title: IL-10 signaling remodels adipose chromatin architecture to limit thermogenesis and energy expenditure [ATAC-Seq]

city: Los Angeles

first: Peter

last: Tontonoz

state: CA

address: 675 Charles E Young

institute: UCLA

summary: Signaling pathways that promote adipose tissue thermogenesis are well characterized, but the physiologic limiters of energy expenditure are largely unknown.  Here we show that ablation of the anti-inflammatory cytokine IL-10 improves insulin sensitivity, protects against diet-induced obesity, and elicits the browning of white adipose tissue.  Mechanistic studies define bone marrow cells as the source of the IL-10 signal and mature adipocytes as the target cell type mediating these effects.  IL-10 receptor alpha is highly enriched in mature adipocytes and is induced in response to cold, obesity and aging.  ATAC-seq and RNA-seq reveal that IL-10 represses the transcription of thermogenic genes in adipocytes by altering chromatin accessibility and inhibiting ATF and PGC-1alpha recruitment to key enhancer regions.  These findings identify the IL-10 axis as a critical and potentially targetable regulator of thermogenesis, and expand our understanding of the links between inflammatory signaling and adipose tissue function in the setting of obesity.

overall_design: Immortalized brown/beige-like preadipocyte cell line (iBAd Cells) was used for ATAC-Seq and mRNA-Seq. For RNA-Seq, triplicate experiments were performed, for ATAC-Seq individual samples were sequenced after 5 days of differentiation with either control treatment, or including IL-10 overnight prior to addition of Isoproterenol for 5-6 hours. Inguinal White adipose tissue was used for RNA-Seq from either WT or IL-10-/- animals, where 11 IL10-/- mice and 9 WT mice were seperately pooled for library construction and sequencing. Primary SVF on day 9 of differentiation or primary adipose was used for ATAC-Seq. ATAC-Seq individual samples were sequenced with either control treatment, or including IL-10 overnight prior to addition of FSK. Inguinal White adipose tissue from 10 weeks old chow-fed mice were used for isolation of primary adipose nuclei.

first: Brandon

last: Thomas

first: Prashant

last: Rajbhandari

first: An-Chieh

last: Feng


title: Single-nucleus analysis of accessible chromatin in developing mouse forebrain reveals cell-type-specific transcriptional regulation

status: Public on Jan 27 2018

city: La Jolla

first: Rongxin

last: Fang

country: USA

institute: Ludwig Institute for Cancer Research

type: Genome binding/occupancy profiling by high throughput sequencing

type: Expression profiling by high throughput sequencing

summary: Analysis of chromatin accessibility can reveal transcriptional regulatory sequences, but heterogeneity of primary tissues poses a significant challenge in mapping the precise chromatin landscape in specific cell types. Here we report single-nucleus ATAC-seq, a combinatorial barcoding-assisted single-cell assay for transposase-accessible chromatin that is optimized for use on flash-frozen primary tissue samples. We apply this technique on the mouse forebrain through eight developmental stages. Through analysis of more than 15,000 nuclei, we identify 20 distinct cell populations corresponding to major neuronal and non-neuronal cell types. We further define cell-type-specific transcriptional regulatory sequences, infer potential master transcriptional regulators and delineate developmental changes in forebrain cellular composition. Our results provide insight into the molecular and cellular dynamics that underlie forebrain development in the mouse and establish technical and analytical frameworks that are broadly applicable to other heterogeneous tissues.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: We performed combinatorial single-nucleus ATAC-seq for 7 stages of embryonic forebrain development from E11.5 to P0 and adult forebrain. Data analysis pipeline used in this study: https://github.com/r3fang/snATAC


title: Enhancer invasion shapes MYCN dependent transcriptional amplification in neuroblastoma

status: Public on Jan 29 2018

city: Boston

first: James

last: Bradner

state: MA

address: 450 Brookline

department: Medical Oncology

institute: Dana-Farber Cancer Institute

summary: Amplification of the locus encoding the oncogenic transcription factor MYCN is a defining feature of high-risk neuroblastoma. Here we present the first dynamic chromatin and transcriptional landscape of MYCN perturbation in neuroblastoma. At oncogenic levels, MYCN associates with E-box binding motifs in an affinity-dependent manner, binding to strong canonical E-boxes at promoters and invading abundant weaker non-canonical E-boxes clustered at enhancers. Loss of MYCN leads to a global reduction in transcription, which is most pronounced at MYCN target genes with the greatest enhancer occupancy. These highly occupied MYCN target genes show tissue-specific expression and are linked to poor patient survival. The activity of genes with MYCN-occupied enhancers is dependent on the tissue-specific transcription factor TWIST1, which co-occupies enhancers with MYCN and is required for MYCN-dependent proliferation. These data implicate tissue-specific enhancers in defining often highly tumor-specific ‘MYC target gene signatures’ and identify disruption of the MYCN enhancer regulatory axis as a promising therapeutic strategy in neuroblastoma.

overall_design: Refer to individual Series


title: ZBTB2 as a novel reader for unmethylated DNA [RNA-seq]

city: Zürich

first: Ino

last: Karemaker

address: Winterthurerstrasse 190

country: Switzerland

institute: University of Zurich

summary: DNA methylation is a dynamic epigenetic modification that plays a key role in various cellular processes. Proteins that bind to DNA depending on its methylation status are thought to play an important role in DNA methylation-mediated gene expression. Using a variety of genomics and proteomics approaches, we identified ZBTB2 as a novel reader of unmethylated DNA. ZBTB2, which forms a complex with ZBTB25 and ZNF639, preferentially binds at CpG island promoters in mouse embryonic stem cells, from where it regulates genes that are involved in the exit from pluripotency. Binding of ZBTB2 to target genes is mostly associated with gene activation. Furthermore, ZBTB2 is intricately interwoven with DNA methylation, as we found not only that its binding to DNA is methylation-sensitive, but also that ZBTB2 regulates the turnover of methylated DNA. Summarising, we propose that ZBTB2 is a DNA methylation-sensitive transcription factor that is involved in cellular differentiation.

overall_design: RNA-seq samples of wildtype ESCs and Zbtb2 KO ESCs

middle: D


title: Gene expression in EBV-positive versus EBV-loss clones derived from endemic Burkitt lymphoma cell lines

status: Public on Jan 25 2018

city: Durham

first: Wenbin

last: Wei

address: Stockton Road

department: Biosciences

country: United Kingdom

institute: Durham University

summary: Whilst the association of Epstein-Barr virus (EBV) with Burkitt lymphoma (BL) has long been recognized, the precise role of the virus in BL pathogenesis is not fully resolved. EBV can be lost spontaneously from some BL cell lines, and these EBV-loss lymphoma cells reportedly have a survival disadvantage. We have generated an extensive panel of EBV-loss clones from multiple BL backgrounds and examined their phenotype comparing them to their isogenic EBV-positive counterparts. Whilst loss of EBV from BL cells is rare, it is consistently associated with an enhanced predisposition to undergo apoptosis and reduced tumorigenicity in vivo. We investigated whether there were common gene expression changes between EBV-positive and loss clones derived for four endemic Burkitt lyphoma cell lines that could explain the apoptosis sensitivity of clones that had lost EBV.

overall_design: 14 samples from 14 different clonal cell lines were analysed. These comprised two EBV-positive and two EBV-loss clones from each of  Mutu-BL, Akata-BL and Awia-BL and one EBV-positive and EBV -loss clone from Elijah -BL

first: Leah

last: Fitzsimmons

first: Andrew

middle: J

last: Boyce

first: Deborah

last: Croom-Carter

first: Rosemary

last: Tierney

first: Marco

last: Herold

first: Andreas

last: Strasser

first: Gemma

middle: L

last: Kelly

first: Martin

last: Rowe


title: Expression analysis of Fudulus heteroclitus populations exposed to arsenic during hypo-osmotic shock

city: Hanover

first: Thomas

middle: H.

