTON

first: Francois

last: Cherbonneau

state: MA

address: 185 CAMBRIDGE STREET, CPZN3200

institute: Massachusetts General Hospital

summary: Purpose: We develop a new methodology to monitor live cells in vitro as well as in vivo. Method: Each population of HEK 293T cells was transfected with the TRACE-seq construct (GFP-C-YTHDF1/GBP1-CD9) and a negative control construct (GFP-C-YTHDF1/GBP1-CD9) which contains no mRNA catcher. The analysis was made in duplicate with microvesicles (MVs) mRNA content isolated from cell culture media and their corresponding cell lysate. Samples were generated with the SMART-seq2 protocol and the Nextera kit (Illumina). Data were collected using 50 × 8 × 50 reads on a HiSeq. Reads were aligned to hg19 using STAR and counting reads associated genes were detected with the FeatureCounts module. Also, a QC qPCR was made on these samples and the cDNA library quality was analyzed by a High sensitivity DNA chip assay. Results: From the 3604 genes selected for this analysis, RNA-seq data showed a non negligeable increase of the correslation between MVs mRNA content and cell lysate from the HEK293T transfected with the TRACE construct. Conclusion: Our methodology bring a representative part of the transcriptome from the cell into MVs which is a usefull method for in vivo analysis. Moreover, our study represents the first detailed analysis of a new undestructive, fully compatible for  in vivo, monitoring process of the cells transcriptome.

overall_design: HEK 293T cells transfected with two construct: TRACE one (GFP-C-YTHDF1/GBP1-CD9) and the negatif control construct (GFP/GBP1-CD9). Analyses made on microvesicles from the cells media and the own cells lysate of each samples.

first: Robert

last: Kitchen


title: Innate sensor Dhx9 orchestrates TCR signaling and controls CD8 T cell immunity against viral infection [ATAC-Seq]

status: Public on Nov 19 2020

city: Xi’an

first: Anjun

last: Jiao

state: Shaanxi

address: No. 28, Xianning West Road, Xi'an City, Shaanxi Province

institute: Department of Pathogenic Microbiology and Immunology, School of Basic Medical Sciences, Xi’an Jiaotong University

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Nuclear sensor molecules of innate cells recognize pathogenic DNA or RNA to activate innate immunity and defense against bacteria and virus infection. Although some sensors exhibit their preliminary functions in T cells, it is still unclear about the underlying mechanism. Here we found a nuclear sensor-Dhx9, was enhanced its expression in effector CD8 T cells. T cell specific deletion of Dhx9 caused an impaired CD8 T cell expansion, memory formation and virus clearance. Mechanically, Dhx9 could directly regulate id2 transcription and in turn control effector CD8 T cell differentiation. Importantly, Dhx9 could bind to Lck protein, mediate Zap70 phosphorylation, and promoting TCR downstream signaling - ERK pathway to protect effector CD8 T cells from apoptosis. The DSRM and DEXDc-Helicase domain is required for Lck binding and Id2 transcription, respectively. Therefore, we revealed a novel regulatory mechanism that innate nuclear sensor regulates T cell fate and function through modulating adaptive pathways.

overall_design: Examination of wt and Dhx9 deficient CD8 T cells from the spleens 8 days post infection

first: Baojun

last: Zhang


title: Landscape of RNA NAD+ capping in Arabidopsis unopened flower bud

summary: Eukaryotic messenger RNAs (mRNAs) possess a 5’-end N7-methyl guanosine (m7G) cap that promotes their translation and stability. However, it was recently demonstrated that eukaryotic mRNAs can also carry a 5' end nicotinamide adenine dinucleotide (NAD+) cap that promotes mRNA decay mediated by the NAD+ decapping enzyme DXO1. However, the dynamic regulation of NAD+ capping in plant remains unknown. Here, we describe the global landscape of NAD+-capped RNAs in Arabidopsis thaliana, and demonstrate that DXO1 is responsible for removal of these 5’-end modifications and facilitates mRNA degradation in plant transcriptomes. We also reveal that in the absence of DXO1 NAD+-capped mRNAs are unstable and processed into smRNAs. Furthermore, we find that Abscisic Acid (ABA) remodel the landscape of RNA cap epitransciptome, and the mRNA lost their NAD+ cap contribute to their stability under ABA. Overall, our results support a link between ABA response and RNA NAD+ capping.

overall_design: NADseq, GMUCT and small RNA profiling in unopened flower bud of Arabidopsis thaliana

first: Lee

last: Vandivier

first: Matthew

last: Willmann

first: Brian

last: Gregory


title: Innate sensor Dhx9 directly binds to multiple genes related to effector CD8 T cell differentiation


title: Innate sensor Dhx9 directly binds to multiple genes related to effector CD8 T cell differentiation [ChIP-Seq]

summary: We provide a set of genes that are directly bound by Dhx9 in CD8 T cells from LCMV infected mice.

overall_design: Examination of binding peaks in genome of wt GP33+CD8 T cells from the spleens 8 days post infection


title: Expression data from P4H-TM deficient mouse cortexes after 24h of pMCAO

status: Public on Nov 12 2020

city: Oulu

first: Nadiya

last: Byts

address: Aapistie 7C

department: FBMM

country: Finland

institute: Biocenter Oulu

summary: Genetic deletion of transmembrane prolyl-4-hydroxylase (P4H-TM) in mice leads to increased inflammatory microgliosis and neutrophil infiltration in the cortex after permanent midle cerebral artery occlusion (pMCAO)

overall_design: Mice underwent pMCAO on a left side and 24h later cortexes from wild-type (WT) and P4H-TM deficient (KO) mice were collected for further mRNA extraction and microarray analysis. Corresponding contralateral cortex (c) was used as a control for inpsilateral injured (i) one. mRNA from 4 individual mice were pooled per experimental condition. Comparison was done between four experimental conditions: WTc, WTi, KOc, KOi.

first: Johanna

last: Myllyharju


title: Landscape of RNA NAD+ capping in Arabidopsis seedling with ABA treatment

first: Xiang

last: Yu

overall_design: NADseq, GMUCT and total RNAseq in Arabidopsis transcriptome with ABA treatment


title: Cortical transcriptomic changes in the newborn rat brain after exposure to surgery and anesthesia in the third trimester

status: Public on Nov 10 2020

city: St. Louis

first: Arvind

last: Palanisamy

state: MO

address: 660 S Euclid Ave

department: Anesthesiology

institute: Washington University School of Medicine

summary: We report changes in the cortical transcriptome of newborn rat pups after exposure to anesthesia and surgery in the third trimester using unbiased RNA-seq analysis.

overall_design: We performed RNA-seq analysis to examine differential gene expression in the cortex of newborn pups (day 0) after in utero exposure to anesthesia and subcutaneous pump implantation surgery on embryonic day 18  (n = 2 pups each of either sex from 3 independent dams)  or no treatment (2 pups each of either sex from 3 independent dams). With an FDR adjusted p < 0.05, none of the genes were determined to be significantly differentially expressed in the newborn brain after exposure to anesthesia and surgery in the third trimester.


title: ARID1A mutations promote P300-dependent endometrial invasion through super-enhancer hyperacetylation (diff_H3K4me3_H3K27me3_ChIP)

city: Grand Rapids

first: Ronald

middle: L.

last: Chandler

state: MI

department: Obstetrics, Gynecology and Reproductive Biology

institute: Michigan State University

summary: Endometriosis affects 1-in-10 women and is characterized by the presence of abnormal endometrium at ectopic sites. ARID1A mutations are observed in deeply invasive forms of the disease, often correlating with malignancy. To identify epigenetic dependencies driving invasion, we utilized an unbiased approach to map chromatin state transitions accompanying ARID1A loss in the endometrium. We show that super-enhancers marked by high H3K27-acetylation are strongly associated with ARID1A binding. ARID1A loss leads to H3K27-hyperacetylation and increased chromatin accessibility and enhancer RNA transcription at super-enhancers, but not typical enhancers, indicating that ARID1A normally prevents super-enhancer hyperactivation. ARID1A co-localizes with P300 at super-enhancers, and genetic or pharmacological inhibition of P300 in ARID1A-mutant endometrial epithelia suppresses invasion and induces anoikis through rescue of super-enhancer hyperacetylation. Among hyperactivated super-enhancers, SERPINE1 (PAI-1) was identified as an essential target gene driving ARID1A-mutant endometrial invasion. Broadly, our findings provide rationale for therapeutic strategies targeting super-enhancers in ARID1A-mutant endometrium.

overall_design: Human 12Z endometriotic epithelial cells were treated with lentiviral particles expressing shRNA targeting ARID1A (shARID1A) or non-targeting control (shNONtg) and assayed by chromatin immunoprecipitation (ChIP) for H3K4me1 followed by sequencing.

first: Mike

last: Wilson

first: Jake

middle: J

last: Reske


title: SWI/SNF complex mutations promote thyroid tumor progression and resistance to redifferentiation therapy (ATAC-seq, Fig2)

city: New York

first: Richard

last: Koche

state: New York

institute: Memorial Sloan Kettering Cancer Center

summary: Recurrent mutations in genes encoding subunits of the SWI/SNF chromatin remodeling complex occur commonly in human cancers of different lineages, including advanced thyroid cancers. Thyroid-specific loss of Arid1a, Arid2 or Smarcb1 in mouse BrafV600E-mutant tumors promotes disease progression and decreased survival, associated with lesion-specific effects on chromatin accessibility and differentiation. Swi/Snf loss leads to a repressive chromatin state at thyroid lineage transcription factors and their target binding sites, resulting in impaired expression of genes required for thyroid hormone biosynthesis. Thyroid differentiation and response to radioiodine therapy is restored by MAPK inhibitors in mouse BrafV600E-mutant thyroid cancers, which is abrogated in the context of Swi/Snf loss. MAPK inhibitors also failed to restore thyroid differentiation and radioiodide incorporation in patients with BRAFV600E or RAS-mutant tumors with SWI/SNF mutations. SWI/SNF complexes are central to the maintenance of differentiated thyroid function, and their loss confers RAI refractoriness and resistance to MAPK inhibitor-based redifferentiation therapies.

overall_design: ATAC-seq of SWI/SNF KO and WT thyroid tissue

first: Mahesh

last: Saqcena

middle: P

first: James

last: Fagin


title: ARID1A mutations promote P300-dependent endometrial invasion through super-enhancer hyperacetylation (diff_H3K4me1_ChIP)


title: ARID1A mutations promote P300-dependent endometrial invasion through super-enhancer hyperacetylation (diff_H3K27ac_CUTNRUN)

overall_design: Human 12Z endometriotic epithelial cells were transfected with a combination of siARID1A, siP300, both, or non-targeting siRNA control and 1 uM A-485 or 0.1% DMSO vehicle and assayed 72 hours post-transfection via Cleavage Under Targets and Release Using Nuclease (CUT&RUN).


