title: Improving Alignment of Ambiguously Mapping Small RNAs

city: University Park

first: Michael

last: Axtell

address: 208 Mueller Lab

institute: Penn State University

summary: Small RNA sequencing in WT Arabidopsis tissue for analysis of miRNA expression paired with qRT-PCR data.

overall_design: High-throughput sequencing of small RNA in WT Arabidopsis inflorescence from 3 replicate libraries.


title: Regulation of normal B cell differentiation and malignant B cell survival by OCT2

status: Public on May 25 2016

city: Bethesda

first: Louis

middle: M.

last: Staudt

state: MD

address: 9000 Rockville Pike, Bldg 10, Rm 4N114

department: Lymphoid Malignancies Branch

country: USA

institute: National Cancer Institute

type: Genome binding/occupancy profiling by high throughput sequencing

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: The RNA-binding protein TTP is a global post-transcriptional regulator of feedback control in inflammation [iCLIP-seq]

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

country: United Kingdom

institute: Altos Labs

type: Expression profiling by high throughput sequencing

summary: RNA-binding proteins (RBPs) facilitate post-transcriptional control of eukaryotic gene expression at multiple levels. The RBP tristetraprolin (TTP/Zfp36) is a signal-induced phosphorylated anti-inflammatory protein guiding unstable mRNAs of pro-inflammatory proteins for degradation and preventing translation. Using iCLIP, we have identified numerous mRNA targets bound by wild-type TTP and by a non-MK2-phosphorylatable TTP mutant (TTP-AA) in 1h LPS-stimulated macrophages and correlated their interaction with TTP to changes at the level of mRNA abundance and translation in a transcriptome-wide manner. The close similarity of the transcriptome of TTP-deficient and TTP-expressing macrophages upon short LPS stimulation suggested an effective inactivation of TTP by MK2 under these conditions whereas retained RNA-binding capacity of TTP-AA to 3’UTRs caused profound changes in the transcriptome and translatome, altered NF-κB-activation and induced cell death. Increased TTP binding to the 3'UTR of feedback inhibitor mRNAs, such as Ier3, Dusp1 or Tnfaip3, in the absence of MK2-dependent TTP neutralization resulted in a strong reduction of their protein synthesis contributing to the deregulation of the NF-κB-signaling pathway. Taken together, our study uncovers a role for TTP in NF-κB-signaling and highlights the importance of fine-tuned TTP activity-regulation by MK2 in order to control feedback signaling during the inflammatory response.

overall_design: Comparison of the transcriptomes of TTP knockout macrophages inducibly expressing GFP, GFP-TTP or GFP-TTP-AA (S52A, S178A) phosphorylation mutant during 1h LPS stimulation. 3 biological replicates per genotype and condition.

first: Christopher

last: Tiedje

first: Manuel

middle: D

last: Diaz-Muñoz

first: Philipp

last: Trulley

first: Helena

last: Ahlfors

first: Kathrin

last: Laaß

first: Perry

middle: J

last: Blackshear

first: Martin

last: Turner

first: Matthias

last: Gaestel


title: Genome-wide gene/genome dosage imbalance regulates gene expressions in synthetic Brassica napus and derivatives (AC, AAC, CCA, CCAA)

city: Wuhan

first: Chen

last: Tan

address: Huazhong Agricultural University

country: China

institute: College of Plant Science and Technology, Huazhong Agricultural University

summary: Gene/genome dosage balance is an essential evolutionary mechanism for organisms to ensure a normal function, but the underlying causes of dosage-imbalance regulation remains poorly understood. Herein, the serial Brassica hybrids/polyploids with different dosages of A and C subgenomes were synthesized to investigate the effects of genome dosages on gene expressions and interaction using RNA-Seq. The expression changes of A- and C-subgenome genes were in accordance with dosage alterations. Dosage-dependent and -independent genes were grouped according to the correlations between dosage variations and gene expressions. Expression levels of dosage-dependent genes were strongly correlated with dosage changes that mainly contributed to dosage effects, while those of dosage-independent genes gave weak correlations with dosage variations that mostly facilitated dosage compensation. More protein–protein interactions were detected for dosage-independent genes than dosage-dependent ones, as predicted by the dosage balance hypothesis. Dosage-dependent genes more likely impacted the expressions by trans effects, whereas dosage-independent genes preferred to play by cis effects. Furthermore, dosage-dependent genes were mainly associated with the basic biological processes to maintain the stability of the growth and development, while dosage-independent genes were more enriched in the stress response related processes to accelerate adaptation. The present comprehensive analysis of gene expression dependent/independent on dosage alterations in Brassica polyploids provided new insights into gene/genome dosage-imbalance regulation of gene expression.

sample_organism:
Brassica napus
Brassica rapa
Brassica oleracea

overall_design: The newly expanded third leaves without petioles from six plants of each genotype were collected and immediately stored in liquid nitrogen for RNA extraction. Six samples with two replicates were sequenced via  Illumina HiSeqTM 2000.

first: Qi

last: Pan

first: Cheng

last: Cui

first: Xian-hong

last: Ge

first: Zai-yun

last: Li


title: The RNA-binding protein TTP is a global post-transcriptional regulator of feedback control in inflammation [Ribo-seq]


title: The lncRNA LUST promotes CCICs self-renewal stimulating the wnt/b-catenin signaling activation

city: New York

middle: J.

last: Walsh

state: NY

address: One Gustave L. Levy Pl.

department: Structural and Chemical Biology

institute: Mount Sinai School of Medicine


title: Active and inactive enhancers co-operate to exert localized and long-range control of gene regulation [RNA-seq]

status: Public on May 26 2016

first: Jane

last: Skok

address: 550 First Ave, Smillow 307

institute: NYU

summary: We report that in developing B cells individual enhancers of Igk make up super-enhancer cluster where contacts between its components rely on all constituents. Reduction of interaction frequency in enhancer knock-out cells is associated with deminished transcriptional output of enhancers and Igk locus. Moreover, we find that Igk enhancer MiEk has an effect on levels of CBFb enrichment on Tcrb enhancer, Eb afffecting Tcrb recombination and T cell development.

overall_design: Examination of expression, chromatin accessibility, histone modifications and nuclear organization in developing wild-type and Igk and Tcrb enhancer deficient B and T lymphocytes.

first: Valentina

last: Snetkova


title: The RNA-binding protein TTP is a global post-transcriptional regulator of feedback control in inflammation [RNA-seq]

last: Krueger


title: Global RNA recognition patterns of post‐transcriptional regulators Hfq and CsrA revealed by UV crosslinking in vivo

city: Irvine

first: Lei

state: CA

address: University of California, Irvine

institute: University of California, Irvine

summary: The molecular roles of many RNA‐binding proteins in bacterial post‐transcriptional gene regulation are not well understood. Approaches combining in vivo UV crosslinking with RNA deep sequencing (CLIP‐seq) have begun to revolutionize the transcriptome‐wide mapping of eukaryotic RNA‐binding protein target sites. We have applied CLIP‐seq to chart the target landscape of two major bacterial post‐transcriptional regulators, Hfq and CsrA, in the model pathogen Salmonella Typhimurium. By detecting binding sites at single‐nucleotide resolution, we identify RNA preferences and structural constraints of Hfq and CsrA during their interactions with hundreds of cellular transcripts. This reveals 3′‐located Rho‐independent terminators as a universal motif involved in Hfq–RNA interactions. Additionally, Hfq preferentially binds 5′ to sRNA‐target sites in mRNAs, and 3′ to seed sequences in sRNAs, reflecting a simple logic in how Hfq facilitates sRNA–mRNA interactions. Importantly, global knowledge of Hfq sites significantly improves sRNA‐target predictions. CsrA binds AUGGA sequences in apical loops and targets many Salmonella virulence mRNAs. Overall, our generic CLIP‐seq approach will bring new insights into post‐transcriptional gene regulation by RNA‐binding proteins in diverse bacterial species.

sample_organism: Salmonella enterica subsp. enterica serovar Typhimurium str. SL1344

overall_design: To detect the binding sites for Salmonella Hfq and CsrA we collected three biological replicates of Salmonella strain SL1344 where either the hfq or csrA gene was fused to a 3xFLAG tag. Half of each replicate culture was irradiated with UV light (254 nm, 800 mJ/cm2) (+CL), while the other half was left untreated (-CL). Bacteria were lysed and the FLAG-tagged protein was immunoprecipitated using a monoclonal anti-FLAG antibody. The samples were treated with benzonase nuclease, calf intestine phophorylase, and polynucleotide kinase in the presence of radioactive gamma-ATP. Samples were separated with SDS-PAGE followed by transfer to nitrocellulose membranes. Radioactively labelled RNA-protein complexes were eluted from membranes and treated with Proteinase K. Purified RNA was used as input for library preparation using the NEB Next Small RNA Library kit.

first: Erik

last: Holmqvist

first: Patrick

middle: R

last: Wright

first: Thorsten

last: Bischler

first: Lars

last: Barquist

first: Richard

last: Reinhardt

first: Rolf

last: Backofen

first: Jörg

last: Vogel


title: Transcriptional profiles by deep sequencing (RNA-seq) of papillomas generated using the DMBA/TPA protocol from control and transgenic Nanog overexpressing mice

city: Barcelona

first: Manuel

last: Serrano

state: Barcelona

address: Parc Científic de Barcelona, C/ Baldiri Reixac 10

department: Molecular Medicine

country: Spain

institute: IRB

summary: NANOG is a key pluripotency factor in embryonic stem cells that is frequently expressed in squamous cell carcinomas (SCCs). However, a direct link between NANOG and SCCs remains to be established. Here, we show that inducible overexpression of NANOG in mouse skin epithelia dramatically promotes the formation of carcinomas upon chemical carcinogenesis. Gene expression analyses in pre-malignant skin indicate that NANOG induces a large set of genes associated to stemness and to epithelial-mesenchymal transition (EMT).

overall_design: 4 papillomas from different control mice (CTR), and 3 papillomas from different transgenic Nanog overexpressing mice (TG)

first: Adelaida

last: Palla

first: Osvaldo

last: Graña


title: The lncRNA LUST promotes CCICs self-renewal stimulating the wnt/b-catenin signaling activation [RNA-Seq]

first: Martin

summary: Comprehensive RNA-seq experiments in CD24bright/CD44bright (CCICs), CD24dim/CD44dim (more differentiated counterpart) cells and colonospheres delineate the role of the lncRNA LUST  in promoting CCICs self-renewal.

overall_design: RNA-Seq study from HT-29 CD44bright/CD24bright and CD24dim/CD44dim sorted cells subpopulation

first: S

last: Di Cecilia

first: F

last: Zhang

first: W

first: M


title: The lncRNA LUST promotes CCICs self-renewal stimulating the wnt/b-catenin signaling activation [RIP-Seq]

summary: Comprehensive RNA-seq experiments in CD24bright/CD44bright (CCICs), CD24dim/CD44dim (more differentiated counterpart) cells and colonospheres delineate the role of the lncRNA LUST  in promoting CCICs self-renewal

overall_design: Identification of lncRNAs binding to Beta-catenin in two different colon cancer cell lines: HT-29 and CaCo-2


title: The DNA affinity purification DAP-Seq assay [ChIP-seq]

city: La Jolla

first: Joseph

last: Ecker

address: 10010 North Torrey Pines Road

department: Genomic Analysis Laboratory

institute: HHMI-Salk-Institute

summary: As 5-15% of higher eukaryotes genes are transcription factors (TFs), the lack of transcription factor binding site (TFBS) information for most factors in most organisms limits the study of gene regulation. Here we describe a next-generation sequencing method, DNA affinity purification (DAP-Seq), an in vitro gDNA/TF interaction assay that produces whole-genome TFBS annotation for any factor from any organism. Like ChIP-Seq, DAP-Seq resolves TFBS as discrete peaks at genomic locations which allows for accurate motif prediction direct assignment of functionally relevant target genes, and shows better overlap with ChIP-Seq peaks than indirect motif assignment approaches. We applied DAP-Seq to a set of 50 transcription factors in eight Arabidopsis thaliana and one Zea Mays families to gain novel biological insight into TFBS architectures, functions, evolution and methylation-sensitivity.  Overall, DAP-Seq offers a low-cost high-throughput approach to identify TFBS in native sequence context for any organism complete with all DNA chemical modifications.