last: Hampton

state: NH

address: Dewey Field Road, HB 7550

department: Microbiology and Immunology

institute: Geisel School of Medicine at Dartmouth

summary:
Gene expression was measured in the gills of saltwater (SW) acclimated Atlantic Killifish, Fundulus heteroclitus, over a time course of freshwater (FW) exposure, in which half the fish were exposed to arsenic.
Fish were sampled from three populations, two from Maine, USA (ME) and one from Virginia, USA (VA)

overall_design:
72 samples comparing gene expression in killifish fish gills from three populations during acclimation to fresh water: 24 from Northeast Creek, Maine (ME1), 24 from Horseshoe Creek, Maine (ME2), and 24 King’s Creek, Virginia (VA).  Half of the freshwater acclimation time course experiments included exposure to 100 micrograms per liter arsenic, as follows. For each population, groups of 4 fish were exposed to: seawater (0 hours freshwater), seawater and arsenic (0 hours freshwater), freshwater for 1 hour, freshwater for 1 h with arsenic,  freshwater for 24 hours, freshwater for 24 hours with arsenic.
Gene expression measurements were made with custom NimbleGen platform GPL14994, which has 135,000 probes that map to 34,713 gene models and 16,104 unique Fundulus heteroclitus genes based on a high-quality killifish reference genome (PMID:28201664)

first: Joseph

middle: R

last: Shaw

middle: H

first: Stephen

middle: P

last: Glaholt


title: IL-10 signaling remodels adipose chromatin architecture to limit thermogenesis and energy expenditure

status: Public on Jan 28 2018

first: Peter

last: Tontonoz

department: Pathology

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


title: Single-cell nanobiopsy reveals compartmentalization of mRNAs within neuronal cells

status: Public on Jan 24 2018

city: Santa Cruz

first: Nader

last: Pourmand

address: 1156 High Street

department: Department of Biomolecular Engineering

institute: University of California at Santa Cruz

summary: In highly polarized cells such as neurons, compartmentalization of mRNA and of local protein synthesis enables remarkably fast, precise, and local responses to external stimuli. These responses are highly important for neuron growth cone guidance, synapse formation, and regeneration following injury. Because an altered spatial distribution of mRNA can result in mental retardation or neurodegenerative diseases, subcellular transcriptome analysis of neurons could be a useful tool for studying these conditions, but current techniques, such as in situ hybridization, bulk microarray, or RNA-Seq, impose tradeoffs between spatial resolution and multiplexing. To obtain a comprehensive analysis of the cell body versus neurite transcriptome from the same neuron, we have recently developed a label-free, single-cell nanobiopsy platform based on scanning ion conductance microscopy (SICM), that uses electrowetting within a quartz nanopipette to extract cellular material from living cells with minimal disruption of the cellular membrane and milieu. In this study, we used this platform to collect samples from the cell bodies and neurites of human neurons and analyzed the mRNA pool with multiplex RNA-Seq. The minute volume of a nanobiopsy sample allowed us to extract samples from several locations in the same cell and to map the various mRNA species to specific subcellular locations. In addition to previously identified transcripts, we discovered new sets of mRNAs localizing to neurites, including nuclear genes such as Eomes and Nap1l3. In summary, our single-neuron nanobiopsy analysis provides opportunities to improve our understanding of intracellular mRNA transport and local protein composition in neuronal growth, connectivity, and function.

overall_design: Analysis of mRNA compartmentalization in cell bodies and neurites of human iPS-derived neuronal cells by nanobiopsy sampling and RNA-Sequencing.

first: Eszter

middle: N

last: Tóth

first: Akshar

last: Lohith

first: Akiyoshi

last: Fukamizu


title: Sonic hedgehog fusion-negative rhabdomyosarcoma orgiginates from endothelial progenitors

city: Memphis

first: David

last: Finkelstein

state: TN

address: 332 N. Lauderdale St.

department: Computational Biology

institute: St Jude Children's Research Hospital

summary: Genetic fate mapping was preformed on aP2-Cre;tdTomato and aP2-Cre;tdTomato;SmoM2/+ animals and endothelial progenitor cells identified as the cell of origin of FN-RMS in aP2-Cre;SmoM2/+ animals

overall_design: mononucular cells were isolated from aP2-Cre;tdTom sternocleidomastoid muscle and aP2-Cre;tdTom;SmoM2/+ tumors and discreet populations collected by FACS

first: Catherine

last: Drummond

first: Jason

middle: A

last: Hanna

first: Matthew

last: Garcia

first: Daniel

last: Devine

first: Alana

last: Heyrana

first: Jennifer

last: Peters

first: Jerold

middle: E

last: Rehg

first: Victoria

middle: C

last: Frohlich

first: Mark

last: Hatley


title: Commensal microbiota modulate gene expression in the skin

city: Philadelphia

first: Elizabeth

last: Grice

state: PA

address: 1007 BRB II/III, 421 Curie Blvd

department: Dermatology

institute: University of Pennsylvania

summary:
Background: The skin harbors complex communities of resident microorganisms, yet little is known of their physiological roles and the molecular mechanisms that mediate cutaneous host-microbe interactions. Here, we profiled skin transcriptomes of mice reared in the presence and absence of microbiota to elucidate the range of pathways and functions modulated in the skin by the microbiota.

Results: A total of 2820 genes were differentially regulated in response to microbial colonization and were enriched in gene ontology (GO) terms related to the host-immune response and epidermal differentiation. Innate immune response genes and genes involved in cytokine activity were generally upregulated in response to microbiota and included genes encoding toll-like receptors, antimicrobial peptides, the complement cascade, and genes involved in IL-1 family cytokine signaling and homing of T cells. Our results also reveal a role for the microbiota in modulating epidermal differentiation and development, with differential expression of genes in the epidermal differentiation complex (EDC). Genes with correlated co-expression patterns were enriched in binding sites for the transcription factors Klf4, AP-1, and SP-1, all implicated as regulators of epidermal differentiation. Finally, we identified transcriptional signatures of microbial regulation common to both the skin and the gastrointestinal tract.

Conclusions: With this foundational approach, we establish a critical resource for understanding the genome-wide implications of microbially mediated gene expression in the skin and emphasize prospective ways in which the microbiome contributes to skin health and disease.

overall_design: mRNA profiles of dorsal skin from 7 conventionally-raised specific pathogen free (SPF) mice and 9 germ free (GF) C57BL/6 mice generated by sequencing on the Illumina HiSeq 2000.

first: Jacquelyn

middle: S

last: Meisel


title: Pluripotent stem cell models of Blau syndrome reveal an IFN-<gamma>-dependent inflammatory response in macrophages

city: Kyoto

first: Akira

last: Niwa

address: 53, Shogoin-Kawahara-cho, Sakyo-ku

department: Department of Clinical Application

country: Japan

institute: CiRA, Kyoto University

summary:
Background: Blau syndrome, or early-onset sarcoidosis, is a juvenile-onset systemic granulomatosis associated with a mutation in Nucleotide-binding oligomerization domain 2 (NOD2). The underlying mechanisms of Blau syndrome leading to autoinflammation are still unclear, and there is currently no effective specific treatment for Blau syndrome.
Objectives: To elucidate the mechanisms of autoinflammation in Blau syndrome, we sought to clarify the relation between disease associated-mutant NOD2 and the inflammatory response in human samples.
Methods: Blau syndrome-specific induced pluripotent stem cells (iPSCs) lines were established. To precisely evaluate the in vitro phenotype of iPSC-derived cells, the disease-associated NOD2 mutation of iPSCs was corrected using a CRISPR/Cas9 system. We also introduced the same NOD2 mutation into a control iPSC line. These isogenic iPSCs were then differentiated into monocytic cell lineages, and the status of NF-κB pathway and proinflammatory cytokine secretion were investigated.
Results: We focused on the signals that upregulate the expression of NOD2, especially IFN-γ signaling. IFN-γ treatment of NOD2-mutant macrophages induced ligand-independent NF-κB activation and proinflammatory cytokine production. IFN-γ treatment acted as a priming signal through the up-regulation of NOD2 protein and recruitment of NOD2 on the basement membrane. Conversely, the production of proinflammatory cytokines by MDP, a ligand of NOD2, was decreased in mutant macrophages.
Conclusions: Our data support the significance of ligand-independent autoinflammation in the pathophysiology of Blau syndrome. Our comprehensive isogenic disease-specific iPSC panel provides a useful platform for probing therapeutic and diagnostic clues for the treatment of Blau syndrome patients.