title: ARID1A mutations promote P300-dependent endometrial invasion through super-enhancer hyperacetylation (diff_P300_ChIP)

overall_design: Human 12Z endometriotic epithelial cells were treated with lentiviral particles expressing shRNA targeting ARID1A (shARID1A) or non-targeting control (shNONtg) and assayed by chromatin immunoprecipitation (ChIP) for P300 followed by sequencing.


title: ARID1A mutations promote P300-dependent endometrial invasion through super-enhancer hyperacetylation (diff_H3Kac_ChIP)

overall_design: Human 12Z endometriotic epithelial cells were treated with lentiviral particles expressing shRNA targeting ARID1A (shARID1A) or non-targeting control (shNONtg) and assayed by chromatin immunoprecipitation (ChIP) for H3K27ac and H3K18ac followed by sequencing.


title: ARID1A mutations promote P300-dependent endometrial invasion through super-enhancer hyperacetylation


title: ARID1A mutations promote P300-dependent endometrial invasion through super-enhancer hyperacetylation (P300_ChIP)

overall_design: Wild-type Human 12Z endometriotic epithelial cells were assayed by chromatin immunoprecipitation (ChIP) for P300 followed by sequencing.


title: ARID1A mutations promote P300-dependent endometrial invasion through super-enhancer hyperacetylation (RNA)

overall_design: Human 12Z endometriotic epithelial cells were transfected with a combination of siARID1A, siP300, both, or non-targeting siRNA control and 100 nM A-485 or 0.1% DMSO vehicle and assayed 72 hours post-transfection via RNA-seq.


title: Atg7-dependent autophagy regulates the naive to primed transition by selective Nanog degradation in mouse embryonic stem cells

status: Public on Nov 07 2020

city: Wuhan

first: Jing-Jing

state: Hubei

address: 1 Lion Rock Street

department: Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction

institute: Huazhong Agricultural University

summary: Autophagy is a conserved cellular mechanism to degrade unwanted cytoplasmic proteins and organelles to recycle their components, and it is proved to be critical for embryonic stem cell (ESC) self-renewal and somatic cell reprogramming. However, the role of autophagy in embryonic development remains elusive, and no information exists regarding its functions during the transition from naive to primed pluripotency. Here by using an in vitro transition model of ESCs to epiblast-like cells (EpiLCs), we describe that the dynamic changes in Atg7-dependent autophagy is required for the naive to primed transition, and it is also necessary for germline specification. RNA-seq and ATAC-seq profiling reveal that Nanog acts as a barrier to prevent pluripotency transition, and autophagy-dependent Nanog degradation is important for dismantling the naive pluripotency expression program through decommissioning of naive-associated active enhancers. Mechanistically, we found that autophagy adaptor protein Sqstm1 (p62) is nucleus located during the pluripotency transition period and it is preferentially associated with ubiquitinated Nanog for selective protein degradation. In vivo, loss of autophagy by Atg7 depletion disrupts peri-implantation development and we observed increased chromatin association of Nanog, which affects neuronal differentiation through activation of a subset of neuroectodermal development-associated enhancers. Taken together, our findings illuminate regulatory mechanisms underlying the naive to primed transition and reveal that autophagy-dependent regulation of Nanog is essential for exit from the naive state and marks distinct cell fate allocation during lineage specification.

overall_design: The ChIP-Seq experiments of the Control and Atg7 KO groups were taken at each time point from the 4-day ESC to EpiLC transition.

first: Jilong

last: Zhou

first: Hainan

last: He

first: Jingjing

first: Yi-Liang

last: Miao


title: Effects of EGF, cetuximab, trastuzumab and afatinib on gene expression in gastric cancer cell lines

status: Public on Nov 02 2020

city: Braunschweig

first: Robert

last: Geffers

address: Inhoffenstr. 7

department: Dep. Molecular Bacteriology

country: Germany

institute: HCI - Helmholtz Centre for Infection Research

summary: The antibody trastuzumab in combination with platin-fluoropyrimidine chemotherapy is the standard of care for patients with HER2-positive recurrent or metastatic gastric cancer. Alternative treatment options and biomarkers for prediction of therapy response are needed since some patients do not respond to trastuzumab therapy or develop acquired resistance. We compared the molecular effects of trastuzumab and other HER-targeting drugs cetuximab and afatinib. We also tested the hypothesis that the treatment-dependent regulation of a gene indicates its importance for response, and therefore it may be used as biomarker for patient stratification.   The four gastric cancer cell lines Hs746T, MKN1, MKN7 and NCI-N87 were treated with EGF, cetuximab, trastuzumab or afatinib for 4 or 24 hours. The effect on gene expression was then measured by RNA sequencing and the resulting candidate biomarkers were tested in an available cohort of gastric cancer patients or were functionally analyzed in vitro.  Treatment with afatinib resulted in the highest number of regulated genes, followed by cetuximab and trastuzumab. Although trastuzumab had little effects on gene expression, we identified BMF, HAS2 and SHB as candidate biomarkers for trastuzumab response. HAS2 and SHB were confirmed as potential predictive markers for trastuzumab therapy response in clinical specimens. AREG, EREG and HBEGF were identified as candidate biomarkers for afatinib and cetuximab treatment. The functional analysis confirmed HBEGF as resistance factor for cetuximab.  By confirming HAS2, SHB and HBEGF as biomarkers for anti-HER therapies, we provide evidence that the regulation of gene expression after treatment can be used for biomarker discovery.

overall_design: Gastric cancer cell lines were treated with EGF, cetuximab, trastuzumab and afatinib in biological triplicates

first: R

first: K

last: Ebert

first: B

last: Luber

first: G

last: Zwingenberger

last: Arnold

first: S

last: Keller


title: Genome-wide maps of small RNA transcription in young and old mouse oocytes

status: Public on Nov 05 2020

city: Tzur Hadassa

first: Michael

last: Klutstein

state: Other

address: Yasmin 273 Tzur Hadassa

department: Institute of Dental Sciences

country: Israel

institute: Hebrew University

summary: We report the abundance of different small RNAs in oocytes of young (8 weeks old) and older (9 months old) mouse females

overall_design: Pools of 50 oocytes from young and older females were prepared for small RNA seq using the SMARTer® smRNA-Seq kit. The first batch was sequenced paired end, one sample 2X42 and one sample 2X38 bp, the second to fourth batch was sequenced single end 76 bp.

first: Peera

last: Wasserzug Pash

first: Rachel

last: Rothman


title: Nrl-dependent and independent rod photoreceptor development in zebrafish

status: Public on Oct 27 2020

first: Fei

last: Liu

address: No. 7 Donghu South Road, Wuchang District

institute: Institute of Hydrobiology, Chinese Academy of Sciences

overall_design: Refer to individual Series


title: Age influences aggressive behaviour: an RNA-seq analysis in bovines

status: Public on Oct 30 2020

city: madrid

first: Paulina

middle: G.

last: Eusebi

state: madrid

address: Ave. Puerta del hierro

department: Animal Production

country: Spain

institute: Universidad Complutense de Madrid

summary: Here we used a RNA-seq approach to investigate differential gene expression in the prefrontal cortex of bovines from the aggressive selected Lidia breed at different age stages: young three-year old and adult four-year-old bulls. The number of up and down-regulated genes in the adult group were 50 and 193, respectively. These genes were associated with diverse functions, including neural plasticity and behavior. A cross-species comparative analysis retrieved 29 genes in common with previous studies on aggressive behaviors. Particularly, we detected changes in the regulation of networks such as the synaptogenesis, involved in the synapses maintenance and refinement, and the glutamate receptor pathway, which acts as excitatory driver in aggression responses. Furthermore, the Wnt//β- catenin signaling pathway was present in both up and down-regulated genes in the adult group, and has been implicated in adult altered behaviors and cognitive disorders. At the regulatory level, we identified four upstream regulators, all involved in behavioral processes and neurophsychiatric disorders including aggressive demonstrations. Our results on an animal model in which the aging process enhance the intensity of aggressive responses provide, first- time ever, insights into candidate genes and networks implicated in the molecular mechanisms leading to the maturation of the cattle brain, with a focus on agonistic components.

overall_design: 16 non-castrated male Lidia bovines 16 non-castrated bovine males of the Lidia breed, eight three-year-old and eight four-year-old, for the young and adult groups respectively

middle: G

first: Natalia

last: Sevane

first: Manuel

last: Pizarro

first: Javier

last: Cañón

first: Susana

last: Dunner


title: Nrl-dependent and independent rod photoreceptor development in zebrafish [RNA-Seq]

summary: NRL (Neural retina leucine zipper) is a key regulator of the fate determination and gene expression of rod photoreceptors in the retina of many vertebrates. In this study, we observed a unique retinal phenotype in the nrl knockout zebrafish model characterized by reduced rods with shortened outer segments and gradually increased green-cones in adulthood. By tracing and comparing the developmental processes of WT and nrl knockout rods, we found there might be two waves of rod genesis distinguished as nrl-dependent and independent with different starting times. To reveal the underlying cellular and molecular mechanisms, bulk and single-cell RNA-seq were performed. The changes in gene expression and cell proportion of rods and cones agreed well with our histological and immunofluorescence studies. Interestingly, we found that rods exhibited noticeable heterogeneities in the gene expression patterns, and a part of rods in nrl knockout zebrafish misexpressed the green-cone genes, reflecting a hybrid status of rod and green-cone. Furthermore, we identified mafba as a novel regulator of rod genesis, which was responsible for the development of rods in nrl knockout zebrafish. Our study will largely improve the current understanding of the developmental processes and regulatory mechanisms of rods in zebrafish and probably other species, and may facilitate future studies in the fields of retinal development and retinal degeneration.

overall_design: Two WT and two nrl-KO samples (two biological replicates) were analyzed. Each sample represented a pool of RNAs from three zebrafish retinas at the age of 2 months. The differentially expressed genes between WT and nrl-KO groups were identified.

first: Yayun

last: Qin

first: Yuwen

last: Huang


title: Resistance to the herbicide 2,4-D in Sisymbrium orientale conferred by a deletion mutation in the degron tail of IAA2

status: Public on Oct 08 2020

city: Fort Collins

first: Todd

last: Gaines

state: CO

address: 1177 Campus Delivery

department: Agricultural Biology

institute: Colorado State University

summary: Resistance to the phenoxy synthetic auxin herbicide 2,4-D is conferred by a deletion mutation in the degron tail of IAA2. An RNA-Seq experiment was used to assemble a reference transcriptome for the weed species Sisymbrium orientale, to quantify gene expression, and to identify sequence variants in the alignments. A 27 bp deletion was identified in the IAA2 gene. Forward genetics supported the role of this deletion mutation in resistance. The IAA2 deletion allele was transformed into Arabidopsis thaliana to confirm that it conferred 2,4-D resistance.