overall_design: Identification of binding sites for 53 transcription factors (52 Arabidopsis thaliana, 1 Zea mays) by direct sequencing of affinity purified genomic DNA fragments.  Comparison to ChIP-Seq for ABI5 (AT2G36270).

first: Ronan

middle: C

last: O'Malley

first: Shao-shan

last: Huang

first: Liang

last: Song

first: Anna

last: Bartlett

first: Mary

last: Galli

first: Andrea

last: Gallavotti

last: Nery


title: Adenomatous polyps and treatment with Sulindac

status: Public on May 24 2016

city: San Francisco

first: Dvir

last: Aran

institute: University of California San Francisco

summary: Parainflammation is associated with cellular senescence and is apparent in a pure primary epithelial tissue, such as gut epithelial organoids. APC-mutated human and mouse polyps represent early neoplastic lesions, which mostly do not progress, possibly due to senescence-associated PI, thus representing a good source for testing the parainflammation signature we characterized.

overall_design: We prepared organoid cultures from APC-mutated normal gut epithelium (MIN) and adenomatous polyps of APCmin/+ mice (Adenoma) and analyzed them via RNA-seq on three biological replicates. We also prepared organoids from  adenomatous polyps of APCmin/+ mice, treated them with the NSAID Sulindac and performed RNA-seq on three biological replicates

first: Yinon

last: Ben-Neriah

first: Audrey

last: Lasry


title: Regulation of DNA methylation landscape in human somatic cell reprogramming by miR-29 family (RNA-seq)

city: New Haven

first: Eriona

last: Hysolli

state: CT

address: 10 Amistad Street

department: Genetics

institute: Yale University

summary: Reprogramming to pluripotency after overexpression of OCT4, SOX2, KLF4 and MYC is accompanied by global genomic and epigenomic changes. Histone modification and DNA methylation states in iPSCs have been shown to be highly similar with embryonic stem cells (ESCs). However, epigenetic differences still exist between iPSCs and ESCs. In particular, aberrant DNA methylation states found in iPSCs are a major concern for using iPSCs in a clinical setting. Thus, it is critical to find factors that regulate DNA methylation states in reprogramming. Here, we found that the miR-29 family is an important epigenetic regulator during human somatic cell reprogramming. Our global DNA methylation and hydroxymethylation analysis shows that DNA demethylation is a major event mediated by miR-29a depletion during early reprogramming, and that iPSCs derived from miR-29a depletion are epigenetically closer to ESCs. Our findings uncover an important miRNA-based approach to generate clinically robust iPSCs.

overall_design: Bisulphite converted gDNAs of D551 fibroblasts transduced for 3 days with overexpression of DNMTs, TETs, TDG and OSKM or miR29a/b/c and control sponge were hybridized into Illumina Infinium HumanMethylation 450K Beadchip.

first: Yoshiaki

last: Tanaka

first: In-Hyun

last: Park


title: Regulation of DNA methylation landscape in human somatic cell reprogramming by miR-29 family (MeDIP-seq)

type: Methylation profiling by high throughput sequencing


title: Regulation of DNA methylation landscape in human somatic cell reprogramming by miR-29 family


title: NOVA2 and NOVA1 HITS-CLIP in wild-type mouse cortex (E18.5)

first: Yuhki

last: Saito

address: 1230 York avenue

institute: The Rockefeller University

summary: Mouse cortex samples from E18.5 wild-type were subjected to HITS-CLIP to detect NOVA2 and NOVA1 binding positions on RNA

overall_design: Mouse cortex samples were UV-irradiated and subjected to NOVA2 and NOVA1 HITS-CLIP (detailed desription in accompanying paper)

first: Robert

middle: B

last: Darnell


title: Multiple levels of transcriptional regulation by PLZF in NKT cell development [mouse_ChIP-seq]

city: Chicago

first: Aiping

last: Mao

state: IL

address: 929 East 57th street

department: Pathology

institute: University of Chicago

summary:
Using biotinylation-based ChIP-seq and microarray analysis of both NKT and PLZF-transgenic thymocytes, we identified several layers of regulation of the innate-like NKT effector program.
The transcription factor PLZF is induced during the development of innate and innate-like lymphocytes to direct their acquisition of a T helper effector program, but the molecular mechanisms involved are poorly understood. Using biotinylation-based ChIP-seq and microarray analysis of both NKT and PLZF-transgenic thymocytes, we identified several layers of regulation of the innate-like NKT effector program: first, PLZF bound and regulated genes encoding cytokine receptors as well as homing and adhesion receptors; second, PLZF bound and activated T helper-specific transcription factor genes that in turn control T helper specific programs; finally, PLZF bound and suppressed the transcription of Bach2, a potent general repressor of effector differentiation in naive T cells. These findings reveal the architecture of the transcriptional program recruited by PLZF and elucidate how a single transcription factor can drive the developmental acquisition of a broad effector program.

overall_design: Examination of PLZF binding sites in both NKT and PLZF-transgenic thymocytes

first: Albert

last: Bendelac


title: The RNA-binding protein TTP is a global post-transcriptional regulator of feedback control in inflammation

first: Felix

last: Krueger


title: Multiple levels of transcriptional regulation by PLZF in NKT cell development


title: Genome-wide DNA methylation and mRNA and the microRNA transcriptome profiling between oxidative and glycolytic skeletal muscles [miRNA-seq]

city: Chengdu

first: Linyuan

last: Shen

state: Sichuan

address: 211# Huiming Road

department: College of Animal Science and Technology

institute: Sichuan Agricultural University

type: Non-coding RNA profiling by high throughput sequencing

summary: The physiological, biochemical and functional difference of oxidative and glycolytic muscles play important roles in human metabolic health as well as in meat quality of animal. To explore this aspect, we firstly provided a genome-wide landscape of DNA methylomes and their relationship with mRNA and miRNA transcriptome for PMM (psoas major muscle; oxidative) and LDM (longissimus dorsi muscle; glycotic) in this study. We observed hypomethylation of subtelomeric regions and high mitochondria content contributed to fast replicative senescence in PMM. The DMRs in promoter (478) and gene bodies (5718) were main enriched in GTPase regulator activity and signaling cascade mediated pathway. Integration analysis revealed methylation status within gene promoter (or gene body) and miRNA promoter have significant negative correlation with mRNA and miRNA expression, respectively. Which included numerous genes were closely related to distinct phenotypic traits between LDM and PMM. For instance, the hypermethylation and down-expression of HK-2 and PFKFB4 decreased the glycolytic potential in PMM. In addition, we validated the promoter hypomethylation and up-expression of miR-378 results in silencing the target gene expression and inducing promoted capillaries biosynthesis in PMM. Together, these results provide a thorough understanding of muscle metabolism and development from an genomic and epigenetic perspective.

overall_design: The aim of our study is to identify miRNAs that play essential roles in the phenotypic variations in oxidative and glycolytic skeletal muscles, and integrate mRNA transcriptome and methylation data to explore the underlying molecular mechanisms.

first: Zhendong

last: Tan

first: Mingzhou

last: Li

first: Shunhua

last: Zhang

first: Li

last: Zhu


title: Genome-wide DNA methylation profiling between oxidative and glycolytic skeletal muscles [MeDIP-seq]

overall_design: The aim of our study is to identify different methylated genes that are linked to the phenotype of physiological, biochemical and functional variation in oxidative and glycolytic skeletal muscles

first: Qiong

last: Yang

first: Xuewei


title: Allele-specific methylome analysis reveals cis-regulation of DNA methylation in Nasonia [WGBS-seq]

city: Ithaca

first: Andrew

last: Clark

address: 227 Biotech Building

institute: Cornell University

summary: Wasps of the genus Nasonia are non-social parasitoids that are emerging as a model for studies of epigenetic processes in insects. To study the transmission pattern of DNA methylation and changes of methylation during species divergence, we scored genome-wide methylation and allele-specific methylation (ASM) percentages at single CpG level in both parental and reciprocal F1s of Nv and Ng. These closely related cross-fertile species are genetically tractable and inbred in nature, allowing accurate estimation of ASM. Among the ~7000 differentially methylated CpGs (DMCpGs) between Nv and Ng parents, 4364 were covered with >8x for both Nv and Ng alleles in reciprocal F1s and they clustered in 178 genes (DM-genes) with >4 DMCpGs per gene. Surprisingly, all 178 DM-genes are differentially methylated between the two alleles in F1s, remembering the parental methylation status with nearly 100% fidelity, suggesting the presence of strong cis-elements driving the target of gene body methylation. The 100% cis-regulation in F1 suggests the methylation machinery is conserved and DNA methylation is targeted by cis-motifs in Nasonia. In addition, we discovered that total and allele-specific expression is positively correlated with ASM in a subset of the DM-genes. Unlike in mammals, we did not find any parent-of-origin DMRs in Nasonia, with paternal or maternal-specific methylation, and there seemed to be no epigenetic reprogramming with two waves of de- and re-methylation. These results will shed light on the mechanism of DNA methylation in insects and the evolution of methylation in animals.

overall_design: Whole-genome bisulfite sequencing of 24-hour reciprocal F1 adults of Nasonia vitripennis and Nasonia giraulti using Illumina sequencing.

middle: G

first: John

middle: H

last: Werren

first: Xu

last: Wang


title: RNA-Seq of SLNCR1 over-expression in the melanoma cell line A375

city: Boston

first: Carl

last: Novina

state: MA

address: 450 Brookline Ave

department: Cancer Immunology

institute: Dana-Farber Cancer Institute

summary: RNA-Seq was used to profile transcriptional changes induced by overexpression of the long non-coding RNA SLNCR1, as well as mutant version SLNCR1 delta conserved and SLNCR1 conserved.

overall_design: The A375 melanoma cell line was transfected with pcDNA3.1 (-) expressing either full length SLNCR1, SLNCR1 delta conserved, or SLNCR1 conserved.

first: Karyn

last: Schmidt


title: RNA-Seq of siRNA-mediated knockdown of the lncRNA SLNCR1 in WM1976 melanoma cells

summary: RNA-Seq was used to profile transcriptional changes induced by siRNA knockdown of the long non-coding RNA SLNCR1.