overall_design: RNA-sequencing was conducted to identify the genes expressed in reponse to stimulation in different manners between WT and MT cells

first: Sanami

last: Takada

first: Megumu

last: Saito


title: Ribosome Incorporation into Somatic Cells Promotes Lineage Transdifferentiation towards Multipotency [RNA-Seq]

city: Koto-ku

first: Yutaka

state: Tokyo

institute: National Institute of Advanced Industrial Science and Technology (AIST)

summary:
Recently, we reported that bacterial incorporation induces cellular transdifferentiation of human fibroblasts. However, the bacterium-intrinsic cellular- transdifferentiation factor remained unknown. Here, we found that cellular transdifferentiation is caused by ribosomes. Ribosomes, isolated from both prokaryotic and eukaryotic cells, induce the formation of embryoid body-like cell clusters. Numerous ribosomes are incorporated into both the cytoplasm and nucleus through trypsin-activated endocytosis, which leads to cell-cluster formation. Although ribosome-induced cell clusters (RICs) express several stemness markers and differentiate into derivatives of all three germ layers in heterogeneous cell populations, RICs fail to proliferate, alter the methylation states of pluripotent genes, or contribute to teratoma or chimera formation. However, RICs express markers of epithelial–mesenchymal transition without altering the cell cycle, despite their proliferation obstruction. These findings demonstrate that incorporation of ribosomes into
host cells induces cell transdifferentiation and alters cellular plasticity.

overall_design: RNA-seq was conducted for ribosome-induced cell clusters (RICs) at day 0, 2, 5, 8, 11, 14.

first: Naofumi

last: Ito

first: Kaoru

last: Katoh

first: Hiroko

last: Kushige

first: Terumasa

last: Umemoto

first: Yu

last: Matsuzaki

first: Hiroshi

last: Kiyonari

first: Daiki

last: Kobayashi

first: Minami

last: Soga

first: Mariko

last: Yamane

first: Takumi

last: Era

first: Norie

last: Araki

first: Yasuhide

last: Furuta

first: Toshio

last: Suda

first: Yasuyuki

last: Kida

first: Kunimasa

last: Ohta


title: Sirtuin-1 is required for proper trophoblast differentiation and placental development

first: Mana

last: Parast

state: California

address: 9500 Gilman drive #0695

institute: University of California San Diego

summary: In this study we identified that Sirt1 is important for mouse trophoblast stem cell (TSC) differentiation. The transcriptome of wild-type and Sirt1-null TSC were analyzed to identify dysregulation of signaling pathways.

overall_design: In order to identify gene expression changes in TSC in the absence of Sirt1, we performed genome-wide expression profiling of Wild-type and Sirt1-null TSCs derived from a littermate pair of embryos on the 129S1/SvlmJ genetic background.

first: Kanaga

last: Rajan

first: Marwa

last: Khater

first: Francesca

last: Soncin

first: Don

last: Pizzo

first: Matteo

last: Moretto-Zita

first: Jonathan

last: Pham

first: Orysya

last: Stus

first: Pooja

last: Iyer

first: Veronique

last: Tache

first: Louise

last: Laurent

middle: M


title: Ribosome Incorporation into Somatic Cells Promotes Lineage Transdifferentiation towards Multipotency [Array]

first: Yutaka

last: Saito

summary: Recently, we reported that bacterial incorporation induces cellular transdifferentiation of human fibroblasts. However, the bacterium-intrinsic cellular- transdifferentiation factor remained unknown. Here, we found that cellular transdifferentiation is caused by ribosomes. Ribosomes, isolated from both prokaryotic and eukaryotic cells, induce the formation of embryoid body-like cell clusters. Numerous ribosomes are incorporated into both the cytoplasm and nucleus through trypsin-activated endocytosis, which leads to cell-cluster formation. Although ribosome-induced cell clusters (RICs) express several stemness markers and differentiate into derivatives of all three germ layers in heterogeneous cell populations, RICs fail to proliferate, alter the methylation states of pluripotent genes, or contribute to teratoma or chimera formation. However, RICs express markers of epithelial–mesenchymal transition without altering the cell cycle, despite their proliferation obstruction. These findings demonstrate that incorporation of ribosomes into host cells induces cell transdifferentiation and alters cellular plasticity.

overall_design: Gene expression was measured for ribosome-induced cell clusters (RICs) and control human dermal fibroblasts (HDFs).

first: Naofumi

last: Ito

first: Kaoru

last: Katoh

first: Hiroko

last: Kushige

first: Terumasa

last: Umemoto

first: Yu

last: Matsuzaki

first: Hiroshi

last: Kiyonari

first: Daiki

last: Kobayashi

first: Minami

last: Soga

first: Mariko

last: Yamane

first: Takumi

last: Era


title: DNA microarray analysis in hippocampus of Wild-type (WT), AtrxΔE2, and 5-aminolevulinic acid (5-ALA) treated AtrxΔE2 mice.

city: Kamakura

first: Satoshi

last: Kondo

state: Kanagawa

address: Tebiro 6-10-1

department: New Projects Development Division

institute: Toray Industries,Inc.

summary: 5-ALA that can be metabolized to porphyrins, protoporphyrin IX and hemin antagonizes decreased synaptic plasticity and cognitive deficits seen in ATR-X model (AtrxΔE2) mice. Our findings suggest a potential therapeutic strategy to target G-quadruplexes and decrease cognitive impairment associated with ATR-X syndrome.

overall_design: We administered 5-ALA (3mg/kg, p.o.) to AtrxΔE2 mice daily between post-natal day 30 to 90.

first: Norifumi

last: Shioda

first: Kohji

last: Fukunaga


title: Role of hypoxia in Diffuse Large B-cell Lymphoma: Metabolic repression and selective translation of HK2 facilitates development of DLBCL

status: Public on Jan 23 2018

city: Baltimore

first: Supriyo

last: De

state: Maryland

address: 251 Bayview Blvd

department: Laboratory of Genetics and Genomics

institute: NIA-IRP, NIH

summary: Published molecular profiling studies in patients with lymphoma suggested the influence of hypoxia inducible factor-1 alpha (HIF1α) targets in prognosis of DLBCL. Yet, the role of hypoxia in hematological malignancies remains unclear. We observed that activation of HIF1α resulted in global translation repression during hypoxic stress in DLBCL. Protein translation efficiency as measured using 35S-labeled methionine incorporation revealed a ≥50% reduction in translation upon activation of HIF1α. Importantly, translation was not completely inhibited and expression of clinically correlated hypoxia targets such as GLUT1, HK2, and CYT-C was found to be refractory to translational repression under hypoxia in DLBCL cells. Notably, hypoxic induction of these genes was not observed in normal primary B-cells. Translational repression was coupled with a decrease in mitochondrial function. Screening of primary DLBCL patient samples revealed that expression of HK2, which encodes for the enzyme hexokinase 2, was significantly correlated with DLBCL phenotype. Genetic knockdown studies demonstrated that HK2 is required for promoting growth of DLBCL under hypoxic stress. Altogether, our findings provide strong support for the direct contribution of HK2 in B-cell lymphoma development and suggest that HK2 is a key metabolic driver of the DLBCL phenotype.ne incorporation revealed a ≥50% reduction in translation upon activation of HIF1α. Importantly, translation was not completely blunted and expression of clinically correlated hypoxia targets such as GLUT1, HK2, and CYT-C was found to be refractory to translational repression under hypoxia in DLBCL cells. Notably, hypoxic induction of these genes was not observed in normal primary B-cells. Translational repression was coupled with decrease in mitochondrial function. Screening of DLBCL patient samples identified that expression of HK2, which encodes for the enzyme hexokinase 2, was significantly correlated with DLBCL phenotype. Genetic knockdown studies show that HK2 is required for promoting growth of DLBCL under hypoxic stress. Altogether, our findings provide more definitive proof of direct contribution of HK2 in development of B-cell lymphoma and suggest that HK2 is a key metabolic driver of DLBCL phenotype.