overall_design: Young leaves were sampled for RNA extraction and preparation of Illumina Hi-Seq libraries. Samples were collected only from untreated plants.

first: Marcelo

last: Figueiredo

first: Anita

last: Kuepper

first: Christopher

last: Preston

first: Eric

last: Patterson


title: Zebrafish as a model to understand the impact of inactivity and neuromuscular electrical stimulation on neuromuscular plasticity in Duchenne Muscular Dystrophy

status: Public on Oct 01 2020

city: Orono

first: Benjamin

last: King

state: ME

address: 5735 Hitchner Hall

department: Molecular and Biomedical Sciences

institute: University of Maine

summary: Skeletal muscle plasticity is imperative for functional adaptation to changing demands. Although a great deal is known about the structural and functional plasticity of healthy skeletal muscle, far less is known about plasticity in diseased muscle. Here, we created an experimental design to better understand how dystrophin-deficient zebrafish muscle responds to inactivity versus activity. We tested two inactivity paradigms, intermittent versus extended inactivity, and found that extended inactivity, defined as 72 hours in low-dose tricaine, worsens muscle structure and decreases survival. Strikingly, extended inactivity followed by a single session of neuromuscular electrical stimulation (NMES) further worsens muscle structure and obliterates its ability to regenerate. Next, we tested four unique NMES paradigms that varied in their frequency and voltage, and found that each NMES paradigm differentially affected muscle structure, function, and survival. However, one NMES paradigm, which we refer to as endurance NMES, improved all three components of muscle health. Accompanying these improvements, we observed increased sarcomere lengths as well as larger surface areas, volumes, and filament indices of muscle nuclei. Additionally, time-lapse imaging reveals less degeneration and improved regeneration of the fast-twitch muscle fibers. RNAseq analysis highlights two potential mechanisms, one in which gene expression levels are returned to wild-type levels as well as remodeling of the extracellular matrix. Collectively, these data provide a new methodology with which to study neuromuscular plasticity in healthy versus diseased muscle. In addition, our data clearly indicate that, at least in the zebrafish model, NMES is beneficial whereas inactivity is deleterious for dystrophic muscle.

overall_design: Gene expression was assayed using RNA sequencing in dmd mutant and wild-type sibling control whole zebrafish larvae at 7 days post fertilization with and without neuromuscular electrical stimultation (NMES). Each biological replicate consisted of two zebrafish larvae. For wild-type siblings, there were 4 biological replicates for the control group and 3 biological replicates for the NMES group. For the dmd mutants, there were 8 biological replicates in the dmd mutant control group, and 10 biological replicates in the dmd NMES group.

first: Elisabeth

last: Kilroy

first: Kaylee

last: Brann

first: Claire

last: Schaffer

first: Davon

last: Varney

first: Kodey

last: Silknitter

first: Jordan

middle: N

last: Miner

first: Ahmed

last: Almaghasilah

first: Tashawna

last: Spellen

first: Alexandra

last: Lewis

first: Karissa

last: Tilbury

first: Joshua

middle: B

last: Kelley

first: Clarissa

last: Henry


title: Effects of intracellular amyloid toxicity on the transcriptome of MC65 human nerve cells

status: Public on Oct 22 2020

city: San Diego

first: April

middle: Elizabeth

last: Williams

state: California

address: 10010 N Torrey Pines Rd

department: IGC

institute: Salk Institute for Biological Studies

summary: Amyloid beta (Aβ) accumulates within neurons in the brains of early stage Alzheimer's disease (AD) patients. However, the mechanism underlying its potential toxicity remains unclear. To elucidate this, a transcriptomic study was carried out using a nerve cell model of toxic intracellular aggregation of Aβ. The neuroprotective compound CMS121 was included as a validating tool. It was found that intracellular Aβ induces profound changes in the omics landscape of nerve cells that are associated with a cancer-like metabolic reprogramming, the oxytosis/ferroptosis cell death program, and physiological alterations detrimental to mitochondrial function. Our findings have implications for the understanding of the basic biology of proteotoxicity, aging and AD as well as for the development of future therapeutic interventions designed to target the oxytosis/ferroptosis programmed cell death pathway in the AD brain.

overall_design: We carried out a study of the whole transcriptome of human MC65 nerve cells in conditions of non-induced and induced Aβ toxicity in the absence and presence of 1uM CMS121. The MC65 is a human nerve cell line that contains the C99 fragment of the amyloid precursor protein (APP) under the control of a tetracycline-sensitive promoter. The parent cell line is SK-N-MC from a human neuroblastoma, and it has an electrically excitable membrane typical of neurons. When tetracycline is withdrawn, MC65 nerve cells express C99 that is converted to Aβ by γ-secretase and the cells die within 3 days due to Aβ accumulation and aggregation within cells. In our study, cells were harvested at day 2 after Aβ induction, prior to any cell death, and the whole RNA was isolated and sequenced.

first: Antonio

last: Currais

first: Ling

last: Huang


title: Nrl-dependent and independent rod photoreceptor development in zebrafish [scRNA-Seq]

first: Fei

last: Liu

overall_design: The retinal single-cell suspensions for WT and nrl-KO zebrafish were prepared. Each suspension represented a pool of retinal cells from three retinas at the age of 5 months. scRNA-seq was performed using the 10X Genomics technology. The retinal cell clusters and the corresponding gene expression patterns were identified.

first: Yayun

last: Qin

first: Pan

last: Gao


title: Estrogen receptor beta repurposes EZH2 to suppress oncogenic NFκB signaling in triple negative breast cancer

status: Public on Sep 30 2020

city: Rochester

first: Aditya

last: Bhagwate

state: MN

address: 200 1st Street SW

institute: Mayo Clinic

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


title: Triple-Sequencing of dendritic cells coinfected with Aspergillus fumigatus and cytomegalovirus

city: Jena

first: Bastian

last: Seelbinder

address: Adolf-Reichwein-Straße 23

department: Systems Biology and Bioinformatics

institute: Hans-Knoell-Institute Jena

summary: Human dendritic cells were coinfected with Aspergillus fumigatus and human cytomegalovirus. Single-cultures or single-infections served as controls. RNA was isolated and processed for next generation sequencing.

sample_organism:
Homo sapiens
Human betaherpesvirus 5
Aspergillus fumigatus

overall_design: 4 replicates per condition (DC alone, Afu alone, DC + Afu, DC + CMV, DC + Afu + CMV)

first: Julia

last: Wallstabe

first: Jürgen

last: Löffler

first: Sascha

last: Schäuble


title: Portal fibroblasts with mesenchymal stem cell features form a reservoir of proliferative myofibroblasts in liver fibrosis

status: Public on Oct 21 2020

city: Oslo

first: Trine

last: Folseraas

state: Oslo

address: Sognsvannsveien 7

department: Transplantation Medicine

country: Norway

institute: Oslo University Hospital Rikshospitalet

summary: Based on the identification  of a transcriptomic signature, including Slit2,  characterizing portal mesenchymal stem cells  (PMSC)  and derived myofibroblast (MF), we examined  the gene expression profile of in liver tissue derived  from multiple human liver disorders, including  primary sclerosing cholangitis (PSC) (n=12), non-alcoholic steatohepatitis (NASH)  (n=7) and other liver diseases (i.e., primary biliary cholangitis, autoimmune hepatitis, alcoholic liver disease and haemochromatosis) (n=8) and  compared them to healthy controls (tumor free tissue from livers with metastasis from colorectal cancer) (n=5). We found that  SLIT2 was overexpressed in the liver of patients with NASH, PSC and other chronic liver diseases. We also examined the microarray data of the human liver tissue samples for the transcriptomic signatures and found that in the different types of liver diseases the gene signature of PMSCs/PMSC-MFs was increased compared to normal liver, and correlated with the expression of ACTA2, COL1A1 and vWF.

overall_design: The RNA used for the microarray experiments was extracted from  fresh frozen tissue obtained from explanted livers or diagnostic liver biopsies from 1) normal human liver tissue (tumor free tissue from livers with metastasis from colorectal cancer) (n=5)  and 2) liver tissue from patients with chronic liver diseases, including primary sclerosing cholangitis (PSC) (n=12), non-alcoholic steatohepatitis (n=7) or other liver diseases (i.e., primary biliary cholangitis, autoimmune hepatitis, alcoholic liver disease and haemochromatosis) (n=8). The liver specimens were provided by the Norwegian biobank for primary sclerosing cholangitis, Oslo, Norway. The Affymetrix Human Gene 1.0 st array was used.

last: Folseraas.


title: Estrogen receptor beta repurposes EZH2 to suppress oncogenic NFκB signaling in triple negative breast cancer [RNA-Seq]

summary: The purpose of this study was to elucidate the transcriptome of ERβ expressing MDA-MB-231 cells following treatment with veh, E2, TNFα, or E2+TNFα.

overall_design: ERβ-expressing MDA-MB-231 cells were treated with ethanol vehicle or 1nM E2 for 5 days, followed by 3 hours of 20ng/ml TNFα in replicates of 3.

first: K

last: Aspros

first: J

last: Hawse


title: Comparison manual and two types of bioprinted kidney organoids by single cell RNA-seq

status: Public on Sep 01 2020

city: Parkville

first: Kynan

middle: Thomas

last: Lawlor

state: VIC

address: Murdoch Children's Research Institute, Flemington Road

institute: Murdoch Children's Research Institute

summary: We compared kidney organoids generated manually ('Man') to those generated by bioprinting single cell deposition ('R0') and thin bioprinted lines ('R40').

overall_design: 4 independent pools of iPS cells were differentiated (Takasato et al, 2015; Howden et al, 2019) and used to generate kidney organoids. Each pool was used to generate each of the 3 types of conformations: Manual organoids, bioprinted organoids composed of a single deposition of cells (R0) and bioprinted organoids composed a thin line of cells (R40). R0 and R40 were composed of 110k starting cells, while Manual was 230k. Each pool of cells was used to set up all 3 conditions, giving a total of 4 replicates per condition. Organoids were dissociated after 12 days of growth and each replicate was antibody labelled using a hash-tag oligo (HTO) barcode (BioLegend). Barcoded cells were pooled by conformation to generate and used to generate 3 10x single cell libraries. Each library represents a single conformation (Manual, R0, R40) and contains the 4 barcoded replicates. HTO sequences were separated by SPRI fractionation and used to geneate a separate HTO library. Standard 10x mRNA libraries and HTO libraries were sequenced separately, mapped and counted to give a count matrix then combined in Seurat for final analysis. Methods are described in detail in a paper under review, which will be referenced here once published.