overall_design: The WM1976 melanoma short-term culture was transfected with either scrambled or SLNCR1-targeting siRNAs


title: Dual function of Med12 in PRC1-dependent gene repression and ncRNA-mediated transcriptional activation

city: Mainz

first: Sergi

last: Sayols

address: Ackermannweg 4

country: Germany

institute: Institute of Molecular Biology, Mainz

summary: Mediator is regarded a general co-activator of RNA-Polymerase II dependent transcription but not much is known about its function and regulation in mouse pluripotent embryonic stem cells (mESC). One means of controlling Mediator  function is provided by binding of the Cdk8 module (Med12, Cdk8, Ccnc and Med13) to Mediator. Here we report that the Cdk8 module subunit Med12 operates together with PRC1 to silence developmental key genes in the pluripotent state. At the molecular level, PRC1 is required to assemble ncRNA containing Med12-Mediator complexes at promoters of repressed genes. In the course of cellular differentiation the H2A-ubiquitin binding protein Zrf1 abrogates PRC1-Med12 binding and facilitates the recruitment of Cdk8 into Mediator. Remodeling of the Mediator-associated protein complex converts Mediator into a transcriptional enhancer that mediates ncRNA-dependent activation of Polycomb target genes

overall_design: RNAseq of  pluripotent (control, shNMC, shRing1b, shMed12, shCdk8, shZrf1) and early differentiating (control, shNMC, shMed12, shCdk8, shZrf1) stem cells in triplicates. Control would be normal E14TG2A mESCs. shNMC refers to E14TG2A cells stably transfected with a short hairpin that has no mammalian targets (Non Mammalian Control). All the other samples are indeed stably transfected with short hairpins against the indicated genes.

first: Thaleia

last: Papadopoulou

first: Richly

last: Holger


title: Gene expression profile in the reicprocal F1 hybrids of two jewel wasp species Nasonia vitripennis and Nasonia giraulti [RNA-seq]

summary: We quantified genome-wide total and allele-specific expression in two non-social parasitoids wasp species Nasonia vitripennis and Nasonia giraulti and their reciprocal F1 hybrids. No parent-of-origin effect in allelic expression was found for >8,000 informative genes, suggesting lack of genomic imprinting in adult Nasonia. Gene expression divergence between Nv and Ng could be attributed to both significant cis- and trans- regulatory changes during evolution.

overall_design: Profiling of expression levels in reciprocal F1 adults of Nasonia vitripennis and Nasonia giraulti using Illumina mRNA-seq


title: Sonic hedgehog signaling promotes basal radial glia and 1 intermediate progenitor cell

status: Public on May 23 2016

city: Memphis

first: David

last: Finkelstein

state: TN

address: 332 N. Lauderdale St.

department: Computational Biology

institute: St Jude Children's Research Hospital

summary: The unique mental abilities of humans are rooted in the immensely expanded and folded neocortex, which reflects the expansion of neural progenitors, especially basal progenitors including basalradial glia (bRGs, also called outer RGs) and intermediate progenitor cells (IPCs). Here, we show that constitutively active Shh signaling expanded basal progenitors and induced folding in the otherwise smooth mouse neocortex, whereas the loss of Shh signaling decreased the number of basal progenitors and the size of the neocortex. SHH signaling was strongly active in the human fetal neocortex but not in the mouse embryonic neocortex, and blocking SHH signaling in humancerebral organoids decreased the number of bRGs. Mechanistically, Shh signaling increased theinitial generation and self-renewal of bRGs as well as increasing IPC proliferation. Thus, robust SHH signaling in the human fetal neocortex may contribute to basal progenitor expansion and neocortical growth and folding.

overall_design: 2 normal WT lateral cortical embrynonic E14.5 samples are compared to 3 medial samples in mouse

first: Lei

first: Shirui

last: Hou

first: Young-Goo

last: Han


title: Allele-specific transcriptome and methylome analysis reveals cis-regulation of DNA methylation and lack of genomic imprinting in Nasonia

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

overall_design: Refer to individual Series


title: Fra-1 is a key driver of colon cancer metastasis and a Fra-1 classifier predicts disease-free survival

city: Amsterdam

first: Oscar

last: Krijgsman

state: Outside the US or Canada

address: Plesmanlaan 121

department: Molecular Oncology and Immunology

country: Netherlands

institute: Netherlands Cancer Institute

summary:
Background and Aim: Fra-1 (Fos-related antigen-1) is a member of the AP1 (activator protein-1) family of transcription factors. We have recently shown that Fra-1 is necessary for breast cancer cells to metastasize in vivo, and that breast cancer outcome can be predicted by a classifier comprising genes that are expressed in a Fra-1-dependent fashion. Here, we show that Fra-1 plays an important role also in colon cancer progression.

Methods: We compared proliferation rates of parental and Fra-1-depleted colon cancer cells in vitro under 2D, 3D, and attachment-free conditions and in vivo upon subcutaneous and intravenous injections into mice. We also compared RNA expression profiles of colon cancer cells with and without Fra-1 expression.

Results: Fra-1 depletion impair colony outgrowth of human colon cancer cells in soft agar and in suspension, whereas it does not affect proliferation on 2D culture plates. Consistent with this, upon subcutaneous injection into mice, tumors formed by Fra-1-depleted colon cancer cells are only three times smaller than those produced by control cells. In contrast, when injected intravenously, Fra-1 depletion causes 200-fold reduction in tumor burden. Consistent with the more aggressive characteristics of Fra-1-proficient tumors, the prognosis of colon cancer patients can be predicted by a Fra-1 classifier generated by comparing RNA profiles of parental and Fra-1-depleted colon cancer cells.

Conclusions: Our results demonstrate that Fra-1 is an important determinant of the metastatic potential of human colon cancer cells, and suggest that a Fra-1 classifier can be used as a prognostic predictor in colon cancer patients.

overall_design: HT29 cell line, two shRNAs against Fra-1, one empty vector control, three biological replicates

first: Sedef

last: Iskit

first: Daniel

last: Peeper


title: CENP-T and CENP-C bridge adjacent CENP-A nucleosomes on young alpha-satellite dimers

city: Seattle

first: Jorja

last: Henikoff

state: WA

address: 1100 Fairview AV N, A1-162

department: Basic Sciences

institute: Fred Hutchinson Cancer Research Center

summary: We find that CENP-T acts as a bridge between two well-positioned CENP-A nucleosomes that are present on young alpha-satellite dimers that dominate functional human centromeres. CENP-T is centered over the CENP-B box, where it interacts with the CENP-B/CENP-C complex. Upon cross-linking, the entire CENP-A/CENP-C/CENP-T-containing complex is recovered as a nuclease-protected particle over an alpha-satellite dimer that comprises the fundamental unit of kinetochore chromatin. Our work reveals that CENP-A/CENP-C and CENP-T branches of kinetochore assembly are physically integrated.

overall_design: We used chromatin immunoprecipitation with sequencing (ChIP-seq) in human cells to determine the relative positions of CENP-A and CENP-T particles at functional centromeres.

first: Jitendra

last: Thakur

first: Steven


title: HCT116 MYC 3' TBE1 (WT) and KO RNA-Seq

city: Hershey

first: Yuka

last: Imamura Kawasawa

state: PA

address: 500 University Dr.

department: College of Medicine, Pharmacology

institute: Penn State University

summary: mRNA was sequenced from HCT116 MYC 3' TBE1 (WT) and KO cells to identify genes differentially expressed after deletion of the MYC 3' TBE1

overall_design: mRNA levels from two biological replicates of HCT116 MYC 3' TBE1 (WT) and KO cells were examined

first: Sherri

middle: A

last: Rennoll

first: Wesley

middle: M

last: Raup-Konsavage

middle: I

last: Kawasawa

first: Gregory

middle: S

last: Yochum


title: RNA-seq analysis of growth factor response of NHEKs to antimicrobial petide LL-37 and dsRNA mimic Poly(I:C)

city: San Diego

first: Christopher

last: Adase

address: 2525 John Hopkins Ct

department: Dermatology

institute: UCSD

summary: In this study, we analyzed how non-coding double stranded RNA (dsRNAs) act as a damage associated molecular pattern (DAMP) in the skin, and how the human cathelicidin AMP LL-37 might influence growth factor production in response to this DAMP.

overall_design: Each sample's RNA was isolated form a single biological source of P6 NHEKs. In total there are 4 samples (non-replicates); Control (PBS treated), 1.75uM LL-37 treated, 0.1ug/ml Poly(I:C) treated, and co-treated with 1.75uM LL-37 and 0.1ug/ml Poly(I:C).

first: Andrew

middle: W

last: Borkowski

first: Ling-juan

last: Zhang

first: Michael

last: Williams

first: Emi

last: Sato

first: James

last: Sanford

first: Richard

middle: L

last: Gallo


title: Histone deacetylase inhibition enhances antimicrobial peptide but not inflammatory cytokine expression upon bacterial challenge

city: Paris

first: Brice

last: Sperandio

address: 28, rue du Docteur Roux

department: Biologie Cellulaire et Infection

country: France

institute: Institut Pasteur

summary: We report expression of genes from human Caco-2 cells (subclone TC7) after inhibition of histone deacetylases by trichostatin A, upon an E. coli bacterial challenge

overall_design: RNAseq on cells pretreated or not with 5 uM TSA and challenged or not for 2 hours with E. coli

first: Natalie

last: Fischer


title: Genome-wide maps of SIX1, SIX2, and active loci in human fetal kidneys generated from ChIP-seq data.

city: Edinburgh

first: GUDMAP

last: Developers

address: Crewe Road

institute: IGMM MRC Human Genetics Unit

summary: SIX2 is expressed by the self-renewing nephron progenitors in the human fetal kidney. We have also discovered that SIX1 is expressed in nephron progenitor population of the human fetal kidney, which is in contrast to the mouse. We performed ChIP-seq  of SIX1 and SIX2 in order to identify the target genes of each factor and compare the role that each factor plays in transcriptional regulation of the nephron progenitors. We additionally performed ChIP-seq for p300 and H3K27ac in order to identify active loci and complement the transcription factor data.

overall_design: Microdissection techniques were used to isolate cells from the renal cortex of the human fetal kidney at 16-17 weeks of gestation.  Specific antibodies for SIX, SIX2, p300 & H3K27ac were used for chromatin immunoprecipitation, and the recovered DNA subjected to high-throughput sequencing.

first: Lori

last: O'Brien

first: Qiuyu

last: Guo

last: McMahon


title: RNA-seq analysis of the human fetal kidney.

summary: Micro-dissection and FACS were used to isolate tissue and cells, respectively, for RNA-seq analyses of the developing human kidney. Specifically, ITGA8 and ROR2 antibodies were used to isolate cells of the nephron progenitor lineage, and representative tissue samples of the outer cortex and whole kidney were collected, to obtain information on the transcripts expressed within these cell populations. These data will further our understanding of human kidney development.

overall_design: Total RNA was obtained from micro-dissected and FACS isolated cells from 15-17 week human fetal kidneys. ITGA8 and ROR2 were used to enrich for nephron progenitors. Micro-dissected samples were representative of the outer cortex and whole kidney. Isolated RNA was subjected to high throughput sequencing analyses to identify expressed transcripts.

first: Peter

last: Whitney


title: MIRA-Seq analysis of bovine dermal fibroblasts isolated from two breeds of cattle, Holstein (dairy breed) and Angus (beef breed).

city: Burlington

first: David

last: Kerr

state: VT

address: 570 Main Street

department: Animal and Veterinary Sciences

institute: University of Vermont

summary: We report the results of MIRA-Seq based high-throughput profiling of the bovine dermal fibroblast methylome from two different breeds of cattle (n=4/breed) to determine the breed-dependent differences in methylation.

overall_design: Fibroblasts were isolated from 4 animals of each cattle breed (Angus and Holstein). Cells were not treated.