overall_design: Two human lymphoma cell lines, HLY-1 and SUDHL2, were cultured under conditions of hypoxia (N=3 biological replicates per cell line) or normoxia ((N=3 biological replicates per cell line), and gene expression microarray analysis employed to examine the global gene expression differences under these conditions.

first: Kavita

last: Bhalla

first: Susan

last: Jaber

first: Nahid

last: Nanaji

first: Karen

last: Underwood

first: Afshin

last: Beheshti

first: Ari

last: Landon

first: Binny

last: Bhandary

first: Paul

last: Bastian

first: Andrew

last: Evens

first: John

last: Haley

first: Brian

last: Polster

first: Ronald

middle: B

last: Gartenhaus


title: Testicular MTHFR deficiency may explain sperm DNA hypomethylation associated with high dose folic acid supplementation

city: Montreal

first: Jacquetta

last: Trasler

state: Quebec

address: 1001 Decarie Boul, Block E, ES1.4380

department: Pharmacology & Therapeutics; Human Genetics

country: Canada

institute: McGill University

type: Methylation profiling by high throughput sequencing

summary: Supplementation with high doses of folic acid, an important mediator of one-carbon transfers for DNA methylation, is used clinically to improve sperm parameters in infertile men. We recently detected an unexpected loss of DNA methylation in the sperm of idiopathic infertile men after 6 months of daily supplementation with 5mg folic acid (>10× the daily recommended intake-DRI), exacerbated in men homozygous for a common variant in the gene encoding an important enzyme in folate metabolism, methylenetetrahydrofolate reductase (MTHFR 677C>T). To investigate the epigenomic impact and mechanism underlying effects of folic acid on male germ cells, wildtype and heterozygote mice for a targeted inactivation of the Mthfr gene were fed high-dose folic acid (10× the DRI) or control diets for six months. No changes were detected in general health, sperm counts or methylation of imprinted genes.  Reduced representation bisulfite sequencing revealed sperm DNA hypomethylation in Mthfr+/- mice on the 10× diets. Wildtype mice demonstrated sperm hypomethylation only with a very high dose (20×) of folic acid for 12 months. Testicular MTHFR protein levels decreased significantly in wildtype mice on the 20× diet but not in those on the 10× diet, suggesting a possible role for MTHFR deficiency in sperm DNA hypomethylation. In-depth analysis of the folic acid-exposed sperm DNA methylome suggested mouse/human susceptibility of sequences with potential importance to germ cell and embryo development. Our data provide evidence for a similar cross-species response to high dose folic acid supplementation, of sperm DNA hypomethylation, and implicate MTHFR downregulation as a possible mechanism.

overall_design: 27 total samples, n=3-5 for each group compared treated (10FS/20FS) to specific Control samples (CD)

first: Mahmoud

last: Aarabi

last: Christensen

first: Donovan

last: Chan

first: Daniel

last: Leclerc

first: Mylene

last: Landry

first: Lundi

last: Ly

first: Rima

last: Rozen


title: Transcript dynamics in classically and alternatively activated macrophages

city: Ansan

first: Young Gyu

last: Chai

state: Gyeonggi-do

address: 55 Hanyangdaehak-ro

department: Molecualr & Life Science

country: South Korea

institute: Hanyang Univ.

summary: Using RNA-seq, we have reported that signaling crosstalk among IFN-γ and LPS is integrated at the level of transcriptome and have associated with TFs and TcoFs changes. In addition, we also argue that the transcriptional differences between BMDMs and RAW264.7 macrophage cell line as well as IL-4 and IL-13 on M2 macrophages activation.

overall_design: Examination of effects of IFN-γ mediated LPS treated versus only LPS treated BMDMs, IL-4 and IL-13 treated BMDMs and LPS treated BMDMs and RAW264.7 macrophage cell line, were generated by deep sequencing on an Illumina HiSeq 2500 (101 cycles PE lane).


title: Novel molecular and phenotypic insights into congenital lung malformations

first: Michael

middle: Patrick

last: Morley

address: 3400 Civic Center Blvd, Bldg 421

department: Penn Cardiovascular Institute

institute: Perelman School of Medicine at the University of Pennsylvania

summary: The purpose of this study was to identify gene expression changes associated with congenital lung malformations.

overall_design: Lung tissue was collected from infants undergoing lobectomy for a congenital lung lesion. From each patient, a sample was obtained from the congenital lung lesion, and an adjacent section of unaffected lung (labeled healthy).

middle: T

last: Swarr

first: Edward

last: Morrisey


title: Targeting multiple effector pathways in pancreatic ductal adenocarcinoma with a G-quadruplex-binding small molecule

city: Cambridge

first: Giovanni

last: Marsico

address: Robinson Way

institute: CRUK Cambridge Institute

summary: Human pancreatic ductal adenocarcinoma (PDAC) involves the dysregulation of multiple signalling pathways. A novel approach to the treatment of PDAC is described, involving the targeting of cancer genes in PDAC pathways having over-representation of G-quadruplexes, using the quadruplex-binding compound CM03. This has been designed by computer modelling, is a potent inhibitor of cell growth in PDAC cell lines and has anti-cancer activity in PDAC models, with a superior profile compared to gemcitabine, a commonly used therapy.   Whole-transcriptome RNA-seq methodology has been used to analyse for the first time the effects of a quadruplex-binding small molecule on gene expression. This has revealed the down-regulation of a large number of genes, rich in putative quadruplex elements and involved in essential pathways of PDAC survival, metastasis and drug resistance.   The changes produced by CM03 represent a global response to the complexity of human PDAC, and may be applicable to other currently hard-to-treat cancers.

overall_design: 12 libraries for MIA PACA-2 cell line (6 and 24 hrs treatment with CM03 drug plus untreated control); 12 libraries for PANC-1 cell line (6 and 24 hrs treatment with CM03 drug plus untreated control); 28 libraries for MIA PACA-2 cell line (6 and 24 hrs treatment at 3 different concentrations with gemcitabine drug plus untreated control); 52 libraries for PANC-1 cell line (6 and 24 hrs treatment at 5 different concentrations with gemcitabine drug plus untreated control). Each condition has 3 or 4 biological replicates. See details below for concentration, treatment and replicates.

first: Chiara

last: Marchetti

first: Katherine

middle: G

last: Zyner

first: Stephan

last: Ohnmacht

first: Mathew

last: Robson

first: Shozeb

last: Haider

first: Jennifer

last: Morton

first: Tam

last: Vo

first: Sarah

last: Laughlin-Toth

first: Ahmed

last: Ahmed

first: Gloria

last: Di Vita

first: Ingrida

last: Pazitna

first: Mekala

last: Gunaratnam

first: Rachael

last: Besser

first: Ana

last: Gomes Andrade

first: Seckou

last: Diocou

first: Jeremy

last: Pike

first: David

last: Tannahill

first: R

last: Pedley

first: T

last: Evans

first: W

last: Wilson

first: Shankar

last: Balasubramanian

first: Stephen

last: Neidle


title: Transcriptional impact of conditional loss of Notch2 from adult NSCs

city: Basel

first: Verdon

last: Taylor

address: Mattenstrasse 28

department: Department Biomedicine

institute: University of Basel

summary: Adult NSCs maintenance depends on intakt Notch signaling. The role of Notch2 has only been marginally studied in adult neurogenesis. To adress the role of the Notch2 paralogue we conditionally deleted Notch2 from the adult SVZ and analysed the Notch2-deficient cells at distinct time points.