middle: T

first: Jessica

last: Vanslambrouck

first: Melissa

middle: H

last: Little


title: Estrogen receptor beta repurposes EZH2 to suppress oncogenic NFκB signaling in triple negative breast cancer [ChIP-Seq]

summary: The purpose of this study was to elucidate the effects of ERβ on the genomic distribution of NFκB and RNA Polymerase II phospho-Ser2, as well as changes in  H3K27me3, H3K27ac, in MDA-MB-231 triple negative breast cancer cells.

overall_design: ERβ-expressing MDA-MB-231 cells were treated with ethanol vehicle or 1nM E2 for 5 days, followed by 30 minutes of 20ng/ml TNFα in replicates of 2.


title: Saliva cell type DNA methylation reference panel for epidemiology studies in children

city: Ann Arbor

first: Kelly

last: Bakulski

address: 1415 Washington Heights

department: Department of Epidemiology

institute: University of Michigan

summary: We profiled the DNA methylation of saliva cell types, to develop a tool for epidemiologic studies. Saliva was collected from 22 children, 21 participants with samples usable for DNA methylation, and sorted into immune and epithelial cells, using size exclusion filtration and magnetic bead sorting. DNA methylation was measured using the Illumina MethylationEPIC BeadChip. Saliva immune and epithelial cells have distinct DNA methylation profiles, which can influence whole saliva epidemiologic measures.

overall_design: 60 samples from 21 individuals: 18 large cell fractions, 20 CD45+ cell fractions, 18 whole saliva, 4 Oragene kit

first: Lauren

last: Middleton

first: John

last: Dou

first: Jonah

last: Fisher

first: Jonathan

last: Heiss

first: Allan

middle: C

last: Just

last: Faul

first: Erin

last: Ware

first: Colter

last: Mitchell

first: Justin

last: Colacino


title: Transcriptional response of Mycobacterium tuberculosis in mouse peritoneal macrophages

status: Public on Sep 24 2020

city: Moscow

first: Artem

last: Grigorov

address: Ulitsa Miklukho-Maklaya, 16/10

department: Genomics and Postgenomic Technologies

country: Russia

institute: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry

summary: This study was conducted to describe transcriptomic changes of M. tuberculosis during macrophage infection.

overall_design: Six samples, representing two groups: in vivo macrophage infection (mice of the I/St inbred strain) by Mycobacterium tuberculosis H37Rv and control (Mycobacterium tuberculosis H37Rv cells were incubated in Dubos broth). In each group, three independent biological samples were sequenced.

first: Taiana

last: Kondratieva

first: Konstantin

last: Majorov

first: Tatyana

last: Azhikina

first: Alexander

middle: S

last: Apt


title: ATAC-seq of human dendritic cells

status: Public on Sep 22 2020

city: Stanford

first: Juliana

last: Idoyaga

address: 299 Campus Dr, Fairchild Science Building

department: Microbiology & Immunology

institute: Stanford University

summary: Human dendritic cells (DCs) comprise subsets with distinct phenotypic and functional characteristics, but the transcriptional programs that dictate their identity remain elusive. Here, we collected global chromatin accessibility profiles across resting and stimulated human DC subsets by means of the assay for transposase-accessible chromatin using sequencing (ATAC-seq). We uncovered chromatin accessibility specific for each subset and discovered previously undescribed transcriptional regulators of DC function. By comparing responses of plasmacytoid DCs to type I interferon -producing and -not producing conditions, we identified genetic programs related to their function. Finally, by intersecting chromatin accessibility with genome-wide association studies, we recognized DC subset-specific enrichment of heritability in autoimmune diseases. Our results unravel the basis of human DC subset heterogeneity and provide a framework for their analysis in disease pathogenesis.

overall_design: We performed ATAC-seq to measure chromatin accessibility in primary human dendritic cells (pDCs, cDC1, cDC2, CD11chi tDCs, CD11clo tDCs) and CD14+ monocytes, as well as stimulated pDCs.

first: Rebecca

last: Leylek


title: RNA interference-independent reprogramming of DNA methylation in Arabidopsis

city: Bunkyo-ku

first: Taiko

middle: Kim

last: To

state: Tokyo

address: 7-3-1 Hongo

department: Biological Science

country: Japan

institute: University of Tokyo

type: Methylation profiling by high throughput sequencing

summary: DNA methylation is important for silencing transposable elements (TEs) in diverse eukaryotes including plants. In plant genomes, TEs are heavily methylated at cytosines of both CG and non-CG (or CH, where H can be A, T, or C) contexts. The role of RNA interference (RNAi) in establishing TE-specific DNA methylation has been extensively studied, although the significance of RNAi-independent pathways in DNA-methylation reprograming remains unexplored. Here, we directly investigated transgenerational de novo DNA methylation of TEs after the loss of DNA methylation. Our analyses identified very potent and precise RNAi-independent pathways for recovering CH methylation in most TE genes, or coding regions within TEs. Characterization of a small subset of TE genes that did not show CH methylation recovery revealed the impacts of H3K9 demethylation and replacement of histone H2A variants. These chromatin components are conserved between plants and animals and may contribute to chromatin reprograming in a conserved manner.

overall_design: Examination of DNA methylation (WGBS), transcription (RNA-seq), and localization of histone variants (ChIP-seq) in Arabidopsis WT, several DNA methylase and histone modification mutants, and their genetic hybrids.

middle: K

first: Yuichiro

last: Nishizawa

first: Soichi

last: Inagaki

first: Yoshiaki

last: Tarutani

first: Sayaka

last: Tominaga

first: Kazuya

last: Takashima

first: Kae

last: Kato

first: Mayumi

last: Takahashi

first: Atsusi

last: Toyoda

first: Asao

last: Fujiyama

first: Tetsuji

last: Kakutani


title: Structure of the full SARS-CoV-2 RNA genome in infected cells

city: Cambridge

first: Silvi

last: Rouskin

address: 455 Main Street

institute: Whitehead Institute for Biomedical Research

summary:
Purpose: SARS-CoV-2 is a betacoronavirus responsible for the COVID-19 pandemic. Currently, limited understanding of the molecular and structural biology of the virus hampers the development of antiviral drugs.
Methods: Vero cells were infected with SARS-CoV-2 and treated with dimethyl sulfate (DMS). RNA was extracted, reverse-transcribed using TGIRT-III, PCR-amplified, and sequenced using an Illumina iSeq100. Reads were trimmed with TrimGalore and mapped to the SARS-CoV-2 genome with Bowtie2. Mutations were quantified with DREEM, and the frequencies were used as folding constraints in RNAstructure. The output structures were visualized with VARNA.
Results: A data-driven secondary structure of the full RNA genome was generated, including structures previously lacking experimental validation, such as the TRSs. An alternative structure was identified for the frameshift element that differs drastically from previous in vitro models. The canonical frameshift element pseudoknot was not detected; in its place was an alternative stem partially overlapping with the canonical pseudoknot. Two alternative structures containing this stem were identified.
Conclusions: The genome-wide structure of SARS-CoV-2 provides a basis for developing therapeutics targeted to key regulatory structures. In particular, the model suggests that frameshifting is caused by a novel 75 nt stem rather than the canonical pseudoknot, which will guide the search for therapeutics that disrupt frameshifting..

sample_organism: Severe acute respiratory syndrome coronavirus 2

overall_design: Secondary structure of SARS-CoV-2 RNA genome in two cultures of infected Vero cells.

first: Tammy

last: Lan

first: Matthew

middle: F

last: Allan

first: Lauren

last: Malsick

first: Stuti

last: Khandwala

first: Sherry

last: Nyeo

first: Yu

last: Sun

first: Junjie

last: Guo

first: Mark

last: Bathe

first: Anthony

last: Griffiths


title: Actin R256 mono-methylation is a conserved post-translational modification involved in transcription

status: Public on Aug 26 2020

city: Smithville

first: Bin

last: Liu

state: TX

address: 1808 Park Road 1C

department: Epigenetics and Molecular Carcinogenesis

institute: University of Texas MD Anderson Cancer Center

summary: Nuclear actin has been elusive due to the lack of molecular mechanisms. From actin-containing chromatin remodeling complexes, we discovered an arginine mono-methylation mark on evolutionarily conserved R256 residue of actin (R256me1). Actin R256 mutations in yeast affect nuclear functions, and cause diseases in human. Interestingly, we show that an antibody specific for actin R256me1 preferentially stain nuclear actin over cytoplasmic actin in yeast, mouse and human cells. We also show that actin R256me1 is regulated by protein arginine methyl transferase-5 (PRMT5) in HEK293 cells. Genome-wide survey of actin R256me1 mark provides a landscape for nuclear actin correlated with transcription. Further gene expression and protein interaction studies uncover extensive correlations between actin R256me1 and active transcription. The discovery of actin R256me1 mark suggests a fundamental mechanism to distinguish nuclear actin from cytoplasmic actin through post-translational modification (PTM), and potentially implicates an actin PTM mark in transcription and human diseases.

overall_design: Three biological replciates for either WT or R256C ( a pathogenic mutant in human ⍺ actin causing cerebral and aortic aneurysms) were collected for RNA extraction and subjected to RNA-Seq analysis and gene set enrichment analysis  to identify differentially expressed gene sets

first: Xuetong

last: Shen

first: Prabodh

last: Kapoor

first: Yuan

last: Zhong


title: Zfp281 functions as a barrier in embryonic stem cells to extraembryonic endoderm stem cells differentiation.

status: Public on Aug 19 2020

city: New York

first: Jianlong

last: Wang

department: Department of Medicine

institute: Columbia University Irving Medical Center

overall_design: Refer to individual Series


title: Transcriptome of chronic myelomonocytic leukemia

status: Public on Aug 14 2020

first: Mrinal

middle: M.

last: Patnaik

address: 200 First Street SW

department: Division of Hematology

summary: We report the gene expression of human chronic myelomonocytic leukemia by performing whole transcriptome shotgun sequencing of peripheral blood mononuclear cells of patients with chronic myelomonocytic leukemia.

overall_design: Comparison of gene expression between proliferative and dysplastic chronic myelomonocytic leukemia.

middle: M

first: Jaime

middle: I

last: Davila


title: Direct reprogramming of fibroblasts identifies signatures of vascular dysfunction in physiological aging and Hutchinson-Gilford Progeria Syndrome

status: Public on Sep 03 2020

first: April

last: Williams

summary: Our goal was to identify gene expression and functional differences between directly reprogrammed vascular cells derived from young and old individuals, as wells as from healthy and Hutchinson-Gilford Progeria Syndrome (HGPS) donors.  We provided a full characterization of reprogrammed endothelial and smooth muscle cells by comparing their gene expression with both the original fibroblasts and primary vascular cells, showing that reprogrammed cells express key vascular cell-identity genes and contribute to the formation of in vitro 3D vascular structures. We identified and validated biomarkers of vascular dysfunction in the context of physiological and accelerated aging typical of HGPS patients, and we demonstrated their direct contribution in the modulation of vascular permeability.

overall_design:
We performed RNAseq on directly reprogrammed vascular cells (induced vascular endothelial cells (iVECs) and induced smooth muscle (iSM) cells). We compared directly reprogrammed cells with the original fibroblasts as well as with primary vascular cells. Furthemore, we compared directly reprogrammed cells obtained from healthy young vs. old donors (N=3 per group) as wells as from Hutchinson Gilford Progeria Syndrome (HGPS) (N=8) vs. healthy (N=3) donors.