first: Aimee

last: Benjamin

first: Benjamin

last: Green

first: Stephanie

last: McKay

first: Brian

last: Crooker

middle: E


title: Breed-dependent differences in transcriptomic response of dermal fibroblasts to lipopolysaccharide

summary: We report the breed-dependent differences in the innate immune response of dermal fibroblasts following exposure to lipopolysaccharide (LPS). Skin biopsies were collected from Angus (a beef breed) and Holstein (a dairy breed) animals and isolated fibroblasts were exposed to LPS to investigate differences in gene expression between these two cattle breeds.

overall_design: Fibroblast were isolated from 4 animals of each breed (Angus and Holstein) and treated with LPS for 0 hrs, 2 hrs, and 8 hrs. RNA was extracted at each of these times points.


title: Differential responsiveness of Holstein and Angus dermal fibroblasts to LPS challenge occurs without major differences in the methylome.


title: Finding missing proteins from epigenetically manipulated human cells

city: Guangzhou

first: Gong

last: Zhang

address: Huang-Pu Avenue West 601

department: Institute of Life and Health Engineering

institute: Jinan University

summary: We performed transcriptome sequencing to assess the genes turn on in the cell under epigenetically manipulation. MHCC97H, A549 and HCT116 were treated by GSK126, S2101 and TSA respectively. We detected the missing proteins in the cells under epigenetically manipulation.

overall_design: A549 and MHCC97H cells were cultured in the complete DMEM medium and epigenetically manipulation (1 μM TSA, 50 μM GSK126 and 100μM S2101 respectively); HCT116 cells were maintained in complete RPMI 1640 medium and epigenetically manipulation (1 μM TSA, 50 μM GSK126 and 100μM S2101 respectively). We detected and analyzed the missing protein with transcriptome and proteome.

first: Xinlei

last: Lian


title: High-throughput functional comparison of promoter and enhancer activities

first: Thomas

middle: Andy

last: Nguyen

address: 77 Avenue Louis Pasteur

institute: Harvard Medical School

summary: Promoters initiate RNA synthesis, and enhancers stimulate promoter activity. Whether promoter and enhancer activities are encoded distinctly in DNA sequences is unknown. We measured the enhancer and promoter activities of thousands of DNA fragments transduced into mouse neurons. We focused on genomic loci bound by the neuronal activity-regulated co-activator CREBBP, and we measured enhancer and promoter activities both before and after neuronal activation. We find that the same sequences typically encode both enhancer and promoter activities. However, gene promoters generate more promoter activity than distal enhancers, despite generating similar enhancer activity. Surprisingly, the greater promoter activity of gene promoters is not due to conventional core promoter elements or splicing signals. Instead, we find that particular transcription factor binding motifs are intrinsically biased toward the generation of promoter activity, while others are not. While the specific biases we observe may be dependent on experimental or cellular context, our results suggest that gene promoters are distinguished from distal enhancers by specific complements of transcriptional activators.

sample_organism:
Mus musculus
unidentified plasmid

platform_organism:
Mus musculus
unidentified plasmid

overall_design: 44 RNA-seq samples, 8 ChIP-seq samples, 8 DNA-seq samples

last: Jones

first: Andrew

last: Snavely

first: Andreas

last: Pfenning

first: Rory

last: Kirchner

first: Martin

last: Hemberg

first: Jesse

last: Gray


title: Genome-wide DNA methylation and mRNA and the microRNA transcriptome profiling between oxidative and glycolytic skeletal muscles

first: Linyuan

last: Shen


title: The Proteasome Stress Regulon is Controlled by a Pair of NAC Transcription Factors in Arabidopsis

status: Public on May 21 2016

city: Cold Spring Harbor

first: Nicholas

last: Gladman

address: 1 Bungtown Rd

department: ARS

institute: USDA

summary: We report the expression differences between 5DAG Arabidopsis Col-0, rpn10-1, and rpn12a-1 seedlings treated with 100 uM MG132 for 0, 3, or 24 hours.

overall_design: Examination of 3 different Arabidopsis seedling genotypes (Col-0 (wild-type), and the exon trap mutants rpn10-1 and rpn12a-1) treated with 100 uM MG132 for either 0, 3, or 24 hours.

middle: P

first: Richard

last: Vierstra


title: Ablation of the epithelial-specific splicing factor Esrp1 results in ureteric branching defects and reduced nephron number

city: Los Angeles

first: Yi

last: Xing

address: 650 Charles E. Young Dr. South

department: Department of Microbiology, Immunology, & Molecular Genetics

institute: UCLA

summary: Evaluation of splicing in ureteric epithelium in Esrp1;Esrp2 double knockout mice compared to Esrp1+/+;Esrp2-/- control mice.

overall_design: Ureteric epithelial cells were isolated from E18.5 embryos using FITC-DBA lectin and fluorescence activated cell sorting (FACS)

first: Russ

last: Carstens


title: Overexpression of a B3 Transcription Factor CsFUS3 Promotes Somatic Embryogenesis of citrus through the Abscisic Acid and Gibberellin

city: Wuhan

first: Liu

last: Zheng

institute: Huazhong Agricultural University

summary: To future examine more globally genes that may be responsive to CsFUS3 accumulation, we used RNA-seq to compare the transcriptome profiles of the overexpression line with that of empty vector line at two representative time points during SE. After removing low-quality reads, about 13 million reads per sample were mapped against sweet orange reference genome. Using FDR≤0.001 and Abs [log(fold-change)] ≥1 as a selection threshold, a total of 329 genes were found differentially expressed in the CsFUS3 OE line after 6 weeks of induction, including 227 up-regulated and 102 down-regulated genes. Of the 1,895 DEGs at 12 weeks, 1,165 were up-regulated and 730 down-regulated in OE line, indicating a large transcriptional reprogramming in the transgenic callus at later stages of SE RNA-Seq transcriptome profiling revealed that a number SE-promotive TF genes were up-regulated in the overexpression line. In addition, the expression of pathway genes involved in biosynthesis, deactivation, and signaling of ABA and GA was altered.

overall_design:
Compare the transcriptome profiles of the overexpression line with that of empty vector line at two representative time points during SE: the onset of the somatic embryo emergence (12 weeks after induction), and the earlier time point when the colour just began to change (6 weeks after induction). Libraries were sequenced by lon Proton platform, two biological replicates for each sample.
Please note that the previous study (PMID 24489955) has done annotation project of Citrus sinensis.The database web link is http://citrus.hzau.edu.cn/. However, this web link is not functional yet. Thus the annotation .gff3 file is provided in the meantime.

first: Guo

last: Wenwu


title: Transcriptome profiles and comparison of Arabidopsis thaliana woody (soc1-ful) and WT (Col-0) stems.

city: Leiden

first: Nicolas

last: Davin

address: Darwingweg 4

department: Botany

institute: Naturalis

summary: The douple mutant Arabidopsis thaliana soc1 ful, in contrast with WT, produces an interfascicular cambium and a large wood cylinder is the flowering stem. We present the RNAseq data for polyA mRNA of different developmental stages of cambium and wood formation in Arabidopsis thaliana. We sequenced 7 stages; 4 in the woody mutant soc1-6 ful-7 (herbaceous, cambium initiation, wood initiation and leaf) and 3 stages in the WT Col-0 (herbaceous, cambium and leaf). The corresponding stem anatomy is also presented in the manuscript indicating the stage of cambium development and the production of secondary xylem.

overall_design: Sequencing of 7 differents stem developmental stage and leaf using 3 biological replicates.

first: Frederic

last: Lens


title: Profiling of Mll1 in mouse epiblast stem cells and H3K4me1/3

city: Ann Arbor

first: Yali

last: Dou

state: MI

address: 1301 Catherine Street

department: Pathology Department

institute: University of Michigan

summary: This study describes the epigenetic profiling of 1) Mll1 in mouse EpiSCs with or without MM401 tratment and 2) H3K4me1 and H3K4me3 in MM401 trated EpiSCs.

overall_design: ChIP-Seq profiles of Mll1 were generated in mEpiSCs with or without MM401; ChIP-Seq profiles of H3K4me1/3 were generated in EpiSCs treated with MM401 for 4 days. For each condition, a sequence profile of genomic DNA (Input) is also included.


title: Natural variation in non-coding regions underlying phenotypic diversity in budding yeast

city: Santiago

first: Francisco

last: Cubillos

address: Obispo Manuel Umaña 050, Edificio de Alimentos, Estación Central

department: Departamento de Ciencia y Tecnología de los Alimentos

country: Chile

institute: Universidad de Santiago de Chile

summary: Determining the different sources of heritable variation underlying quantitative traits in nature is currently at the forefront of genetic studies. To this end, molecular profiling studies in S. cerevisiae have shown that individual gene expression levels are subject to genetic control and this variation can mediate genetic differences on phenotype. Thus, determining how natural variation influences allele specific expression (ASE) and ultimately complex traits represents a useful tool to determine the mechanisms leading to yeast niche adaptation. Here, in order to test the hypothesis that allele-specific expression differences between isolates contributes to the phenotypic diversity in natural populations, we evaluated ASE levels in a grid of six F1 hybrids from the cross of four representative founder strains from major lineages. Genome-wide and across hybrids we quantified ASE for 3,320 genes. We found evidence for abundant genome-wide expression differences between alleles, with levels ranging between 27% up to 61% of the evaluated genes, depending on the cross. We observed that ASE can be explained by allele-specific differences in transcription factor binding to cis-regulatory regions and differences in strain-specific trans-activation can be detected by taking advantage of the shared trans environment of F1 hybrids. Furthermore, modules of genes under cis-regulatory variation with related function are enriched within the different genetic backgrounds, supporting the premise of intraspecies directional regulatory selection in yeast. Finally, we were able to identify two genes, GDB1 and ASN1 exhibiting high expression levels in the Wine/European strain and underlying phenotypic differences for oenological phenotypes due to polymorphisms within non-coding regions, providing direct evidence of the importance of regulatory variation in natural trait diversity.

overall_design: ASE analysis in 6 F1 hybrids in triplicates


title: AR and ERG ChIP-seq in presence or absence of PRMT5

status: Public on May 20 2016

city: Boston

first: Thomas

last: Westerling

address: 450 Brookline Ave

department: Medical Oncology

institute: Dana Farber Cancer institute

summary: Replicates of ChIP seq data using AR and ERG antibodies from VCaP cell lines harboring inducible shRNA constructs for PRMT5 (3 independent, sh1, sh2, sh3) and 1 non targeting control (NTC), all sample stimulated with ligand, either DHT or R1881

overall_design: AR ChIPseq in VCaP cells, 3 replicates each of DHT stimulated cells with one of 4 inducible shRNA constructs, sh1, sh2, sh3 targeting PRMT5 or NTC non targeting controls. Furthermore 2 AR ChIPseq in the same system stimulated with R1881 for targeting constructs sh1, sh2 and NTC. ERG ChIPseq 2 replicates DHT stimulated, each of sh1, sh2, sh3 constructs. Cells where induced (doxycyclin) for shRNA expression on day0, hormone starved on day3 and harvested on day5.