overall_design: Loss of Notch2 was conditionally induced in adult, Notch2 floxed mice. Recombined cells were sorted and processed ex vivo. We used microarrays to detail transcriptional changes upon gene deletion.

first: Anna

last: Engler

first: Chiara

last: Rolando

first: Claudio

last: Giachino

first: Ichiko

last: Saotome

first: Andrea

last: Erni

first: Callum

last: Brien

first: Runrui

last: Zhang

first: Ursula

last: Zimber-Strobl

first: Freddy

last: Radtke

first: Spyros

last: Artavanis-Tsakonas

first: Angeliki

last: Louvi


title: Plasticity of life-long calorie restricted C57BL/6J mice in adapting to a medium-fat diet intervention at old age

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

country: Netherlands

institute: Wageningen University

summary: Calorie restriction (CR) is a dietary regimen that supports healthy aging. In this study we investigated the systemic and liver-specific responses caused by a diet switch to a medium-fat (MF) diet in 24-month-old life-long, CR-exposed mice. This study aimed to increase the knowledge base on dietary alterations of gerontological relevance. Nine-week-old C57BL/6J mice were exposed either to a control, CR or MF diet. At the age of 24 months, a subset of mice of the CR group was transferred to ad libitum MF feeding (CR-MF).The mice were sacrificed at the age of 28 months, then biochemical and molecular analyses were performed. Our results showed that, despite the long-term exposure to the CR regimen, mice in the CR-MF group displayed hyperphagia, rapid weight gain, and hepatic steatosis. However, no hepatic fibrosis/injury or alteration in CR-improved survival was observed in the diet switch group. The liver transcriptomic profile of CR-MF mice largely shifted to a profile similar to the MF-fed animals but leaving ~22% of the 1578 differentially regulated genes between the CR and MF diet groups comparable with the expression of the life-long CR group. Therefore, although the diet switch was performed at an old age, the CR-MF-exposed mice showed plasticity in coping with the challenge of a MF diet without developing severe liver pathologies.

overall_design: Male C57BL/6J mice arrived at 7 weeks of age and allowed to acclimate for 2 weeks, receiving standard AIN-93G upon arrival. At the start of the diet intervention the mice were 9 weeks old and randomly distributed into three intervention groups: 1) Control diet receiving AIN-93W diet ad libitum; 2) CR diet receiving AIN-93W-CR in portions containing 70E% of the mean energy intake of the group of the control mice were provided each day at 15.30; 3) medium fat diet (MF) receiving AIN-93W-MF ad libitum. The long-term dietary invention was continued until a sacrifice at the age of 28 month, but in addition to the three diet groups, at 24 month the animals in the CR diet group were randomly assigned either to remain on the CR diet or undergo a diet switch to the MF diet (CR-MF). We performed various measurements on metabolic parameters and gene expression analysis. This entry represents the microarray data.

first: Fenni

last: Rusli

first: Mark

middle: V

last: Boekschoten

first: Vincenzo

last: Borelli

first: Chen

last: Sun

first: Carolien

last: Lute

first: Aswin

last: Menke

first: Joost

last: van den Heuvel

first: Stefano

last: Salvioli

last: Franceschi

first: Michael

last: Müller

first: Wilma

last: Steegenga


title: Ribosome Incorporation into Somatic Cells Promotes Lineage Transdifferentiation towards Multipotency

first: Yutaka

last: Saito

overall_design: Refer to individual Series


title: The coding and noncoding transcriptome of Neurospora crassa

city: Heidelberg

last: Brunner

address: Im Neuenheimer Feld 328

country: Germany

institute: BZH

type: Non-coding RNA profiling by high throughput sequencing

summary:
We present the NEUTRA database, a catalogue of the Neurospora transcriptome, based on RNA-Seq, RNAPII ChIP-Seq and ribosome fractionation studies.
The database also includes genome-wide binding sites of transcription factors involved in regulation of circadian gene expression.
In a comprehensive analysis of the transcriptome we identified and characterized 1478 long intergenic non-coding RNAs (lincRNAs) and 1056 natural antisense transcripts.
Both classes of long non-coding RNAs accumulate at lower levels than protein-coding mRNAs and they are considerably shorter.
We also confirmed annotated splice sites and identified new splice junctions and alternative splice sites, which are rare in Neurospora.
Our analysis revealed that the vast majority of lincRNAs and antisense transcripts do not contain introns.
NEUTRA is the first database to provide a collection of genome-wide deep sequencing data for the model organism Neurospora crassa and thus provides a tool that facilitates future research.

overall_design: RNA-seq of Neurospora crassa.

first: Ibrahim

last: Cemel

first: Nati

last: Ha

first: Geza

last: Schermann

first: Shusuke

last: Yonekawa


title: Fetal sepsis in utero induces disruption of gene networks involved in cardiac morphogenesis in a nonhuman primate model of infection-induced preterm labor

city: Seattle

first: James

middle: William

last: MacDonald

state: WA

address: 4225 Roosevelt Way NE

department: Environmental and Occupational Health Sciences

institute: University of Washington

summary: Fetal lung and brain injury have been linked with intrauterine exposure to amniotic fluid (AF) cytokines, but injury to other fetal organs are not well characterized. Fetal cardiac injury was suspected based on an ultrasound study suggesting diastolic dysfunction, but not previously confirmed or characterized. In this study we explore the relationship between cytokine exposure due to exposure to group B streptococcus and cardiac gene expression.

platform_organism:
Macaca fascicularis
Macaca mulatta

overall_design: Fetal cardiac tissue was obtained after Cesarean section and fetal necropsy in cases known to have fetal sepsis (N=5) or from saline controls (N=5) in a nonhuman primate model of preterm labor (Macaca nemestrina). Fetal sepsis was induced by choriodecidual inoculation of Group B Streptococcus (N=4) or intra-amniotic inoculation of E. coli (N=1). RNA was extracted from fetal hearts and profiled by microarray. Statistical analysis included Wilcoxon rank sum, single gene analysis, gene set analysis. Results were validated by quantitative RT-PCR.

first: Timothy

last: Mitchell

middle: W

first: Theodor

middle: K

last: Bammler

first: Lakshmi

last: Rajagopal

first: Kristina

last: Adams Waldorf


title: PTSD or resilient - Genetic and Epigenetic association studies in police officers

city: Amsterdam

first: Peter

last: Henneman

state: Noord Holland

address: Meibergdreef 9

department: Clinical Genetics

institute: Academic Medical Center Amsterdam

type: Methylation profiling by genome tiling array

summary: Epigenetic and genetic profiles (N=67) were based on the Illumina Infinium HumanMethylation450 BeadChip. We searched for differentially methylated probes (DMPs) and regions (DMRs). For genetic associations we analyzed the CpG-SNPs present on the array

overall_design: Bisulphite converted DNA obtained from whole blood were analysed using the the Infinium HumanMethylation450 BeadChip array

first: I

last: Krzyzewska

first: J

last: Ensink

first: L

last: Nawijn

first: A

last: Mul

first: S

last: Koch

last: Venema

first: V

last: Shankar

middle: L

last: Frijling

first: D

middle: J

last: Veltman

first: R

last: Lindauer

first: M

last: Olff

last: Mannens

last: van Zuiden

first: P


title: Pancreatic gene expression during recovery after pancreatitis reveals unique transcriptome profiles

first: Ting

last: Wang

address: 1100 Fairview Ave. N.