This series includes the re-analysis of GSM3175518, GSM3175519 and GSM3175520.

first: Simone

last: Bersini

first: Roberta

last: Schutle

first: Hannah

last: Tsai

first: Martin

middle: W

last: Hetzer

first: Maxim

last: Shokhirev

first: Ling

last: Huang


title: Early precursor T cells establish and propagate T cell exhaustion in chronic infection [RNA-seq]

status: Public on Aug 28 2020

city: Clayton

first: Wei

last: Shi

state: Victoria

address: 15 Innovation Walk

department: Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute

institute: Monash University

summary: TCF1+ precursors of exhausted T cells maintain T cell immunity during chronic infection and cancer and mediate the response to immune checkpoint inhibition; however, their developmental requirements are poorly understood. Here, we demonstrate that high antigen load promoted the differentiation of TCF1+ precursor T cells, which acquired hallmarks of T cell exhaustion such as impaired production of IFN within the first days after infection. Transcriptional profiling revealed that exhaustion was established first in precursor T cells while early effector cells retained a polyfunctional profile. Early precursor T cells exposed to high amounts of antigen showed epigenetic imprinting of T cell receptor-dependent transcription factor binding, were dependent on BACH2 and restricted to the generation of T cells displaying features of exhaustion. Overall, we demonstrate that T cell exhaustion manifests first in TCF1+ precursor T cells and is propagated subsequently to the pool of antigen-specific T cells.

overall_design: RNA sequencing of ID3GFP+ and ID3GFP- P14 T cells obtained from LCMV Armstorng and Docile infected mice.

first: Daniel

last: Utzschneider

first: Sarah

last: Gabriel

first: David

last: Chisanga

first: Patrick

last: Gubser

first: Axel

last: Kallies


title: Spleen glia are a transcriptionally unique glial subtype interposed between immune cells and sympathetic axons

first: Tawaun

middle: Autry

last: Lucas

state: CA

address: 1201 Welch Rd

summary: Non-myelinating glia in visceral organs have not been well characterized. It Is likekely they have uniqe functions in the organs in which they reside. This is interesting in immune organs such as the spleen in which glia ensheath nerves that come in close contact with immune cells. However, glia in the spleen have not been well characterized. We report here an in-depth anatomical and transcriptional characterization of spleen glia.

overall_design: Examination of spleen glia enriched genes comapred to whole spleen

middle: A

first: Li

last: Zhu

first: Marion

last: Buckwalter


title: Recruitment of HMGN By H3K27ac Modified Nucleosomes to Regulatory Sites Modulates Transcription Factor Binding to Chromatin

status: Public on Sep 20 2020

city: Bethesda

first: Michael

last: Bustin

state: MD

address: Building 37, Room 3122

department: National Cancer Institute

institute: National Institutes of Health

summary: H3K27ac modified nucleosomes are a major epigenetic mark of active chromatin that can be used to identify cell type specific chromatin regulatory regions which serve as binding sites for transcription factors. Here we show that H3K27ac modified nucleosomes recruit the ubiquitous nucleosome binding proteins HMGN1 and HMGN2 to cell-type specific chromatin regulatory regions. We find that HMGNs bind directly to the acetylated nucleosome and that the H3K27ac residue and linker DNA are necessary and sufficient for the preferential binding of HMGNs to the modified nucleosomes. Loss of HMGNs increases the levels of H3K27me3 and the histone H1 occupancy at enhancers and promoters and alters the interaction of several transcription factors with chromatin.  These experiments identify an additional mechanism whereby the H3K27ac epigenetic mark promotes chromatin decompaction and provide insights into the molecular mechanism whereby HMGN proteins, which bind to chromatin without DNA sequence specificity, modulate cell type specific gene expression.

overall_design: Two biological replicates for wild type and two biological replicates for double knock-out, using ChIP-seq technology

first: Shaofei

last: Zhang

first: Yuri

last: Postnikov

first: Alexei

last: Lobanov

first: Takashi

last: Furusawa

first: Tao

last: Deng


title: Distinct and overlapping roles of STAG1 and STAG2 in cohesin localization and gene expression in embryonic stem cells [RNA-seq]

status: Public on Aug 10 2020

city: Chapel Hill

first: Jill

last: Dowen

state: NC

address: 250 Bell Tower Drive

institute: University of North Carolina at Chapel Hill

summary:
Purpose: Cohesin is a ring-shaped complex that is critical to genome organization and is an important regulator of gene expression. How cohesin accessory proteins contribute to cohesin function remain unclear. The purpose of this study is to assess the roles of cohesin accessory proteins, STAG1 and STAG2, in cohesin localization and gene expression.

Method: The role of STAG1 and STAG2 in gene regulation was assessed by performing RNA-seq in wildtype and CRISPR-Cas9 genome edited STAG knockout mouse embryonic stem cells (mESCs). The redundancy of the STAGs was addressed by using siRNA against STAG1 in a STAG2-knockout background.

overall_design: RNA-seq was performed in wild type mESCs, STAG-knockout CRISPR-Cas9 genome edited mESCs, and STAG-knockout mESCs with siRNA depletion of the other STAG, all in triplicate.

first: Nicole

middle: L

last: Arruda

first: Zachary

last: Carico

first: Megan

last: Justice

first: Ying

last: Liu

first: Junjie

last: Zhou

first: Holden

last: Stefan


title: Atg7-dependent autophagy regulates the naive to primed transition by selective Nanog degradation in mouse embryonic stem cells [WGBS]

status: Public on Jul 22 2020

city: Wuhan

first: Jing-Jing

address: 1 Lion Rock Street

department: Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction

institute: Huazhong Agricultural University

summary: Autophagy is a conserved cellular mechanism to degrade unwanted cytoplasmic proteins and organelles to recycle their components, and it is proved to be critical for embryonic stem cell (ESC) self-renewal and somatic cell reprogramming. However, the role of autophagy in embryonic development remains elusive, and no information exists regarding its functions during the transition from naive to primed pluripotency. Here by using an in vitro transition model of ESCs to epiblast-like cells (EpiLCs), we describe that the dynamic changes in Atg7-dependent autophagy is required for the naive to primed transition, and it is also necessary for germline specification. RNA-seq and ATAC-seq profiling reveal that Nanog acts as a barrier to prevent pluripotency transition, and autophagy-dependent Nanog degradation is important for dismantling the naive pluripotency expression program through decommissioning of naive-associated active enhancers. Mechanistically, we found that autophagy adaptor protein Sqstm1 (p62) is nucleus located during the pluripotency transition period and it is preferentially associated with ubiquitinated Nanog for selective protein degradation. In vivo, loss of autophagy by Atg7 depletion disrupts peri-implantation development and we observed increased chromatin association of Nanog, which affects neuronal differentiation through activation of a subset of neuroectodermal development-associated enhancers. Taken together, our findings illuminate regulatory mechanisms underlying the naive to primed transition and reveal that autophagy-dependent regulation of Nanog is essential for exit from the naive state and marks distinct cell fate allocation during lineage specification.

overall_design: The WGBS experiments of the Control and Atg7 KO groups were taken at time point day 4 from the 4-day ESC to EpiLC transition.

first: Jilong

first: Hainan

last: He

first: Jingjing

first: Yi

last: Miao


title: An Activity-Guided Map of Electrophile-Cysteine Interactions in Primary Human T Cells

status: Public on Jul 30 2020

first: April

last: Williams

address: 10010 N Torrey Pines Rd

summary: Electrophilic compounds originating from nature or chemical synthesis have profound effects on immune cells. These compounds are thought to act by cysteine modification to alter the functions of immune-relevant proteins; however, our understanding of electrophile-sensitive cysteines in the human immune proteome remains limited. Here, we present a global map of cysteines in primary human T cells that are susceptible to covalent modification by electrophilic small molecules. More than 3,000 covalently liganded cysteines were found on functionally and structurally diverse proteins, including many that play fundamental roles in immunology. We further show that electrophilic compounds can impair T cell activation by distinct mechanisms involving the direct functional perturbation and/or degradation of proteins. Our findings reveal a rich content of ligandable cysteines in human T cells and point to electrophilic small molecules as a fertile source for chemical probes and ultimately therapeutics that modulate immunological processes and their associated disorders.

overall_design: Examination of gene expression changes after tretment of primary human T cells with electrophilic immunosuppressive compounds and corresponding controls for 24h

first: Ekaterina

middle: V

last: Vinogradova

first: Benjamin

last: Cravatt

first: Maxim

middle: N

last: Shokhirev


title: Comparison of bioprinted kidney organoids in different conformations

first: Kynan

middle: Thomas

last: Lawlor

summary: Bulk RNA-seq comparison of kidney organoids bioprinted in 3 different conformations with varying starting cell densities. Density is dictated by the ratio of bioprinter tip movement to  the amount of extrusion, where higher ratios spread cells over a larger surface area. We compare organoids printed with no movement ('blob', ratio 0) to those with moderate ('line 3', ratio 20) or high movement ('line 1', ratio 40).

overall_design: Each condition consistes of 3 replicate organoids, where each replicate was generated from a distinct well of differentiated iPSCs.

first: Jessica

last: Vanslambrouck

first: Melissa

last: Little


title: Single cell RNA-Seq identifies mechanisms controlling hypothalamic patterning and differentiation

status: Public on Aug 09 2020

city: Baltimore

first: Dong Won

last: Kim

state: Maryland

address: 733 N.Broadway, MRB building (Rm 332)

department: Neuroscience

institute: Johns Hopkins University

summary: The hypothalamus is a central regulator of many innate behaviors that are essential for survival, but the molecular mechanisms controlling hypothalamic patterning and cell fate specification remain poorly understood. To identify genes that control hypothalamic development, we have used single-cell RNA sequencing (scRNA-Seq) to profile mouse hypothalamic gene expression across 11 developmental time points between embryonic day 10 and postnatal day 45.   This identified genes that delineated clear developmental trajectories for all major hypothalamic cell types and readily distinguished major regional subdivisions of the developing hypothalamus.  We show that this approach can rapidly and comprehensively characterize mutants that control hypothalamic patterning and, in doing so, identify multiple genes that simultaneously repress posterior hypothalamic identity while promoting prethalamic identity.  This argues in favor of a modified columnar organization of hypothalamus and prethalamus.  These data serve as a resource for further studies of hypothalamic development, physiology, and dysfunction.