first: Zineb

last: Mounir

first: Raymond

last: Pagliarini

first: Myles

last: Brown


title: RNA-Seq of over-expression and knockdown of the lncRNA SLNCR1 in melanoma cells

first: Carl

last: Novina

institute: Dana-Farber Cancer Institute

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

overall_design: Refer to individual Series


title: Genome-wide analysis of Alternaria stress-responsive microRNAs in tomato

city: Kolkata

first: Pallob

last: Kundu

state: West Bengal

address: P 1/12 CIT Scheme VIIM

department: Division of Plant Biology

country: India

institute: Bose Institute

summary: MicroRNAs are crucial regulator of reprogramming of gene expression cascade during plant-pathogen interaction. We have used tomato (Pusa Ruby) plant and early blight pathogen, Alternaria for the analysis of tomato miRNA expression profiles in a compatible interaction. Illumina next generation sequencing (NGS) technique based whole transcriptome analysis revealed that, (i) about 188 known miRNAs, ranging from 18nt to 24nt expressed in tomato, which belonged to 124 miRNA families and (ii) both conserved and Solanaceae specific miRNAs were differentially expressed. Most of the miRNAs were down-regulated, and around 7 miRNAs were highly differentially regulated (log2FC ≥ ±3). Furthermore, using stringent selection criteria we could detect approximately 74 putative novel miRNAs. GO terms enrichment and KEGG pathway analyses of predicted targets of differentially expressed miRNAs have been performed to identify the pathways that were perturbed during the infection. Supported by DBT, Govt. of India.

overall_design: miRNA profile of Indian tomato (Solanum lycopersicum) cultivar Pusa Ruby infected with Alternaria fungus.  Two mock infected and two Alternaria fungus infected samples were analyzed.

first: Deepti

last: Sarkar

first: Ranjan

middle: K

last: Maji

first: Zhumur

last: Ghosh


title: Capturing the dynamic nascent transcriptome during acute cellular responses: the serum response

first: Mats

last: Ljungman

state: Michigan

address: NCRC, B520 Room 1346 2800 Plymouth Rd.

summary: Dynamic regulation of gene expression via signal transduction pathways is of fundamental importance during many biological processes such as cell state transitioning, cell cycle progression and stress responses. In this study we used serum stimulation as a cell response paradigm to apply the nascent RNA Bru-seq technique in order to capture early dynamic changes in the nascent transcriptome. Our data provides an unprecedented view of the dynamics of genome- wide transcription during the first two hours of serum stimulation in human fibroblasts. While some genes showed sustained induction or repression, other genes showed transient or delayed responses. Surprisingly, the dynamic patterns of induction and suppression of response genes showed a high degree of similarity, suggesting that these opposite outcomes are triggered by a common set of signals. As expected, early response genes such as those encoding components of the AP-1 transcription factor and those involved in the circadian clock were immediately but transiently induced. Surprisingly, transcription of important DNA damage response genes and histone genes were rapidly repressed.We also show that RNA polymerase II accelerates as it transcribes large genes and this was independent of whether the gene was induced or not. These results provide a unique genome-wide depiction of dynamic patterns of transcription of serum response genes and demonstrate the utility of Bru-seq to comprehensively capture rapid and dynamic changes of the nascent transcriptome.

overall_design: One cell line was used (normal human foreskin fibroblasts; HF1, referred to as "nf" in the sample names). HF1 cells were serum-starved for 48 h. One sample ("starved") was then labeled with bromouridine (2 mM) for 30 min.  For the remaining samples, serum was added and bromouridine added immediately ("30 min serum") or 30 min, 60 min, or 90 min post serum addition ("60 min serum", "90 min serum", and "120 min serum", respectively).  RNA was collected immediately after the 30 min bromouridine labeling period.

first: Killeen

last: Kirkconnell

first: Michelle

middle: T

last: Paulsen

first: Brian

last: Magnuson

first: Karan

last: Bedi


title: The midguts transcriptome of Aedes aegypti (Liverpool strain) females blood fed DENV2 NS1 versus those blood fed BSA

city: beijing

first: jianying

last: Liu

state: beijing

address: Room 4-305,Biotechnology building, Tsinghua University, Beij

institute: tsinghua university

summary: We report the RNA-seq based analyses of the transcriptional changes in the Aedes aegypti midguts transcriptome 4,8,18 hours after blood feeding BSA or DENV2 NS1.

overall_design: Comparison of the midguts transcriptome of Aedes aegypti females at two blood feeding conditions and three time points.

first: Jianying

first: Yang


title: Effects of biobanking on chromatin accessibility

status: Public on May 22 2016

city: Atlanta

first: Chris

last: Scharer

state: GA

address: 1510 Clifton Rd, Suite 3086A

department: Microbiology and Immunology

institute: Emory University

summary: The goal of this study was to test the effects of biobanking on chromatin assessibility, as defined by ATAC-seq.  Human naïve B cells were FACS sorted from PBMCs that were either frozen or processed fresh.  A range of cell numbers were used to identify technical limitations and reproducibility of the ATAC-seq assay on biobanked PBMCs comapred to freshly sorted material. To provide insight into a disease process, naive B cells were isolated from healthy donors or a biorepository of Systemic Lupus Erythematosus undergoing disease flares.  These data identify a unique signature of accessible chromatin in SLE B cells.

overall_design: ATAC-Seq on fresh and frozen B cells

first: Christopher

middle: D

first: Jeremy

last: Boss


title: Acquired tissue-specific promoter bivalency is a basis for PRC2 necessity in adult somatic cells

first: Ramesh

last: Shivdasani

address: 450 Brookline Ave. Dana 720D

institute: Dana Farber Cancer Institute

summary:
To define H3K27me3 modified genes in intestinal stem, progenitor and epithelial cells, we performed chromatin immunoprecipitation coupled with high-throughput sequencing (ChIP-Seq).
We used RNA-sequencing to profile gene expression changes during intestinal stem cell differentiation into mature villus cells, as well as genes perturbed upon loss of PRC2 activity (deletion of Eed) . We find thousands of genes that change in expression including many H3K27me3 marked genes. The deregulated genes reaveal a intestinal tissue specific role of PRC2.

overall_design: H3K27me3, H3K4me2 and RNA Pol2 ChIP-Seq analysis of isolated mouse intestinal stem cells, enterocyte and secretory progenitor cells, and mature villus cells. RNA seq analysis of control mouse villus cells, control intestinal stem cells and Eed-deleted villus.

first: Unmesh

last: Jadhav

first: Kodandaramireddy

last: Nalapareddy

first: Madhurima

last: Saxena

first: Nicholas

last: O’Neill

first: Luca

last: Piniello

first: Guo-Cheng

last: Yuan

first: Stuart

last: Orkin


title: DDX3X regulation of global translation is impaired by medulloblastoma-associated mutations [RiboSeq]

city: Memphis

first: Yong-Dong

last: Wang

address: 262 Danny Thomas Pl, MS1135

institute: St. Jude Children's Research Hospital

summary: Whole-genome sequencing recently identified recurrent missense mutations in the RNA helicase DDX3X in pediatric medulloblastoma (MB) and other tumors. The normal function of DDX3X is poorly understood, and the consequences of its cancer-associated mutations have not been explored. Here we used genomic, biochemical, cell biological, and animal modeling approaches to investigate normal DDX3X function and the impact of cancer-associated DDX3X mutations. Cross-linking immunoprecipitation–high-throughput sequencing (CLIPseq) analyses revealed that DDX3X binds primarily to ~1000 mature mRNA targets at binding sites spanning the full mRNA length; their enrichment in the coding regions suggests that DDX3X plays a role in translational elongation. The association of wild-type DDX3X with polysomes is consistent with this observation. Cancer-associated mutations result in loss of DDX3X from polysomes and accumulation of mutant DDX3X in stress granules (cytoplasmic accumulations of translationally arrested mRNAs). Mutation-dependent redistribution of DDX3X to stress granules is also observed in a Drosophila model system and in MB tumor cells from patients carrying DDX3X mutations. Importantly, mRNAs targeted by DDX3X are enriched in translation factors, suggesting that DDX3X regulates translation both directly and indirectly. Indeed, depletion of DDX3X by RNAi or over-expression of mutant DDX3X significantly impairs global protein synthesis. Ribosome profiling confirmed this observation and showed a 5’ bias in ribosomal occupancy, further confirming the role of DDX3X in translational elongation. Together, our data show that DDX3X is a key regulator of translation and that this function is impaired by cancer-associated mutations. Finally, we found that medulloblastoma-related mutant DDX3X can efficiently bind the wild-type form suggesting that mutant DDX3X could exert a dominant negative effect in vivo.

overall_design: Examination of translation at whole genome scale under conditions of overexpression of WT DDX3X or cancer-associated DDX3X mutant G325E

first: Yasmine

last: Valentin-Vega

first: Yong

first: Matthew

last: Parker

first: Michael

last: Rusch

first: Demelza

last: Smeeth

first: David

last: Finkelstein

first: Deanna

last: Patmore

first: Nam

last: Kim

first: Anderson

last: Kanagaraj

first: Jennifer

last: Moore

first: Brett

last: Winborn

first: Jinghui

last: Zhang

first: Richard

middle: J

last: Gilbertson

first: J

last: Taylor


title: DDX3X regulation of global translation is impaired by medulloblastoma-associated mutations


title: Transcriptome of ESCs, EpiSCs and reverted ES by MM401 (Mll1 inhibitor) #2

first: Yali

last: Dou

summary: This study describes the transcriptome profiling of: 1) mouse ES cells in LIF/KSR medium; 2)EpiSCs in bFGF/serum-free (KSR) medium; 3) EpiSCs treated with MM401/LIF KSR  at D3 and D6 (P2); 4) rES reverted form EpiSC by MM401/LIF KSR treatment at P6, P30 with or without MM401 .

overall_design: RNA-Seq profiling on mouse pluripotent cells. Biological duplicates of each sample are labled as rep1/2.


title: An extensive program of periodic alternative splicing linked to cell cycle progression

status: Public on May 19 2016

city: Shanghai

first: Zefeng

address: 320 Yueyang Road

institute: CAS-MPG Partner Institute for Computational Biology (PICB)

summary: Progression through the mitotic cell cycle requires periodic regulation of gene function at the levels of transcription, translation, protein-protein interactions, post-translational modification and degradation. However, the role of alternative splicing (AS) in the temporal control of cell cycle is not well understood. By sequencing the human transcriptome through two continuous cell cycles, we identify ~1,300 genes with cell cycle-dependent AS changes. These genes are significantly enriched in functions linked to cell cycle control, yet they do not significantly overlap genes subject to periodic changes in steady-state transcript levels. Many of the periodically spliced genes are controlled by the SR protein kinase CLK1, whose level undergoes cell cycle-dependent fluctuations via an auto-inhibitory circuit. Disruption of CLK1 causes pleiotropic cell cycle defects and loss of proliferation, whereas CLK1 over-expression is associated with various cancers. These results thus reveal a large program of CLK1-regulated periodic AS intimately associated with cell cycle control.

first: Daniel

last: Dominguez

first: Benjamin

last: Blencowe


title: Transcriptome analysis of human cumulus cells reveals hypoxia as the main determinant of follicular senescence.

city: New Haven

first: Pasquale

last: Patrizio

address: 150 Sargent drive

department: Ob/Gyn

institute: Yale University

summary: RNA sequencing profiling of cumulus cells revealed that the follicles of older women have a compromised environment with hypoxic stress being the main contributor to age-related oocyte quality decay.

overall_design: Twenty cumulus cells (CC) samples (from a total of 15 patients) were obtained from oocytes of either male factor or egg donor patients. RNA sequencing and bioinformatic tools were used to identify differentially expressed genes (DEGs) between CCs from seven aged and eight young patients (<35 (years old) y.o. vs >40 y.o.). Quantitative-PCR and immunoflourescent staining were used for validation.

first: Emanuela

last: Molinari

first: Haim

last: Bar

first: Anna Marie

last: Pyle


title: De novo transcriptome profiling of cold-stressed siliques during pod filling stages in Indian mustard (Brassica juncea L.)