department: Clinical Research

institute: Fred Hutchinson Cancer Research Center

summary: Histological resolution of the murine pancreas occurs within one week after injury. Whether histological resolution constitutes pancreatic recovery at a molecular level is not known. We performed RNA-sequencing on the recovering pancreas to determine the transcriptomic profile within the histologically recovered pancreas. We show that although there is histological resolution one week after injury in mice, compared to baseline (non-injured pancreas), there are still numerous differentially expressed genes (DEGs) at one and even two weeks after injury. Overall, the findings suggest the actual recovery takes longer than initially thought given the differential transcriptomic profile in the pancreas two weeks after injury compared to the baseline pancreas. There is also the possibility of a novel emerging pancreatic transcriptome upon recovery.

overall_design: Acute pancreatitis was induced by caerulein hyperstimulation in both male and female C57BL/6 mice. Total RNA was extracted from the head of the murine pancreas in mice at baseline (non-injured; n=8), day 7 (post-injury; n=8), and day 14 (post-injury; n=7). Total stranded RNA libraries (ribo-depleted) were generated and sequenced on the Illumina NextSeq 500 NGS platform. RNA-seq data was analyzed for differentially expressed genes between baseline and day 7 and between baseline and day 14.

first: Sohail

middle: Z

last: Husain

first: Kristy

last: Boggs


title: BL-Hi-C is an efficient and sensitive approach for capturing structural and regulatory chromatin interactions

city: Shenzhen

first: Zhengyu

last: LIANG

state: Guangzhou

address: 1088 Xueyuan Avenue

department: Department of Biology

country: China

institute: Southern University of Science and Technology

summary: We propose a novel in situ Hi-C method named Bridge-Linker-Hi-C (BL-Hi-C) for structural and regulatory chromatin interactions capture by restriction enzyme targeting and two-step proximity ligation. It improves the sensitivity and specificity for active chromatin loop detection and can reveal a better enhancer-promoter regulatory architecture than conventional method at a lower sequencing depth and with a simpler protocol.

overall_design: BL-Hi-C was performed in K562 cell for 13 samples: 5 samples for enrichment analysis; 4 samples for enzyme digestion validation and 4 samples for proximity ligation validation.

last: Liang

first: Michael

middle: Q

last: Zhang

first: Yang

last: Chen


title: A heterochromatin gene signature unveils HP1α mediating neuroendocrine prostate cancer development and aggressiveness

status: Public on Jan 22 2018

city: Vancouver

first: Yuzhuo

state: BC

address: 2660 Oak Street

department: Vancouver Prostate Centre

institute: University of British Columbia

summary: Neuroendocrine prostate cancer (NEPC) is a lethal subtype of prostate cancer (PCa) with hyperchromatic nuclei being a distinguishing histopathological feature. Here we show that, underlying this distinct nuclear structure, heterochromatin related genes are significantly enriched in NEPC. Among them, heterochromatin protein 1α (HP1α) expression is increased early in NE transdifferentiation and is consistently elevated in clinical NEPC samples. Functional studies showed that HP1α knockdown attenuates NEPC tumor growth by inhibiting proliferation and survival. Its ectopic expression significantly promotes NE transdifferentiation in adenocarcinoma cells subjected to androgen deprivation treatment. To identify mechanisms underlying the function of HP1α involved in NEPC cell proliferation and adenocarcinoma cell NE transdifferentiation, we analyzed gene expression profiles from stable NEPC cell lines with control or HP1α knockdown, and adenocarcinoma cell lines with control or HP1α overexpression, respectively.

overall_design: NCI-H660 cell was transduced with lentiviral particles delivering control or HP1α shRNA. V16D cell was transduced with lentiviral particles delivering control or HP1α overexpressing plasmid. To induce neuroendocrine phenotype in V16D cell, cells were starved with phenol-red free RPMI-1640  containing 10% Charcoal-Stripped Serum for 24 hours, then cultured with the addition of 10 μM enzalutamide. Total RNA was extracted from stable cell lines, and applied for gene expression profiling analysis using Agilent SurePrint G3 Human Gene Expression v3 8x60K array.

first: Xinpei

last: Ci

first: Anne

last: Haegert

first: Dong

last: Lin


title: KSHV chromatin looping facilitates effective gene expression: high-resolution mapping of long-range chromatin interactions in the KSHV genome [Hi-C]

city: Sacramento

first: Clifford

middle: G.

last: Tepper

address: 4645 2nd Avenue, Room 2300A

department: Biochemistry and Molecular Medicine

institute: UC Davis School of Medicine

summary: The three-dimensional structure of chromatin via formation of genomic loops allows cellular RNA polymerase II to access distal promoters, thus it has a significant impact on the outcome of gene expression. The Kaposi’s sarcoma-associated herpesvirus (KSHV) entire lytic transcriptional program of 80 plus genes is initiated by a single viral transactivator, K-Rta. Here we examined the relationship between the KSHV genomic structure and K-Rta recruitment sites with unbiased Capture Hi-C analyses. The studies showed that KSHV forms a unique genomic structure, which coordinates a domain-specific viral gene expression program via K-Rta recruitment, and identified a number of direct physical, long-range, and dynamic genomic interactions. Chromatin immunoprecipitation-sequencing analyses identified a limited number of K-Rta recruitment sites in the KSHV genome, including the K12 and PAN RNA promoters. KSHV engineered to harbor point mutations in the K-Rta-responsive elements (RE) at these promoters significantly attenuated not only the direct downstream gene, but also distal viral gene expression in a domain-specific manner. Despite the long distance between two RE sites in the linear orientation, efficient recruitment of K-Rta to either promoter required intact K-Rta REs at both promoters. Finally, inducible genomic loops generated during KSHV reactivation occurred preferentially at K-Rta recruitment sites. Mutation of a K-Rta RE inhibited formation of inducible genomic loops, gene expression of its destination of looping genes, and KSHV replication in de novo infected human gingival epithelial cells. These results suggest that the KSHV genome is programed to form both stable and inducible chromatin hubs for effective KSHV gene expression, and partly explains why K-Rta has such a dominant effect on KSHV lytic gene transcription.

overall_design: The overall goal of these studies was to define the long-range chromatin interactions occurring within the KSHV genome as well as between the KSHV and human host cell genomes both during latency and viral reactivation. These were defined in an unbiased manner using Hi-C analysis, which is chromosome conformation capture (3C) combined with next-generation sequencing (NGS). A modified protocol, referred to as Capture Hi-C, was utilized to selectively enrich interactions involving KSHV by performing target enrichment with a custom-designed KSHV genomic capture probe library. Experiments were performed in two cell lines, TREx-(F3H3)-K-Rta BCBL-1 and iSLK.r219 cells, in which KSHV can be reactivated via doxyclyine-induced expression of K-Rta. Accordingly, Capture Hi-C sequencing libraries were prepared from replicate cultures of each cell line cultured in the absence (i.e., latency) or presence (i.e., reactivation) of doxycycline for 24 hours.

first: Yoshihiro

last: Izumiya

first: Mel

last: Campbell


title: Id proteins suppress E2A-driven iNKT cell development prior to TCR selection

city: Durham

first: Yuan

last: Zhuang

state: NC

address: 207 Research Drive

department: Immunology

institute: Duke University

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


title: Circulating miRNA profiles from serum of women at high-risk for breast cancer

city: Burlington

first: Nicholas

middle: H

last: Farina

state: VT

address: 89 Beaumont Avenue

department: Biochemistry

institute: University of Vermont Cancer Center

summary:
Circulating microRNAs (c-miRNAs) have emerged as measurable biomarkers (liquid biopsies) for cancer detection. The goal of our study was to identify novel biomarkers to predict long-term breast cancer risk in cancer-free women.  We evaluated the ability of c-miRNAs to identify women most likely to develop breast cancer by profiling miRNA from serum obtained long before diagnosis.  24 breast cancer cases and controls (matched for risk and age) were identified from women enrolled in the High-Risk Breast Program at the UVM Cancer Center.
We used Affymetrix miRNA v4 microarrays to interrogate miRNAs (miRBase v20) in the serum of cancer-free women at high-risk for breast cancer.  The 24 cases developed breast cancer at least 6 months (average of 3.2 years) and the 24 controls remain cancer-free.