overall_design: Dissection and cell dissociation - Embryos or postnatal mice were collected and dissociated following previously published protocol. Embryos were collected using Hibernate-E media (Thermo Fisher Scientific) with 2% B-27 supplement (Thermo Fisher Scientific) and GlutaMAX supplement (0.5 mM final, Thermo Fisher Scientific). A small incision was made dorsal to the lower jaw to expose the ventral portion of the brain. For samples collected between E10 and E16, tissue residing posterior to the medial ganglionic eminence and anterior to the midbrain and sensory thalamus was dissected to collect developing prethalamus and hypothalamus. Prethalamus was excluded from samples aged E18 and older, with only hypothalamus collected, as previously described (2). Collection of medial ganglionic eminence and posterior structure to the supramammillary nucleus were variable between dissections. Between 8 and 12 embryos of either sex were collected for each embryonic time point. Postnatal mice were collected using Hibernate-A media with 2% B-27 and GlutaMAX (0.5 mM final), and the tissue posterior to the optic chiasm and anterior to the posterior hypothalamus were collected. Eight pups (4 male and 4 female) were collected for P4, P8, and P14 dataset, and 3 male mice were pooled for P45 dataset. E10, E12, and E15; E11 and E13; E14, E16, and P45; E18, P4, and P14 were each generated on the same day.For single-cell phenotyping studies, E12.5 time-mated mice were collected and placed in buffer mentioned above on ice. Tail-tips were collected and rapidly genotyped using GeneAmp Fast PCR mastermix (Thermo Fisher Scientific). Both control and mutant groups were collected on the same day, and E12.5 embryos were pooled from 3 different dams. Following dissection, tissues were dissociated in papain (Worthington Biochemical) as previously described in calcium-free Hibernate media. Tissue debris were removed using OptiPrep density gradient media (Sigma-Aldrich) in postnatal mice following cell dissociation. Number of viable cells was counted manually haemocytometer with Trypan Blue staining, and cell concentration was adjusted following manufacturer’s protocol of 10x Genomics. 10x library generation and data processing - Suspended cells were loaded into 10x Genomics Chromium Single Cell System (10X Genomics), and libraries were generated using v1 (1 library) and V2 chemistry with manufacturer’s instructions. Libraries were sequenced on Illumina MiSeq (1 library) and NextSeq500. Sequencing data were pre-processed through the Cell Ranger pipeline (10x Genomics) with default parameters, aligned to mm10 genome, and matrix files were used for bioinformatic analysis.

first: Dong

first: Parris

last: Washington

first: Sonia

last: Lin

first: Changyu

last: Sun

first: Lizhi

last: Jiang

first: Zoe

last: Wang

first: Seth

last: Blackshaw


title: Identification of ADAR3-bound targets in U87 cells

city: Bloomington

first: Heather

middle: Ann

last: Hundley

state: IN

address: 1001 E 3rd St

department: Biology

institute: Indiana University Bloomington

summary: We report transcripts bound by ADAR3 through RNA immunoprecipitation of U87 cells expressing 3X-FLAG tagged ADAR3 compard to control cells

overall_design: Determining transcripts enriched for ADAR3 binding by deep sequencing 3 biological replicates of RIP-RNA from U87 cells expressing ADAR3 and control cells and total RNA from the same cells

first: Reshma

last: Raghava Kurup

first: Eimile

last: Oakes

first: Aidan

last: Manning


title: Expression data from mouse bone marrow derived macrophages (BMDM)

status: Public on Jul 01 2020

city: St. Louis

first: MALAY

last: HALDAR

state: MO

address: 7751 charing square lane

department: Pathology and Immunology

institute: Washington University in St. Louis

summary:
The transcription factor Spic regulates inflammatory responses and iron metabolism in activated macrophages.
We used microarrays to uncover genes regulated by the transcription factor Spic in bone marrow derived macrophages.

overall_design: To identify Spic-regulated genes, we isolated bulk bone marrow cells from wild type and Spic-deficient mice and cultured them under conditions promoting macrophage differentiation. The resulting BMDMs were exposed to LPS or control and their gene expression were analyzed by microarray.

first: Zahidul

last: Alam

first: Samir

last: Devalaraja

first: Minghong

last: Li

first: Tsun

last: To

first: Ian

last: Folkert

first: Erick

last: Mitchell-Velasquez

first: Mai

middle: T

last: Dang

first: Patricia

last: Young

first: Christopher

middle: J

last: Wilbur

first: Michael

last: Silverman

first: Xinyuan

first: Youhai

middle: H

last: Chen

first: Paul

last: Hernandez

first: Aritra

last: Bhattacharyya

first: Mallar

last: Bhattacharya

first: Matthew

last: Levine

first: Malay

last: Haldar


title: Integrative analysis of coding, non-coding transcriptome and chromatin accessibility reveals a distinct gene expression and epigenetic landscape in human memory B cells

status: Public on Aug 27 2020

city: San Antonio

first: Justin

last: Moroney

state: Texas

address: 7703 Floyd Curl Drive; Medical Building, Rm #5.011V

department: Microbiology, Immunology and Molecular Genetics

institute: UT Health San Antonio

summary: Memory B cells (MBCs) are long-lived and rapidly respond to cognate antigen with production of high-affinity, generally, class-switched antibodies. Here, we have developed an integrative analysis of differentially expressed (DE) mRNAs, miRNAs, lncRNAs, chromatin profiles and cis-regulatory elements to determine how gene expression intersects with epigenetic regulatory elements in human MBCs. Using a multiparameter cell sorting approach, we isolated CD27-IgD+NBCs, CD27+IgD+unswMBCs, CD27+IgG+swMBCs and CD27+IgA+swMBCs as well as total CD27-NBCs and CD27+MBCs from peripheral blood of healthy human subjects of different age, sex and ethnic background. Total RNA was used to construct RNA-Seq libraries and subjected to next generation sequencing for in-depth analysis of B cell transcriptome, including mRNAs, miRNAs and lncRNAs, as well as Ig heavy chain (H) V(D)J, and Ig light chain (L) gene transcripts. Differential expression analysis identified shared mRNA, miRNA and lncRNA profiles that distinguished CD27+IgG+swMBCs and CD27+IgA+swMBCs from CD27-IgD+NBCs, and stratified CD27+IgD+unswMBCs between NBCs and swMBCs. Further, targeted integration using Ingenuity Pathway Analysis (IPA) outlined distinct signaling pathways in human MBCs, while the activity of TFs was inferred from global changes in transcriptional activation. ATAC-Seq was integrated with RNA-Seq to correlate DE RNAs with chromatin accessibility, thereby uncovering distinct profiles of cis-regulatory elements in human MBCs. A deep downregulation of MIR181 was concomitant with upregulation of this microRNA target genes, which accounted for 11 percent of the DE mRNAs in swMBCs, indicating an important role for MIR181 in MBC differentiation and/or maintenance. Further, lncRNA MIAT, a sponge of MIR181, was upregulated in MBCs and inversely correlated with MIR181a and MIR181b expression. This along with chromatin accessibility contributes downregulation of MIR181 and promotes MBC-specific gene expression. Overall, our findings provide evidence for overlapping layers of regulation, including chromatin remodeling, cis-regulatory elements and distinct sets of miRNAs and lncRNAs, which integrate to dictate gene expression profiles and activation pathways characteristic of human MBCs.

overall_design:
Analysis of  mRNA expression and chromatin accessibility profiles in human B cells (total CD27- NBCs and total CD27+ MBCs) isolated from the peripheral blood of 4 donors

These data are related to amplicon sequencing data included in PRJNA658698.

first: Paolo

last: Casali

first: Hong

last: Zan

first: Yidong


title: Expression data from mouse circulating Ly6C-low subset of monocytes (patrolling monocytes)

first: MALAY

last: HALDAR

summary:
The transcription factor Spic regulates genes associated with heme and iron metabolism.
We used microarrays to identify genes regulated by the transcription factor Spic in patrolling monocytes.

overall_design: To identify Spic-regulated genes in patrolling monocytes, we isolated Ly6C-low monocytes from peripheral blood of Spic heterozygous and Spic knockout mice that were previously exposed to LPS and performed gene expression analysis of these cells using microarray.

first: Zahidul

last: Alam

first: Samir

last: Devalaraja

first: Minghong

last: Li

first: Tsun

first: Ian


title: Origin of kidney papillary tumors from renal progenitors

status: Public on Jul 16 2020

city: Firenze

first: Roberto

last: Semeraro

address: Viale Pieraccini, 6

department: Medicina Sperimentale e Clinica

country: Italy

institute: Università degli studi di Firenze

summary:
Purpose: Using single-cell RNA sequencing (scRNAseq), Young et al. (Science, 2018, PMID 30093597) recently identified a canonical cancer transcriptome as the possible cell of origin of papillary renal cell carcinoma (pRCC) that matched a rare proximal convoluted tubular cell population characterized as VCAM1+ SLC17A3+ SLC7A13-, a cell cluster signature defined as PT1. The goal of this study is to compare pRCC transcriptome to human renal progenitor cell (RPC) transcriptome.
Methods: To compare the pRCC cells with PT1 features to our RPC population, we analyzed Young pRCC scRNAseq data set together with scRNAseq data obtained from human RPC cultures.
Results: The PT1 signature that characterizes the cell of origin of pRCC identifies a RPC cluster.
Conclusions: These results suggest that the cell of origin of pRCC corresponds to RPCs.

overall_design: Comparison of scRNAseq data from pRCC cells and RPCs. The same RPCs sample was used to create four libraries, each linked to a different 10x Genomics index.

first: Anna

last: Peired

first: Paola

last: Romagnani


title: Expression data from mouse spleen pre-Red Pulp Macrophages (PreRPM), circulating Ly6C-low subset of monocytes (patrolling monocytes), and bone marrow derived macrophages (BMDM)