city: Delhi

first: Manu

last: Agarwal

address: Chatra Marg, North campus

department: Department of Botany

institute: University of Delhi

summary: Low temperature is a major abiotic stress that impedes plant growth and development. Brassica juncea is an economically important oil seed crop and is sensitive to freezing stress during pod filling subsequently leading to abortion of seeds. To understand the cold stress mediated global perturbations in gene expression, whole transcriptome of B. juncea siliques that were exposed to sub-optimal temperature was sequenced. Manually self-pollinated siliques at different stages of development were subjected to either short (6 h) or long (12 h) durations of chilling stress followed by construction of RNA-seq libraries and deep sequencing using Illumina’s NGS platform. De-novo assembly of B. juncea transcriptome resulted in 133641 transcripts, whose combined length was 117 Mb and N50 value was 1428 bp. We identified 13342 differentially regulated transcripts by pair-wise comparison of 18 transcriptome libraries. Hierarchical clustering along with Spearman correlation analysis identified that the differentially expressed genes segregated in two major clusters representing early (5-15 DAP) and late stages (20-30 DAP) of silique development. Further analysis led to the discovery of sub-clusters having similar patterns of gene expression. Two of the sub-clusters (one each from the early and late stages) comprised of genes that were inducible by both the durations of cold stress. Comparison of transcripts from these clusters led to identification of 283 transcripts that were commonly induced by cold stress, and were referred to as ‘core cold-inducible’ transcripts. Additionally, we found that 689 and 100 transcripts were specifically up regulated by cold stress in early and late stages, respectively. We further explored the expression patterns of gene families encoding for transcription factors (TFs), transcription regulators (TRs) and kinases, and found that cold stress induced protein kinases only during early silique development. We validated the digital gene expression profiles of selected transcripts by qPCR and found a high degree of concordance between the two analyses. To our knowledge this is the first report of transcriptome sequencing of cold-stressed B. juncea siliques. The data generated in this study would be a valuable resource for not only understanding the cold stress signaling pathway but also for introducing cold hardiness in B. juncea.

overall_design: mRNA-seq profilling of cold stressed Brassica juncea silique, Various development stages ( 5, 10, 15, 20, 25 and 30 DAP) of siliques were taken, 18 samples generated from two different durations of stress (6h, 12h) with their respective control (C).

first: Somya

last: Sinha

first: Vivek

last: Raxwal


title: Impact of transposable elements in Pleurotus ostreatus transcriptome profiles

city: Pamplona

first: Raul

last: Castanera

address: Campus Arrosadia, DPTO Producción Agraria

institute: Public University of Navarre

summary: Pleurotus ostreatus, also known as the oyster mushroom, is an active lignin decomposer in the forests. The genomes of the monokaryotic strains PC15 and PC9 have been used to characterize the content and distribution of transposable elements. This study analyzes the impact of transposable element insertions on the global transcriptome of P. ostreatus. The transcriptome of PC15 and PC9 has been analyzed in exponential growth during submerged fermentation in malt-yeast extract-sucrose medium

overall_design: RNAseq of two P. ostreatus strains: PC15 and PC9

first: Lucía

last: Ramírez

first: Raúl


title: Enhancer chromatin marks in Dnmt3aKO/FLT3-ITD lymphoid leukemia

city: Houston

first: Margaret

last: Goodell

state: TX

address: One Baylor Plaza, N1030

institute: Baylor College of Medicine

summary: We mapped H3K4me1 and H3K27ac chromatin marks in Dnmt3aKO/FLT3-ITD and FLT3-ITD lymphoid leukemias

overall_design: Maps of enhancer chromatin marks H3K4me1 and H3K27ac in Dnmt3aKO/FLT3-ITD leukemias and FLT3-ITD leukemias were obtained by sequencing on Illumina HiSeq 2500

last: Rodriguez

first: Wei

last: Li


title: The mir-143-145 cluster plays a pro-tumorigenic role in lung adenocarcinoma by promoting neoangiogenesis

city: Cambridge

first: A

last: Bhutkar

address: 77 Massachusetts Avenue

institute: MIT

summary: A growing body of literature has proposed cell-autonomous tumor suppressor functions for the mir-143~145 cluster in a variety of human cancers, including lung adenocarcinoma, and has reported therapeutic benefits of delivering mir-143 and mir- 145 to tumors. In contrast to these studies, we found that depletion or forced expression of mir-143 and mir-145 in an autochthonous mouse model of lung adenocarcinoma did not affect tumor development. Surprisingly, we observed that loss of mir-143~145 from the tumor microenvironment significantly reduced tumor burden, indicating a non-cell- autonomous role for these miRNAs in promoting tumorigenesis. By examining the expression patterns of different cell populations isolated in vivo from tumor-bearing lungs using an integrated computational approach, we identified a role for mir-145 in stimulating the proliferation of endothelial cells by downregulating an inhibitory kinase, Camk1d, which prevents mitotic entry. As a consequence, tumors in mir-143~145- deficient animals exhibited diminished hallmarks of neo-angiogenesis, increased apoptosis and their expansion appeared limited by the tumor’s ability to co-opt the alveolar vasculature. These findings show that expression of the mir-143~145 cluster in the tumor stroma promotes rather than suppresses tumorigenesis and cautions against the use of these miRNAs as agents in cancer therapeutics.

overall_design: Epcam-positive, CD31-positive, and triple-negative (Epcam-CD31-CD45-) cell populations isolated by flow cytometry from tumor-bearing lungs of K-rasG12D/+, miR-143/145-proficient and -deficient mice. Three independent mice from each genotype were used as biological replicates.

first: Nadya

last: Dimitrova

first: Vasilena

last: Gocheva

first: Arjun

first: Rebecca

last: Resnick

first: Robyn

last: Jong

first: Kathryn

last: Miller

first: Jordan

last: Bendor

first: Tyler

last: Jacks


title: WGBS assessment of global methylation alterations in Dnmt3aKO/FLT3-ITD lymphoid leukemia

summary: We generated global DNA methylation maps of FLT3-ITD and Dnmt3aKO/FLT3-ITD lymphoid leukemias, as well as normal control thymocytes.

overall_design: Whole genome bisulfite sequencing of cells using Illumina HiSeq 2000 or 2500

first: Benjamin


title: DDX3X regulation of global translation is impaired by medulloblastoma-associated mutations [RNA-Seq]

first: Yong-Dong

last: Wang

overall_design: Examination of the whole tarnscriptome under conditions of DDX3X knockdown or overexpression of WT DDX3X or cancer-associated DDX3X mutants

first: Yasmine

last: Valentin-Vega

first: Yong

first: Matthew

last: Parker

first: Michael

last: Rusch

first: Demelza

last: Smeeth

first: David

last: Finkelstein

first: Deanna

last: Patmore

first: Nam

middle: C

last: Kim

first: Anderson

last: Kanagaraj

first: Jennifer

last: Moore

first: Brett

last: Winborn

first: Jinghui

last: Zhang

first: Richard

last: Gilbertson

first: J

last: Taylor


title: Identification of CAMTA1 target genes in the adult mouse hippocampus

city: Heidelberg

first: Carlos

last: Bas Orth

address: Im Neuenheimer Feld 307

department: Anatomy and Cell Biology

institute: Ruprecht-Karls-University Heidelberg

summary: The formation of long-term memory requires signaling from the synapse to the nucleus to mediate neuronal activity-dependent gene transcription. Synapse-to-nucleus communication is initiated by influx of calcium ions through synaptic NMDA receptors and/or L-type voltage-gated calcium channels and involves the activation of transcription factors by calcium/calmodulin signaling in the nucleus. Recent studies have drawn attention to a new family of transcriptional regulators, the so-called calmodulin-binding transcription activator (CAMTA) proteins. CAMTAs are expressed at particularly high levels in the mouse and human brain, and we reasoned that, as calmodulin-binding transcription factors, CAMTAs may regulate the formation of long-term memory by coupling synaptic activity and calcium/calmodulin signaling to memory-related transcriptional responses. This hypothesis is supported by genetic studies that reported a correlation between CAMTA gene polymorphisms or mutations and cognitive capability in humans. Here, we show that acute knock-down of CAMTA1, but not CAMTA2, in the hippocampus of adult mice results in impaired performance in two memory tests, contextual fear conditioning and object-place recognition test. Short-term memory and neuronal morphology were not affected by CAMTA knock-down. Gene expression profiling in the hippocampus of control and CAMTA knock-down mice revealed a number of putative CAMTA1 target genes related to synaptic transmission and neuronal excitability. Patch clamp recordings in organotypic hippocampal slice cultures provided further evidence for CAMTA1-dependent changes in electrophysiological properties. In summary, our study provides experimental evidence that confirms previous human genetic studies and establishes CAMTA1 as a regulator of long-term memory formation.

overall_design: We compared the mRNA expression profile of three groups, i.e. mice infected with a recombinant adeno-associated virus (rAAV) expressing a non-targeting control shRNA, mice infected with a rAAV expressing Camta1 targeting shRNA sequence A, and mice infected with a rAAV expressing Camta1 targeting shRNA sequence B. Three animals were used per group.

last: Bas-Orth

first: Yan-Wei

last: Tan

first: Ana

last: Oliveira

first: Colin

last: Bengtson

first: Hilmar

last: Bading


title: Trancription factor Fli1 occupancy in Dnmt3aKO/FLT3-ITD lymphoid leukemia

first: Margaret

last: Goodell

summary: We mapped binding of FLI1 in Dnmt3aKO/FLT3-ITD leukemias over-expressing Dnmt3a or GFP.

overall_design: Maps of transcription factor FLI1 binding sites in tertiary transplanted Dnmt3aKO/FLT3-ITD leukemias over-expressing Dnmt3a or GFP were obtained by sequencing on Illumna HiSeq 2500

first: Benjamin

last: Rodriguez

first: Wei

last: Li


title: A role for HOX13 proteins in the regulatory switch between TADs at the HoxD locus

city: Yokohama

first: Nayuta

last: Yakushiji-Kaminatsui

state: Kanagawa

address: 1-7-22 Suehiro-cho, Tsurumi-ku

country: Japan

institute: RIKEN Center for Integrative Medical Sciences

summary: During vertebrate limb development, Hoxd genes are regulated following a bimodal strategy involving two topologically associating domains (TADs) located on either side of the gene cluster. These regulatory landscapes alternatively control different subsets of Hoxd targets, first into the arm and, subsequently, into the digits. We studied the transition between these two global regulations, a switch that correlates with the positioning of the wrist, which articulates these two main limb segments. We show that the HOX13 proteins themselves help switch off the telomeric TAD, likely through a global repressive mechanism. At the same time, they directly interact with distal enhancers to sustain the activity of the centromeric TAD, thus explaining both the sequential and exclusive operating processes of these two regulatory domains. We propose a model whereby the activation of Hox13 gene expression in distal limb cells both interrupts the proximal Hox gene regulation and re-enforces the distal regulation. In the absence of HOX13 proteins, a proximal limb structure grows without any sign of wrist articulation, likely related to an ancestral fish-like condition.

overall_design: 4C-seq analysis of Hoxd11 and Hoxd13 3D interactions in proximal and distal limbs from E12.5 wt or Hoxa13-/-;Hoxd13-/- mutant embryos

first: L

last: Beccari

last: Woltering

first: E

last: Rodriguez-Carballo

first: D

last: Duboule


title: Genome-wide analysis of Alternaria stress-responsive transcriptome in tomato

first: Pallob

last: Kundu

summary: Gene expression cascade in a plant is altered in times of stress. Reprogramming of the expression profiles of genes is required for a robust and specific response. Until now, tomato  transcriptomic alteration in  response to Alternaria fungal stress were not known. This study presents the profile of genes that are differentially expressed during Alternaria stress in local tomato cultivar (Pusa Ruby). At least 2944 genes expression was varied and pathways that are altered during this compatible interaction have been identified.  Supported by DBT, Govt. of India.