overall_design: 24 cases and 24 matched controls

first: Jon

middle: E

last: Ramsey

first: Melissa

last: Cuke

first: Thomas

middle: P

last: Ahern

first: David

last: Shirley

first: Janet

last: Stein

first: Gary

first: Jane

last: Lian

first: Marie

last: Wood


title: KSHV chromatin looping facilitates effective gene expression: high-resolution mapping of K-Rta binding sites in the KSHV genome

state: CA


title: Transcriptional profiling of putative J protein Jjj1 in Candida glabrata

status: Public on Jan 21 2018

city: Memphis

first: Katherine

middle: S

last: Barker

address: 881 Madison Avenue, Room 305

department: Clinical Pharmacy

institute: University of Tennessee Health Science Center

summary: In the present study the gene encoding the putative J protein Jjj1 was deleted in a susceptible dose dependent clinical isolate resulting in decreased susceptibility to fluconazole compared to wild type. When JJJ1 was deleted in combination with the transcription factor PDR1 the fluconazole susceptibility was consistently only one fold higher from the strain deleted for PDR1 alone. Expression profiling was conducted to determine which genes have altered expression in the absence of JJJ1 and which of these alterations are dependent on PDR1 expression.

overall_design: We examined the genome-wide gene expression profiles of the parent strain, clinical isloate SM1 and its jjj1Δ and pdr1Δjjj1Δ derivatives in duplicate

first: Sarah

last: Whaley

first: Kelly

last: Caudle

first: Lucia

last: Simonicova

first: Qing

last: Zhang

first: W

last: Moye-Rowley

first: P

middle: D

last: Rogers


title: Skeletal muscle “ex vivo” mitochondrial respiration parallels decline in vivo oxidative capacity, cardiorespiratory fitness and muscle strength: the Baltimore Longitudinal Study of Aging.

first: Supriyo

last: De

address: 251 Bayview Blvd

institute: NIA-IRP, NIH

summary: Mitochondrial function in human skeletal muscle declines with aging, possibly because of damage accumulation caused by reactive oxygen species (ROS) produced during oxidative phosphorylation. However, most evidence for such a decline comes from studies based on indirect methods and the clinical relevance of this decline with respect to physical function has not been clearly delineated. We hypothesized that mitochondrial respiration objectively assessed in permeabilized human muscle fibers declines with aging, correlates with phosphocreatine post exercise recovery rate (kPCr) assessed by 31P magnetic resonance spectroscopy (MRS) and with muscle performance and aerobic fitness. Mitochondrial respiration was assessed by high resolution respirometry in saponin-permeabilized fibers from vastus lateralis muscle biopsies of 38 participants from the Baltimore Longitudinal Study of Aging (BLSA) (21 men, age 24-91 years) who also had available measures of peak oxygen consumption (VO2max) from treadmill tests, gait speed in different tasks, 31P magnetic resonance spectroscopy and isokinetic knee extension strength. Results indicated a significant reduction in mitochondrial respiration in older age (p<0.05) that was independent of potential confounders. Mitochondrial respiratory capacity was also associated with VO2max, muscle strength, phosphocreatine post-exercise recovery rate (kPCr), and time in 400m walk (p < 0.05). A moderate negative trend toward significance (p = 0.074) was observed between mitochondrial respiration and BMI. Finally, transcriptional profiling revealed a downregulation of mitochondrial gene networks with aging (p < 0.05). Overall, our findings reinforce the notion that mitochondrial function declines with age and is implicated in the parallel decline of muscle performance and cardiorespiratory fitness.

overall_design: RNA obtained from the vastus lateralis of a subset of the participants (N=24), aged 25-84 years (mean= 67 ± 15) was analyzed by Illumina HT-12v4 microarrays, with 4 young (Y, 2M/2F), 5 middle-aged (MA, 2M, 3F), and 14 old (O, 11M, 3F) BLSA participants.

first: Marta

last: Gonzalez-Freire

first: Paul

last: Scalzo

first: Jarod

last: D’Agostino

first: Zenobia

middle: A

last: Moore

first: Alberto

last: Diaz-Ruiz

first: Elisa

last: Fabbri

first: Ariel

last: Zane

first: Brian

last: Chen

first: Kevin

last: Becker

first: Elin

last: Lehrmann

first: Linda

last: Zukley

first: Chee

last: Chia

first: Toshiko

last: Tanaka

middle: M

last: Coen

first: Michel

last: Bernier

first: Rafael

last: de Cabo

first: Luigi

last: Ferrucci


title: Gene expression analysis of chimeric antigen receptor-transduced T cells after antigenic stimulation

city: Tokyo

first: Yuki

last: Kagoya

address: 35 Shinanomachi, Shinjuku

department: School of Medicine

institute: Keio University

summary:
Adoptive transfer of anti-CD19 chimeric antigen receptor (CAR)-engineered T cells has shown impressive clinical responses in patients with refractory B-cell malignancies.  However, therapeutic effects of CAR-T cells targeting other hematologic malignancies and solid tumors are not yet satisfactory.  Although inefficient tumor trafficking and multiple immunosuppressive molecules impede CAR-T cell effector responses, signals delivered by the current CAR constructs may still be insufficient to fully activate antitumor T cell functions.  Optimal T cell activation and proliferation requires multiple signals including T cell receptor (TCR) engagement (signal 1), costimulation (signal 2), and cytokine engagement (signal 3).  CAR genes developed to date contain a CD3z domain and costimulatory domain(s), but not a domain to transmit signal 3.
In this study, we have developed a novel CAR construct capable of inducing cytokine signaling in an antigen-dependent manner.  The new generation CD19 CAR encodes a cytoplasmic domain of IL-2RB and STAT3-binding YXXQ motif together with CD3z and CD28 domains (28-DIL2RB-z (YXXQ)).  The 28-DIL2RB-z (YXXQ) CAR-T cells showed antigen-dependent JAK-STAT, especially the STAT3-mediated pathway activation, which promoted their proliferation and prevented terminal differentiation.  The 28-DIL2RB-z (YXXQ) CAR-T cells demonstrated superior in vivo persistence and antileukemia effects compared with the currently used CARs in multiple tumor models.

overall_design: Human CD8+ T cells derived from four different healthy donors were retrovirally transduced with individual anti-CD19 CAR genes (28-z, BB-z and 28-DIL2RB-z (YXXQ)) and were stimulated with the CD19+ B-cell acute lymphoblastic leukemia cell line NALM-6.  RNA was collected 4 hours, 24 hours and 72 hours after stimulation and gene expression profiles were analyzed by Affymetrix Human Gene 2.0 ST Array (gene-level analysis).

first: Naoto

last: Hirano


title: Next Generation Sequencing Facilitates Quantitative Analysis of ChIP-sequence of U2OS cells expressing FANCD2-3x FLAG under mild replication stress by low dose aphidicolin (APH)

status: Public on Jan 20 2018

city: Kyoto

first: Masamichi

last: Ishiai

state: Kyoto

address: Yoshidakonoe-cho, Sakyo-ku

department: Radiation Biology Center

institute: Kyoto University

summary: Genome-wide distribution of FANCD2 protein in chromatin under replication stress.