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


title: LIN28B Regulates Transcription and Potentiates MYCN-induced Neuroblastoma in a let-7-independent Manner [ChIP-seq]

city: Hangzhou

first: Ting

last: Tao

state: Zhejiang

address: 3333 Binsheng Road

institute: Children's Hospital, Zhejiang University School of Medicine

summary: LIN28B is highly expressed in neuroblastoma and promotes tumorigenesis at least in part through inhibition of let-7 microRNA biogenesis. Here, we report that overexpression of either wild-type LIN28B, or a LIN28B mutant that is unable to bind and inhibit let-7, accelerates the onset and increases the penetrance of MYCN-induced neuroblastoma, potentiates the invasion and migration of transformed sympathetic neuroblasts and drives the distant metastases in vivo. Genome-wide ChIP-seq analysis and co-immunoprecipitation experiments show that LIN28B indirectly binds active gene promoters in neuroblastoma cells through an interaction with ZNF143 and activates the expression of downstream targets, including GSK3B and L1CAM, which are involved in neuronal cell adhesion and migration. These findings reveal an unexpected, let-7-independent function for LIN28B in transcriptional regulation during neuroblastoma pathogenesis.

overall_design: ChIP-Seq for LIN28B, ZNF143 in BE2C, CHP134 and Kelly neuroblastoma cells

first: Hui

last: Shi

first: Luca

last: Mariani

first: Brian

last: Abraham

first: Adam

middle: D

last: Durbin

first: Mark

last: Zimmerman

first: John

last: Powers

first: Pavlos

last: Missios

first: Kenneth

last: Ross

first: Antonio

middle: R

last: Perez-Atayde

first: Martha

last: Bulyk

first: Richard

last: Young

first: George

middle: Q

last: Daley

first: A

last: Look


title: Expression data from mouse spleen pre-Red Pulp Macrophages (PreRPM)

first: MALAY

last: HALDAR

summary:
The transcription factor Spic regulates inflammatory responses in PreRPM.
We used microarrays to identify genes regulated by the transcription factor Spic in PreRPMs.

overall_design: To identify Spic-regulated genes in PreRPMs, we isolated PreRPMs from wild type and Spic-knockout spleens using FACS and compared gene expression profile of these cells using microarrays.

first: Zahidul

last: Alam

first: Samir

last: Devalaraja

first: Minghong

last: Li

first: Tsun

middle: K

last: To

first: Ian

last: Folkert

first: Erick

last: Mitchell-Velasquez

first: Mai

last: Dang

first: Patricia

first: Christopher

last: Wilbur

first: Michael

last: Silverman

first: Xinyuan

first: Youhai

last: Chen

first: Paul

last: Hernandez

first: Aritra

last: Bhattacharyya

first: Mallar

last: Bhattacharya

first: Matthew

last: Levine

first: Malay

last: Haldar


title: Clonal relations in the mouse brain revealed by single-cell and spatial transcriptomics

status: Public on Jun 29 2020

city: Solna

last: Ratz

address: Solnavägen 9

department: CMB

country: Sweden

institute: Karolinska Institute

summary: The mammalian brain contains many specialized cells that develop from a thin sheet of neuroepithelial progenitor cells. Single-cell transcriptomics revealed hundreds of molecularly diverse cell types in the nervous system, but the lineage relationships between mature cell types and progenitor cells are not well understood. Here we show in vivo barcoding of early progenitors to simultaneously profile cell phenotypes and clonal relations in the mouse brain using single-cell and spatial transcriptomics. By reconstructing thousands of clones, we discovered fate-restricted progenitor cells in the mouse hippocampal neuroepithelium and show that microglia are derived from few primitive myeloid precursors that massively expand to generate widely dispersed progeny. We combined spatial transcriptomics with clonal barcoding and disentangled migration patterns of clonally related cells in densely labelled tissue sections. Our approach enables high-throughput dense reconstruction of cell phenotypes and clonal relations at the single-cell and tissue level in individual animals and provides an integrated approach for understanding tissue architecture.

overall_design: Lentiviral delivery of a transcribed barcode into neuroepithelial progenitor cells using in utero microinjection into the mouse forebrain at embryonic day 9.5 followed by RNA-seq analysis of barcoded cells at postnatal day 11-14.

first: Leonie

last: von Berlin

first: Ludvig

last: Larsson

first: Marcel

last: Martin

first: Jakub

middle: O

last: Westholm

first: Gioele

last: La Manno

first: Joakim

last: Lundeberg

first: Jonas

last: Frisén


title: Combinatorial single-cell analyses of granulocyte-monocyte progenitor heterogeneity reveals an early uni-potent neutrophil progenitor

city: Singapore

first: Nicholas

last: Ang

address: 8a Biomedical Grove, Singapore 138648

country: Singapore

institute: Singapore Immunology Network (SIgN)

summary: Granulocyte-monocyte progenitors (GMPs) have been previously defined for their potential to generate various myeloid progenies such as neutrophils and monocytes. Although studies have proposed lineage heterogeneity within GMPs, it is unclear if committed progenitors already exist among these progenitors and how they may behave differently during inflammation. By combining single-cell transcriptomic and proteomic analyses, we identified the early committed progenitor within the GMPs responsible for the strict production of neutrophils, which we designate as proNeu1. Our dissection of the GMP hierarchy led us to further identify a previously unknown intermediate proNeu2 population. Similar populations could be detected in human samples. proNeu1s, but not proNeu2s, selectively expanded during the early phase of sepsis at the expense of monocytes. Collectively, our findings help shape the neutrophil maturation trajectory roadmap and challenge the current definition of GMPs.

overall_design: BM Ly6C-GMP, proNeu1, proNeu2 and cMoP were sorted based on the gating strategy of Figure 4A. preNeus were sorted as described previously.  3 biological replicates were sequenced each. For sepsis-related  samples, mice were sacrificed after 3 days of mid-grade CLP and sorted in the same way as WT controls.

first: Bernett

last: Lee


title: Comparative Transcriptome Analysis of Endemic and Epidemic Kaposi’s Sarcoma (KS) Lesions and the Secondary Role of HIV-1 in KS Pathogenesis

status: Public on Jun 26 2020

city: Philadelphia

first: Priyankara

last: Wickramasinghe

state: PA

address: 3601 Spruce Street

department: Bioinformatics

institute: The Wistar Institute

summary:
In sub-Saharan Africa, endemic Kaposi’s sarcoma (EnKS) is still prevalent despite high incidence of epidemic Kaposi’s sarcoma (EpKS) resulting from the on-going HIV-1 epidemic. While KSHV is clearly the etiologic agent of KS, the mechanisms underlying KS development are not fully understood. For example, HIV-1 co-infection and concomitant immune dysfunction have been associated with EpKS development. However, the direct or indirect role(s) of HIV-1, and therefore of immune suppression, in EpKS remains unclear. How, or whether, EpKS is mechanistically distinct from EnKS is unknown. Thus, the absence of HIV-1 co-infection in EnKS provides a unique control for investigating and deciphering whether HIV-1 plays a direct or indirect role in the EpKS tumor microenvironment. We hypothesized that HIV-1 co-infection would induce transcriptome changes that differentiate EpKS from EnKS, thereby defining the direct intra-tumor role of HIV-1 in KS. Comparison of ART-treated and -naïve patients would further define the impact of ART on the KS transcriptome.
We utilized RNA-seq followed by multiparameter bioinformatics analysis to compare transcriptomes from KS lesions to uninvolved control skin. We provide the first transcriptomic comparison of EpKS versus EnKS, ART-treated vs. –naïve EpKS and male vs. female EpKS to define the roles of HIV-1 co-infection, the impact of ART, and gender on KS gene expression profiles. Our findings suggest that ART-use and gender have minimal impact on transcriptome profiles of KS lesions. Gene expression profiles strongly correlated between EpKS and EnKS patients (Spearman r=0.83, p<10-10). A subset of genes involved in tumorigenesis and inflammation/immune responses showed higher magnitude, but not unique dysregulation in EnKS compared to EpKS. While gender and ART had no detectable contribution, the trend toward higher magnitude of gene dysregulation in EnKS coupled with the absence of HIV-1 transcripts in EpKS may suggest an indirect or systemic effect of HIV-1 to promote KS tumorigenesis.

overall_design: RNA-seq of Kaposi’s sarcoma lesion, control and normal tissues from KS patients of both genders, different KS origins and HIV/ART status.

first: Charles

last: Wood

first: Salum

last: Lidenge


title: Extraction of Distinct Cell Types from within a Genetically Continuous Population

status: Public on Jul 25 2020

city: San Diego

first: April

middle: Elizabeth

last: Williams

department: IGC

institute: Salk Institute for Biological Studies

summary: The brain is composed of millions of diverse neurons that differ systematically in the genes that they express. Analyses of single cell gene expression data reveal clusters which correspond to different cell types that differ in their connectivity and function. Can the connections formed by genetically identical neurons systematically alter their gene expression? Here we address this question  by combining retrograde labeling, single cell gene expression, and rabies-based analyses of connectivity to assess cortical-cortical projection neurons in the mouse primary visual cortex. We find that pyramidal neurons projecting to different cortical targets and with known functional differences differ systematically in their gene expression and connectivity despite forming only a single genetic cluster with continuous variability. These observations demonstrate that single cell gene expression analysis in isolation is insufficient to identify neuron types.

overall_design: We used fluorescent tagged retrograde tracers, single-nuclei sequencing combined by projection profile via retrogradely labeling virus, and rabies-based projection- and layer-dependent input tracing to identify distinctive cell types from V1 L2/3 pyramidal neurons.

first: Euiseok

last: Kim

first: Zhuzhu

last: Zhang

first: Ling

last: Huang

first: Joseph

last: Ecker

first: Edward

middle: M

last: Callaway


title: Integrated analysis of DNA methylation, chromatin accessibility, transcription factor binding, and gene expression in large collections of breast cancer cell lines and patient tumors

status: Public on Jun 11 2020

city: Salt Lake City

first: Katherine

middle: E

last: Varley

state: UT

address: 2000 Circle of Hope, Room 3719

department: Department of Oncological Sciences

institute: Huntsman Cancer Institute University of Utah

overall_design: Refer to individual Series


title: Dynamic regulation of histone modifications and long-range chromosomal interactions during post-mitotic transcriptional reactivation

state: California


title: Dynamic regulation of histone modifications and long-range chromosomal interactions during post-mitotic transcriptional reactivation [EU-RNA-seq]

summary: During mitosis, transcription of genomic DNA is dramatically reduced, before its reactivation occurs during nuclear reformation in anaphase/telophase. Many aspects of the underlying principles that mediate transcriptional memory and reactivation in the daughter cells remain unclear. Here, we used ChIP-seq on synchronized cells at different stages after mitosis to generate genome-wide maps of histone modifications. In combination with EU-RNA-seq and Hi-C analyses, we show that, during prometaphase, promoters, enhancers, and insulators retain H3K4me3 and H3K4me1, while losing H3K27ac. Enhancers harboring mitotic H3K4me1 are associated with cell type-specific transcription factor motifs and mitotic activation of cell identity genes. As cells exit mitosis, promoters regain H3K27ac, which correlates with transcriptional reactivation. Insulators also gain H3K27ac and CCCTC binding factors (CTCF) in anaphase/telophase. This increase may play a role in the establishment of topologically associating domains (TADs). Together, our results show that the genome is reorganized in sequential order, in which histone methylations occur first in prometaphase, histone acetylation and CTCF in anaphase/telophase, transcription in cytokinesis, and long-range chromatin interactions in early G1. Our results provide insights into the histone modification landscape that allows faithful reestablishment of the transcriptional program and TADs during cell division.