overall_design: RNA seq profile of Indian tomato (Solanum lycopersicum) cultivar Pusa Ruby infected with Alternaria. One mock infected and One Alternaria fungus infected samples were analyzed.

first: Deepti

last: Sarkar

first: Ranjan

last: Maji

first: Zhumur

last: Ghosh


title: DNMT3A and TET2 Compete and Cooperate to Repress Differentiation Lineage-Specific Factors in Hematopoietic Stem Cells

city: HOUSTON

first: Jianzhong

last: Su

state: TEXAS

address: 1 Baylor Plaza

institute: Baylor college of Medicine

summary: Mutations in the epigenetic modifiers DNMT3A and TET2 non-randomly co-occur in lymphoma and leukemia despite their epistasis in the methylation-hydroxymethylation pathway. Using double knock-out (DKO) mice in which malignancy development is accelerated, we show that the DKO methylome reflects regions of independent, competitive and cooperative activity. Expression of lineage-specific transcription factors, including the erythroid regulator Klf1 is upregulated in DKO HSCs. DNMT3A and TET2 both repress Klf1 suggesting a model of cooperative inhibition by the epigenetic modifiers. These data demonstrate a dual role for TET2 in promoting and inhibiting HSC differentiation, loss of which, along with DNMT3A, obstructs differentiation leading to transformation.

overall_design: I)Whole genome bisulfite sequencing of Tet2 conditional knockout hematopoietic stem cells,  Tet2 and DNMT3a double knockout hematopoietic stem cells using Illumina HiSeq 2000.II)Whole genome CMS-enriched bisulfite sequencing  of Tet2 conditional knockout hematopoietic stem cells,  Tet2 and DNMT3a double knockout hematopoietic stem cells using Illumina HiSeq 2000.III)Whole genome mRNA profiles of Tet2 conditional knockout hematopoietic stem cells,  Tet2 and DNMT3a double knockout hematopoietic stem cells using Illumina HiSeq 2000. IV)Genome wide histone modification H3K27me3 profiling of Tet2 conditional knockout hematopoietic stem cells,  Tet2 and DNMT3a double knockout hematopoietic stem cells using Illumina HiSeq 2000

first: Xiaotian

last: Zhang

first: Hyun

last: Park


title: CIS is a potent checkpoint in NK cell-mediated tumour immunity

city: Clayton

last: Shi

state: Victoria

address: Wellington Rd

country: Australia

institute: Monash University

summary: Natural killer (NK) cells have evolved to detect and kill aberrant cells with this activity being governed by the cytokine interleukin (IL)-15 and foreign and self-ligands. We have identified CIS (Cytokine-inducible SH2-containing protein; Cish gene) as the critical negative regulator of IL-15 signalling in NK cells. Cish was rapidly induced in response to IL-15 and deletion of Cish rendered NK cells hypersensitive to IL-15, as evidenced by superior proliferation, survival, IFN-γ production and cytotoxicity towards tumours. This was associated with enhanced JAK/STAT signalling in Cish-deleted NK cells. Correspondingly, CIS interacted with the tyrosine kinase JAK1, inhibiting its enzymatic activity and targeting JAK for proteasomal degradation. Cish-/- mice were resistant to melanoma, prostate and breast cancer metastasis in vivo, and this was intrinsic to NK cell activity. This study has uncovered a potent checkpoint in NK cell-mediated tumour immunity and holds promise for novel immunotherapies directed at blocking CIS function.

overall_design: Transcriptional profiling of ex vivo and in vitro CIS-/- nature killer cells

first: Rebecca

middle: B

last: Delconte

first: Tatiana

last: Kolesnik

first: Laura

middle: F

last: Dagley

first: Sandra

last: Nicholson

first: Nicholas

last: Huntington


title: RecG Directs DNA Synthesis During Double-Strand Break Repair

city: Edinburgh

first: Benura

last: Azeroglu

address: Alexander Crum Brown Road

institute: University of Edinburgh

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

overall_design: Refer to individual Series


title: RecG Directs DNA Synthesis During Double-Strand Break Repair [ChIP-seq]

summary: In the bacterium Escherichia coli, RecG directs DNA synthesis during the repair of DNA double-strand breaks by homologous recombination.

overall_design: Examination of RecA binding during double-strand break repair in Escherichia coli in the presence and absence of RecG protein

first: Julia

last: Mawer

first: Martin

last: White

first: Charlotte

last: Cockram

first: A

last: Hasan

first: Milana

last: Filatenkova

first: David

last: Leach


title: RecG Directs DNA Synthesis During Double-Strand Break Repair [Chromosomal marker frequency analysis]

overall_design: Chromosomal marker frequency analysis (MFA) following induction of a DSB in the absence and presence of RecG


title: RNA deep sequencing to compare genome-wide differences between PRMT5/knockdown and control AML cells

city: Duarte

first: Xiwei

last: Wu

state: CA

address: 1500 E Duarte Rd

department: Molecular and Cellular Biology

institute: Beckman Research Institute

summary: We report the pro-leukemic activities of PRMT5 in AML partly via modulating the transcription of key genes. The goal of this experiment is to confirm the changes observed in expression of genes targeted by PRMT5 activities. RNA Deep sequencing was employed to validate and reproduce the changes measured by quantitative reverse transcription polymerase chain reaction (qRT–PCR) techniques.

overall_design: Total RNA samples from MV4-11 cell line (FLT3-ITD AML) following PRMT5 knockdown using specific short hairpin RNA (shRNA) was used to compare gene expression pattern between PRMT5/Knockdown and control (control: MV4-11 cells transduced with scramble control).

first: Somayeh

last: Tarighat

first: Ling

last: Li

first: Guido

last: Marcucci


title: DDX3X regulation of global translation is impaired by medulloblastoma-associated mutations [CLIP-Seq]

first: Yong-Dong

last: Wang

summary: Whole-genome sequencing recently identified recurrent missense mutations in the RNA helicase DDX3X in pediatric medulloblastoma (MB) and other tumors. The normal function of DDX3X is poorly understood, and the consequences of its cancer-associated mutations have not been explored. Here we used genomic, biochemical, cell biological, and animal modeling approaches to investigate normal DDX3X function and the impact of cancer-associated DDX3X mutations. Cross-linking immunoprecipitation–high-throughput sequencing (CLIPseq) analyses revealed that DDX3X binds primarily to ~1000 mature mRNA targets at binding sites spanning the full mRNA length; their enrichment in the coding regions suggests that DDX3X plays a role in translational elongation. The association of wild-type DDX3X with polysomes is consistent with this observation. Cancer-associated mutations result in loss of DDX3X from polysomes and accumulation of mutant DDX3X in stress granules (cytoplasmic accumulations of translationally arrested mRNAs). Mutation-dependent redistribution of DDX3X to stress granules is also observed in a Drosophila model system and in MB tumor cells from patients carrying DDX3X mutations. Importantly, mRNAs targeted by DDX3X are enriched in translation factors, suggesting that DDX3X regulates translation both directly and indirectly. Indeed, depletion of DDX3X by RNAi or over-expression of mutant DDX3X significantly impairs global protein synthesis. Ribosome profiling confirmed this observation and showed a 5’ bias in ribosomal occupancy, further confirming the role of DDX3X in translational elongation. Together, our data show that DDX3X is a key regulator of translation and that this function is impaired by cancer-associated mutations. Finally, we found that medulloblastoma-related mutant DDX3X can efficiently bind the wild-type form suggesting that mutant DDX3X could exert a dominant negative effect in vivo.

overall_design: Examination of RNA targets od endogenous DDX3X

first: Yasmine

last: Valentin-Vega

first: Yong

first: Matthew

last: Parker

first: Michael

last: Rusch

first: Demelza

last: Smeeth

last: Finkelstein

first: Deanna

last: Patmore

first: Nam

last: Kim

first: Anderson

last: Kanagaraj

first: Jennifer

last: Moore

first: Brett

last: Winborn

first: Jinghui

last: Zhang

first: Richard

last: Gilbertson

first: J

last: Taylor


title: Covalent targeting of remote cysteine residues to develop CDK12 and CDK13 inhibitors [ChIP-Seq]

last: Young

address: 9 Cambridge Center

institute: Whitehead Institute for Biomedical Research

summary: Cyclin-dependent kinases 12 and 13 (CDK12 and CDK13) play critical roles in the regulation of gene transcription. However, the absence of CDK12 and CDK13 inhibitors has hindered the ability to investigate the consequences of their inhibition in healthy cells and cancer cells. Here we describe the rational design of a first-in-class CDK12 and CDK13 covalent inhibitor, THZ531. Co-crystallization of THZ531 with CDK12–cyclin K indicates that THZ531 irreversibly targets a cysteine located outside the kinase domain. THZ531 causes a loss of gene expression with concurrent loss of elongating and hyperphosphorylated RNA polymerase II. In particular, THZ531 substantially decreases the expression of DNA damage response genes and key super-enhancer-associated transcription factor genes. Coincident with transcriptional perturbation, THZ531 dramatically induced apoptotic cell death. Small molecules capable of specifically targeting CDK12 and CDK13 may thus help identify cancer subtypes that are particularly dependent on their kinase activities.

overall_design: ChIP-Seq for CDK12 and RNA Pol II during treatment with a transcriptional inhibitor

first: Tinghu

first: Nicholas

last: Kwiatkowski

first: Calla

last: Olson

first: Sarah

last: Dixon Clarke

first: Brian

last: Abraham

first: Ann

last: Greifenberg

first: Scott

last: Ficarro

first: Jonathan

last: Elkins

first: Yanke

last: Liang

first: Nancy

last: Hannett

first: Theresa

last: Manz

first: Bartolomiej

last: Bartowiak

first: Arno

last: Greenleaf

first: Jarrod

last: Marto

first: Matthais

last: Geyer

first: Alex

middle: N

last: Bullock

first: Nathanael

last: Gray

first: Mingfeng

last: Hao


title: HiC of Wild Type Neurospora crassa and mutants deficient in heterochromatin formation

city: Colorado Springs

first: Andrew

middle: David

last: Klocko

state: Colorado

address: 278 Centennial Hall, 1420 Austin Bluffs Pkwy

department: Chemistry and Biochemistry

institute: University of Colorado Colorado Springs

summary: Eukaryotic genomes are organized into chromatin domains with distinct three-dimensional arrangements resulting from nucleic acid and protein factor interactions within the physical constraints of the nucleus. It is of obvious interest to determine interactions between various chromosomal regions defined by these nuclear constraints, and to identify important factors that limit the interactions. We used chromosome conformation capture (3C) followed by high-throughput sequencing (HiC) to improve our understanding of Neurospora crassa genome organization and to examine if known components of heterochromatin machinery influence nuclear organization. In heterochromatin establishment, DIM-5 tri-methylates histone H3 on lysine 9 (H3K9me3), and this mark is subsequently bound by Heterochromatin Protein-1 (HP1). NUP-6 is required to target DIM-5 to incipient A:T rich DNA and the dim-3 mutant of NUP-6 is deficient in DIM-5 localization. We performed HiC on chromatin from wildtype, Δdim-5, Δhpo, and dim-3 strains. The genome configuration of wild type nuclei revealed strong intra- and inter-chromosomal associations between both constitutive and facultative heterochromatic domains, with the strongest interactions among the centromeres, telomeres and interspersed heterochromatin. We found that loss of the H3K9me3 mark, as well as loss of HP1, minimally altered the chromatin organization found at these strongly interacting heterochromatic regions. Surprisingly, dim-3 chromatin was highly disorganized suggesting NUP-6 plays key role(s) in genome structure. Thus, our datasets suggest that while heterochromatic regions are critical to defining the genome conformation in Neurospora, non-canonical protein factors may play a key role in maintaining this organization.

overall_design:
We analyzed a total of four strains of Neurospora crassa by chromatin conformation capture followed by high throughput sequencing (HiC).  A wild type strain, NMF39, serves as the reference for the mutants deficient in heterochromatin formation (delta dim-5; delta hpo; and dim-3). Each strain has at least two replicates, and the NMF39 wild type strain has three replicates. Strains were grown from conidia ~4 hours, crosslinked, and conidia were made into spheroplasts. Genomic DNA from spheroplasts was digested with HindIII, ends were filled in with dNTPs (including biotin-conjugated dCTP), blunt ends were ligated, crosslinks were reversed, DNA was soniciated, ligation products were purified with streptavidin beads, and libraries were prepared for sequencing.