overall_design: EU-RNA-seq during mitosis-G1 in U2OS cells

first: Hyeseon

last: Kang

first: Maxim

last: Shokhirev

first: Martin

middle: W

last: Hetzer


title: Dynamic properties of transcription and co-transcriptional splicing during early stages of Drosophila development

status: Public on Jun 17 2020

city: Lisbon

first: Maria

last: Carmo-Fonseca

address: Av. Egas Moniz

country: Portugal

institute: Instituto de Medicina Molecular

summary: Widespread co-transcriptional splicing has been demonstrated from yeast to human. However, measuring the kinetics of splicing relative to transcription has been hampered by technical challenges. Here, we took advantage of native elongating transcript sequencing (NET-seq) to identify the position of RNA polymerase II (Pol II) when exons become ligated in the newly synthesized RNA. We analyzed Drosophila melanogaster embryos because the genes transcribed initially during development have few and short introns (like yeast genes), whereas genes transcribed later contain multiple long introns (more similar to human genes). Moreover, compared to human, the Drosophila genome is more compact and thus the coverage of NET-seq reads on intragenic regions is higher. We detected spliced NET-seq reads connected to Pol II molecules that were positioned just a few nucleotides downstream of the 3’ splice site. Although the majority of splice junctions were covered by spliced reads, many introns remained unspliced, resulting in a complex range of heterogeneity in splicing dynamics. Introns that show splicing completion before Pol II has reached the end of the downstream exon are necessarily intron-defined. As expected, we found a relationship between the proportion of spliced reads and intron size. However, intron definition was observed at all intron sizes. Both canonical and recursive splicing were associated with a higher Pol II density, suggesting a splicing-coupled mechanism that slows down transcription elongation. We further observed that transcription termination was very efficient for isolated genes but that the presence of an overlapping antisense gene was often associated with transcriptional read-through. Taken together, our data unravels novel dynamic features of Pol II transcription and splicing in the developing Drosophila embryo.

overall_design: Native Elongating Transcript sequencing was performed in whole Drosophila embryos collected at one of two timepoints (2-3 hours post-fertilisation and 4-6 hours post-fertilisation).

first: Pedro

last: Prudêncio

first: Kenny

last: Rebelo

first: Rosina

last: Savisaar

first: Rui

middle: G

last: Martinho


title: Role of ADAR3 in U87 cells

first: Heather

last: Hundley


title: Reduced representation bisulfite sequencing (RRBS) of breast cancer cell lines

summary: Breast cancers can be divided into subtypes with different prognoses and treatment responses based on global gene expression differences.   This study utilized integrated analysis of DNA methylation, chromatin accessibility, transcription factor binding, and gene expression in large collections of breast cancer cell lines and patient tumors to identify transcription factors responsible for the basal-like gene expression program. The results of this study indicate that glucocorticoid receptor (GR) and signal transducer and activator of transcription 3 (STAT3) bind to the same genomic regulatory regions that are specifically open and unmethylated in basal-like breast cancer. These transcription factors cooperate to regulate expression of hundreds of genes in the basal-like gene expression signature and these downstream genes are associated with poor prognosis in patients.

overall_design: To identify regulatory regions specific to basal-like breast cancer, reduced representation bisulfite sequencing (RRBS) was performed on 28 breast cancer cell lines (18 basal-like and 10 luminal) in order to measure DNA methylation across the genome.

first: K-T


title: Transcriptome-wide analysis of RNA editing changes in response to ADAR3 expression in U87 cells

summary: We report  RNA editing changes across the trancriptome in U87 cells that express ADAR3 compared to control cells.

overall_design: Examination of 3 biological replicates of U87 cells expressing ADAR3 compared to control cells

first: Reshma

last: Raghava Kurup

first: Eimile

last: Oakes

first: Aidan

middle: C

last: Manning

middle: A


title: Dynamic regulation of histone modifications and long-range chromosomal interactions during post-mitotic transcriptional reactivation [Hi-C]

first: April

last: Williams

overall_design: Hi-C during mitosis-G1 in U2OS cells

first: Zhichao

last: Xu

first: Jesse

last: Dixon


title: Dynamic regulation of histone modifications and long-range chromosomal interactions during post-mitotic transcriptional reactivation [ChIP-seq]

overall_design: ChIP-seq of H3K4me3, H3K27ac, H3K27ac and CTCF during mitosis-G1 in U2OS and RPE1 cells

middle: N


title: LIN28B Regulates Transcription and Potentiates MYCN-induced Neuroblastoma in a let-7-independent Manner

first: Ting

last: Tao


title: Atg7-dependent autophagy regulates the naive to primed transition by selective Nanog degradation in mouse embryonic stem cells

status: Public on May 20 2020

first: Jing-Jing

last: Zhang

state: Hubei


title: Stress-induced translation inhibition through release of 40S scanning initiation factors

status: Public on Jun 08 2020

city: Edinburgh

first: Stefan

last: Bresson

state: Scotland

address: Max Born Crescent, Swann 5.1

department: Wellcome Trust Centre for Cell Biology

country: United Kingdom

institute: University of Edinburgh

summary: Cellular responses to environmental stress are frequently mediated by RNA-binding proteins (RBPs). Here, we examined global RBP dynamics in Saccharomyces cerevisiae in response to glucose starvation and heat shock. Each stress induced rapid remodeling of the RNA-protein interactome, without corresponding changes in RBP abundance. Consistent with general translation shutdown, ribosomal proteins contacting the mRNA showed decreased RNA-association. Among translation components, RNA-association was most reduced for initiation factors involved in 40S scanning (eIF4A, eIF4B, and Ded1), indicating a common mechanism of translational repression. In unstressed cells, eIF4A, eIF4B, and Ded1 primarily targeted the 5′-ends of mRNAs. Following glucose withdrawal, 5’-binding was abolished within 30sec, explaining the rapid translation shutdown, but mRNAs remained stable. Heat shock induced progressive loss of 5’ RNA-binding by initiation factors over ~16min. Translation shutoff provoked selective 5′-degradation of mRNAs encoding translation-related factors, mediated by Xrn1. These results reveal mechanisms underlying translational control of gene expression during stress.

overall_design:
There are 56 CRAC datasets, and 18 RNAseq datasets. The CRAC datasets were generated using one of three different affinity-tagged proteins: eIF4A, eIF4B, and Ded1. Cells without any tagged proteins ('untagged') were tested as a negative control. For each tagged protein, we tested four different conditions: 1) a 'control' for cells grown at 30° in SC -TRP medium with 2% glucose; 2) a 'mock' shift, in which cells were harvested by filtration, and transferred to identical medium (2% glucose at 30°C); 3) 'GE', in which the cells were collected by filtration and transferred to medium lacking glucose but containing 2% each of glycerol and ethanol; and 4) '42C', in which cells were transferred to medium containing glucose but prewarmed to 42°C. These shifts were performed for either 30 seconds or 16 minutes.  Each CRAC library is associated with a unique number (07-99) to differentiate different replicates. For some analyses (e.g. bedgraph files), different replicates were combined to provide improved coverage across individual transcripts. The relevant replicates are included in the file name. The RNAseq files (A-S) were designed to mimic the conditions used for CRAC and the different conditions and timepoints used are indicated in the file names.

Please note that the processed data generated from merged/multiple samples is linked to the corresponding first sample records (indicated in the file name).

For example, the '08_21_26_71_eIF4B_control_merged*bedgraph' files were generated from the 08, 21, 26 and 71* samples (as indicated in the file name and sample description field) and are linked to the 08_eIF4B_control sample records.

first: David

last: Tollervey


title: Atg7-dependent autophagy regulates the naive to primed transition by selective Nanog degradation in mouse embryonic stem cells [ChIP-seq]

status: Public on May 30 2020

summary: Autophagy is a conserved cellular mechanism to degrade unwanted cytoplasmic proteins and organelles to recycle their components, and it is proved to be critical for embryonic stem cell (ESC) self-renewal and somatic cell reprogramming. However, the role of autophagy in embryonic development remains elusive, and no information exists regarding its functions during the transition from naive to primed pluripotency. Here by using an in vitro transition model of ESCs to epiblast-like cells (EpiLCs), we describe that the dynamic changes in Atg7-dependent autophagy is required for the naive to primed transition, and it is also necessary for germline specification. RNA-seq and ATAC-seq profiling reveal that Nanog acts as a barrier to prevent pluripotency transition, and autophagy-dependent Nanog degradation is important for dismantling the naive pluripotency expression program through decommissioning of naive-associated active enhancers. Mechanistically, we found that autophagy adaptor protein Sqstm1 (p62) is nucleus located during the pluripotency transition period and it is preferentially associated with ubiquitinated Nanog for selective protein degradation. In vivo, loss of autophagy by Atg7 depletion disrupts peri-implantation development and we observed increased chromatin association of Nanog, which affects neuronal differentiation through activation of a subset of neuroectodermal development-associated enhancers. Taken together, our findings illuminate regulatory mechanisms underlying the naive to primed transition and reveal that autophagy-dependent regulation of Nanog is essential for exit from the naive state and marks distinct cell fate allocation during lineage specification.

overall_design: The ChIP-Seq experiments of the Control and Atg7 KO groups were taken at each time point from the 4-day ESC to EpiLC transition.

first: Jilong

last: Zhou

first: Hainan

last: He

first: Jingjing

first: Yi

middle: L

last: Miao


title: Atg7-dependent autophagy regulates the naive to primed transition by selective Nanog degradation in mouse embryonic stem cells [ATAC-seq]

overall_design: The ATAC-Seq experiments of the Control and Atg7 KO groups were taken at each time point from the 4-day ESC to EpiLC transition.


title: Atg7-dependent autophagy regulates the naive to primed transition by selective Nanog degradation in mouse embryonic stem cells [RNA-seq]

overall_design: The RNA-Seq experiments of the Control and Atg7 KO groups were taken at each time point from the 4-day ESC to EpiLC transition.