Supplementary file 'GSE71024_H3K9me3_10kb-bins_wg_forCircos.txt' includes processed data obtained by using the raw data from Sample GSM1686146 of Series GSE68897, and supplementary file 'GSE71024_H3K27me3_10kb-bins_wg_forCircos.txt' includes processed data obtained by using the raw data from Sample GSM1686139 of Series GSE68897.

last: Galazka

first: Eric

middle: U

last: Selker

first: Michael

last: Freitag


title: Transcriptome of Sclerotinia sclerotiorum during vegetative growth, sclerotial development, myceliogenic germination, carpogenic germination, apothecium formation (stipe) and infection

city: Wuhan

first: Xueliang

last: Lyu

state: Hubei

address: Shizishan street NO.1

department: College of Plant Science and Technology

institute: Huazhong Agricultural University

summary: The Solexa/Illumina’s digital gene expression (DGE) system was used to gain insight into the broad range of transcriptional responses during the vegetative growth, sclerotial development, myceliogenic germination, carpogenic germination, apothecium formation (stipe) and infection of Sclerotinia sclerotiorum. We obtained a sequencing depth of approximately 3.3 million clean tags per cDNA library. Tag mapping indicated that these six cDNA libraries in total represented more than 66.7% of all of the genes presented in the predicted transcript databases of the Broad Institute. Thouthands of differentially expressed genes were indentified during the various developmental stages compared to the vegetative growth stage. Our results could increase and deepen the understanding of the vegetative and reproductive development as well as the infection of S. sclerotiorum

overall_design: The S. sclerotiorum strain Ep-1PNA367 was grown or treated under different conditions, and samples were collected for RNA extraction for DGE analysis during the following stages: (i) Vegetative stage: activating hyphal agar discs of the Ep-1PNA367 stain were placed on a cellophane membrane overlaid onto PDA medium at 20°C; subsequently, the mycelia were collected at 12, 24, 36, 48 and 60 h; (ii) Sclerotial formation stage: the colonies growing on the cellophane membrane overlaid onto PDA medium were further incubated under the same conditions and then the cultures were collected at 84, 96, 108, 120 and 132 h; (iii) Early stages of infection: fresh hyphal fragments of the Ep-1PNA367 strain were overlaid onto sterilized cheese cloth, which was overlaid onto the leaves of the A. thaliana ecotype Columbia-0, followed by inoculation at 20°C with 100% relative humility; the cheese cloth with hyphae was then rolled up from the leaves at 9 h and 12 h; (iv) Sclerotial myceliogenic germination stage: sclerotia were surface sterilized with sodium hypochlorite and were sowed on a PDA plate at 20°C to induce myceliogenic germination, and when approximately 50% of sclerotia had germinated, the sclerotia were harvested; (v) Sclerotial carpogenic germination stage: sclerotia were dried at room temperature and were pretreated in a freezer (4-6°C) for up to one month and then were surface sterilized and sowed on wet sterilized sands in a plate at 15°C to induce carpogenic germination; when approximately 50% of the sclerotia germinated (stipes having only emerged from sclerotia), the sclerotia were harvested; and (vi) Early apothecial formation stage: sclerotia were allowed to grow in the same incubator, and the stipes were cut and collected immediately before apothecium formation for RNA extraction. Finally, different time-point samples from the vegetative stage, sclerotial formation stage and infection stage were pooled together respectively according to equal quantities for RNA extraction.

first: Jiasen

last: Chen

first: Jiatao

last: Xie

first: Yanping

last: Fu

first: Daohong

last: Jiang


title: Tracing the formation of hematopoietic stem cells in mouse embryos by single-cell functional and RNA-Seq analyses [10-cell]

city: Beijing

first: Xianlong

last: Li

address: 5, Yiheyuan Rd., Haidian District

department: Biodynamic Optical Imaging Center, College of Life Sciences

institute: Peking University

summary: Hematopoietic stem cells (HSCs) in adult are specified early from the endothelium-derived precursors (e.g., hemogenic endothelium, and pre-HSCs) in mouse mid-gestation embryos, the detailed process, however, is still largely unknown due to their rareness, transience, and current inability to prospectively isolate them efficiently . Here we developed a potent set of surface markers that could capture the earliest emerging HSCs, the CD45- pre-HSCs with high accuracy and purity, as rigorously and functionally verified by single-cell-initiated serial transplantation assays. Then we applied single-cell RNA-Seq technique to analyze five populations related to HSC formation: the CD45- (type 1) and CD45+ (type 2) pre-HSCs as well as endothelial cells in the E11 AGM region; and later mature HSCs in the E12 and E14 fetal livers. Compared to other cell populations, both type 1 and type 2 pre-HSCs have their unique signatures of transcription machinery, transcription factor network, signaling pathway, cell cycle status, metabolism state, and lncRNA expression patterns. Our work paves the way for dissection of the complex molecular mechanisms regulating the step-wise formation of HSCs from endothelial cells, thus informing future efforts on engineering HSCs for clinical application.

overall_design: RNA-Seq of 35 10-cell pooled samples from 5 FACS sorted cell types, i.e., endothelial cells (ECs), T1 and T2 pre-HSCs from E11 AGM region, as well as mature HSCs from E12 and E14 fetal liver

first: Fan

last: Zhou

first: Ping

last: Zhu

first: Jie

first: Meng

last: Ding

first: Fuyin

last: Xiong

first: Xiaona

last: Zheng

first: Zhuan

first: Yanli

last: Ni

first: Lu

last: Wen

first: Bing

last: Liu

first: Fuchou

last: Tang


title: Pitx2 promotes heart repair by transcriptionally activating the antioxidant response after cardiac injury

first: Min

last: Zhang

state: Texas

address: One Baylor Plaza Room 504E

department: Department of Molecular Physiology

summary: Myocardial infarction results in compromised myocardial function with heart failure due to insufficient cardiomyocyte self-renewal. Unlike lower vertebrates, mammalian hearts only have a transient neonatal renewal capacity. Reactivating the primitive reparative ability in the fully mature heart requires an intimate knowledge of the molecular mechanisms promoting early heart repair. Here we identified a novel factor that can sufficiently promote heart muscle repair. By screening an established Hippo-deficient heart regeneration model for renewal promoting factors, we found that PITX2 protein expression in ventricles was induced after cardiomyocyte injury. Moreover, Pitx2-deficient neonatal hearts failed to repair after apex resection. Pitx2-gain-of-function in ventricular cardiomyocytes conferred reparative ability to the adult mouse heart after myocardial infarction. Integrated genomic analyses indicated that Pitx2 activated genes encoding electron transport chain components and reactive oxygen species scavengers. Pitx2 mutant myocardium had elevated reactive oxygen species levels while supplement of antioxidants suppressed the Pitx2-loss-of-function phenotype. Furthermore, PITX2 directly binds NFE2L2 and translocates from cytoplasm to nucleus upon oxidative stress.

overall_design:
RNA-Seq Experiments:
Total mRNA for RNAseq was extracted from the ventricles of Pitx2CKO and Pitx2f/f mice 5 days after P1 apex resection

ChIP-Seq Experiments:
Examination of transcription factors Pitx2 binding sites 4 days post P2 LAD-O

first: James

last: Martin

first: Ge

last: Tao

first: Peter

last: Kahr

first: Yuka

last: Morikawa

first: Lele

first: Zhao

last: Sun

first: Brad

middle: A

last: Amendt


title: Tracing the Formation of Haematopoietic Stem Cells in Mouse Embryos by Single-cell Functional and RNA-Seq Analyses [single-cell]

first: Xianlong

last: Li

summary: Haematopoietic stem cells (HSCs) are derived early from embryonic precursor cells, such as haemogenic endothelial cells and pre-HSCs. However, the identity of precursor cells remains elusive due to their rareness, transience, and inability to be isolated efficiently. Here we employed potent surface markers to capture the nascent pre-HSCs at 30% purity, as rigorously validated by single-cell-initiated serial transplantation assay. Then we applied single-cell RNA-Seq technique to analyse five populations closely related to HSC formation: endothelial cells, CD45- and CD45+ pre-HSCs in E11 aorta-gonad-mesonephros (AGM) region, and mature HSCs in E12 and E14 foetal liver. In comparison, the pre-HSCs showed unique features in transcriptional machinery, apoptosis, metabolism state, signalling pathway, transcription factor network, and lncRNA expression pattern. Among signalling pathways enriched in pre-HSCs, the mTOR activation was uncovered indispensable for the emergence of HSCs but not haematopoietic progenitors from endothelial cells in vivo. By comparing with proximal populations without HSC potential, the core molecular signature of pre-HSCs was identified. Collectively, our work paves the way for dissection of complex molecular mechanisms regulating the step-wise generation of HSCs in vivo, informing future efforts to engineer HSCs for clinical application.

overall_design:
RNA-Seq of 181 single-cell samples from 8 FACS sorted cell types:
1. endothelial cells (samples E11.0_EC_xxxx. CD31+ VE-cadherin+CD41-CD43-CD45-Ter119-);
2. T1 pre-HSCs (samples E11.0_T1_xxxx. CD31+CD45-CD41low c-Kit+CD201high);
3. T1 CD201- cells (samples E11.0_T1CD201neg_xxxx,  CD31+CD45-CD41low c-Kit+CD201low/-) ;
4. T2 pre-HSCs (samples E11.0_T2_35xx. CD31+CD45+c-Kit+CD201high),
5. T2 CD41low (samples E11.0_T2_21xx, E11.0_T2_24xx and E11.0_T2_27xx. CD31+CD45+CD41low);
6. E12 HSCs (samples E12.5_FL_xxxx. Lin-Sca-1+Mac-1lowCD201+);
7. E14 HSCs (samples E14.5_FL_xxxx. CD45+CD150+CD48-CD201+);
8. Adult HSCs (samples Adult_HSC_xxxx. CD45+CD150+CD48-CD201+).

ECs, T1 pre-HSCs, T1 CD201- cells, T2 pre-HSCs, T2 CD41low cells were sorted from E11 AGM region. Mature HSCs were sorted from E12 or E14 fetal liver and adult bonemarrow.

first: Fan

first: Ping

first: Jie


title: Tracing the formation of hematopoietic stem cells in mouse embryos by single-cell functional and RNA-Seq analyses