overall_design: Examination of APH treat and non-treat samples with anti-FLAG Ab-ChIP, in the presence of normal-IgG ChIP and input cotrol

first: Yusuke

last: Okamoto

first: Kazuto

last: Kugou

first: Watal

last: Iwasaki

first: Chinatsu

last: Ohki

first: Hideki

last: Innan

first: Minoru

last: Takata

first: Kunihiro

last: Ohta


title: Mechanistic differences in neuropathic pain modalities revealed by correlating behavior with global expression profiling (RNA-Seq)

status: Public on Jan 19 2018

city: Los Angeles

first: Giovanni

last: Coppola

address: 1524 Gonda, 695 Charles Young Drive South

department: Psychiatry and Neurology

institute: UCLA

summary: Chronic neuropathic pain is a major morbidity of neural injury, yet its mechanisms are incompletely understood. Hypersensitivity to previously non-noxious stimuli (allodynia) is a common symptom. Here, we demonstrate that the onset of cold hypersensitivity precedes tactile allodynia and this temporal divergence was associated with major differences in global gene expression in dorsal root ganglia. Transcripts whose expression correlate with the onset of cold allodynia were nociceptor-related whereas those correlating with tactile hypersensitivity were enriched for immune cell activity. Selective ablation of TrpV1 lineage nociceptors resulted in mice that did not acquire cold allodynia but developed normal tactile hypersensitivity. Whereas depletion of macrophages or T cells reduced neuropathic tactile allodynia but not cold hypersensitivity. We conclude that neuropathic pain is contributed to by reactive processes of sensory neurons and immune cells, each leading to distinct forms of pain hypersensitivity, potentially allowing effective drug development targeted to each pain modality.

overall_design: High temporal analysis of global gene expression in the DRG following spared nerve injury (SNI) correlated with behavior taken at the same time points. Expression and behavioral sensitivity taken at least daily over the first 10 days post SNI.

first: E

last: Cobos

first: C

last: Nickerson

first: F

last: Gao

first: V

last: Chandran

first: I

last: Bravo-Caparrós

first: R

last: González-Cano

first: P

last: Riva

first: N

last: Andrews

first: A

last: Latremoliere

last: Seehus

first: G

last: Perazzoli

middle: R

last: Nieto

last: Joller

first: M

last: Painter

last: Ma

first: T

last: Omura

last: Chesler

first: D

last: Geschwind

last: Rangachari

last: Woolf

last: Costigan


title: Loss of Capicua alters early T cell development and predisposes mice to T cell lymphoblastic leukemia/lymphoma

status: Public on Jan 18 2018

city: Edmonton

first: Qiumin

last: Tan

state: Alberta

address: 6-16 Medical Sciences Building

department: Cell Biology

institute: University of Alberta

summary: We isolated RNAs from hematopoietic stem and progenitor cells (HSPCs). We compared two groups of mice: Cicflox/flox (control) and UBC-cre/ERT2; Cicflox/flox (knockout), two weeks post tamoxifen treatment. Following RNA isolation, RNAseq was performed and gene expression profiles were compared between controls and conditional knockouts.

overall_design: Examination of gene expression in hematopoietic stem and progenitor cells (HSPCs) in two mouse groups.

first: Ying-Wooi

last: Wan

first: Zhandong

last: Liu

first: Huda

middle: Y

last: Zoghbi


title: Dbf4 recruitment by forkhead transcription factors defines an upstream rate-limiting step in determining replication origin firing

city: Montpellier

first: romain

last: forey

address: 142 avenue de la Cardonille

country: France

institute: IGH

summary: Initiation of eukaryotic chromosome replication follows a spatiotemporal program. Current model suggests that replication origins compete for a limited pool of initiation factors. However, it remains to be answered how these limiting factors are preferentially recruited to early origins. Here, we report that Dbf4 is enriched at early origins through its interaction with forkhead transcription factors Fkh1 and Fkh2. This interaction is mediated by Dbf4 C-terminus and was successfully reconstituted in vitro. An interaction defective mutant dbf4ΔC phenocopies fkh alleles in terms of origin firing. Remarkably, genome-wide replication profiles reveal that the direct fusion of the DNA-binding domain of Fkh1 to Dbf4 restores the Fkh-dependent origin firing, but specifically interferes with the pericentromeric origin activation. Furthermore, Dbf4 directly interacts with Sld3 and promotes the recruitment of downstream limiting factors. These data suggest that Fkh1 targets Dbf4 to a subset of non-centromeric origins to promote early replication, in a manner that is reminiscent to the recruitment of Dbf4 to pericentromeric origins by Ctf19.

overall_design: NGS-based analysis of origin activity in HU-arrested S. cerevisiae cells

first: Dingqiang

last: Fang

first: Armelle

last: Lengronne

first: Di

last: Shi

first: Romain

last: Forey

first: Magdalena

last: Skrzypczak

first: Krzysztof

last: Ginalski

first: Changhui

last: Yan

first: Xiaoke

last: Wang

first: Qinhong

last: Cao

first: Philippe

last: Pasero

first: Huiqiang

last: Lou


title: Mitotic chromosomes fold by condensin-dependent helical winding of chromatin loop arrays

city: Worcester

first: Job

last: Dekker

state: Massachusetts

address: 368 Plantation Street

department: Program in Systems Biology

institute: UMass Medical School

summary: Mitotic chromosome morphogenesis occurs through condensin-mediated disassembly of the interphase conformation and formation of extended prophase loop arrays that then shorten by condensin-dependent helical winding

sample_organism:
Gallus gallus
Homo sapiens

platform_organism:
Gallus gallus
Homo sapiens

overall_design: Chromosome organization investigation through Hi-C

first: Johan

last: Gibcus

first: Anton

last: Goloborodko

first: Kumiko

last: Samejima

first: Itaru

first: Natalia

last: Naumova

first: Masato

middle: T

last: Kanemaki

first: Linfeng

last: Xie

first: James

last: Paulson

first: William

middle: C

last: Earnshaw

first: Leonid

last: Mirny


title: Mechanistic differences in neuropathic pain modalities revealed by correlating behavior with global expression profiling

first: Giovanni

last: Coppola

overall_design: Refer to individual Series


title: Identification of cadmium resistance and transcriptome sequence analysis under Cadmium Stress in Verbena bonariensis

city: Chengdu

first: Zhenyu

last: Bai

address: 211 Huimin Road, Wenjiang District

institute: Sichuan Agricultural University

summary: Verbena bonariensis is a species with excellent garden plant, good environmental adaptability and great potential for future development.Cadmium has caused serious heavy metal pollution in the soil, which has posed a great threat to plant growth. In this study, Illumina sequencing technology was used to sequence the transcriptome of Verbena bonariensis leaf under normal and Cd stress, respectively. In total, 95,013 transcripts and 63021 genes with an average length of 923 bp and 1,246 bp were constructed from the clean sequence reads, respectively. And 1037 DEGs were found in response to cadmium treatment, of which 10 were selected for qRT-PCR. In conclusion, this study first identified the Verbena bonariensis as a heavy metal tolerant plant and provided the first large-scale transcriptional data set in response to cadmium stress. Our research will help to understand the mechanism of resistance to Cd in the Verbena bonariensis and provide clues for further studies on the relationships between plants and heavy metals in other Verbenaceae plants.

overall_design: transcriptome of two kinds of plant leaves which were treated under normal and cadmium treatment was sequenced by the Illumina sequencing technology


title: The rate of ageing-associated DNA methylation dynamics is a molecular readout of lifespan variation amongst mammalian species [Macaque_450k]

status: Public on Jan 17 2018

city: London

first: Robert

last: Lowe

address: 4 Newark Street

department: Blizard Institute

institute: QMUL

summary: We used blood samples from Macaque to determine rate of change of methylation with respect to age.

overall_design: Illumina 450K profiling of Macaque at various age.


title: SLFN11 blocks stressed replication forks independently of ATR

city: Bethesda

first: Junko

last: Murai

address: 37 Convent Dr.

institute: NCI

summary: SLFN11 sensitizes cancer cells to a broad range of DNA-targeted therapies. Here we show that, in response to replication stress induced by camptothecin, SLFN11 tightly binds chromatin at stressed replication foci via RPA1 together with the replication helicase subunit MCM3. Unlike ATR, SLFN11 neither interferes with the loading of CDC45 and PCNA nor inhibits the initiation of DNA replication but selectively blocks fork progression while inducing chromatin opening across replication initiation sites. The ATPase domain of SLFN11 is required for chromatin opening, replication block and cell death but not for the tight binding of SLFN11 to chromatin. Replication stress by the CHK1 inhibitor Prexasertib also recruits SLFN11 to nascent replicating DNA together with CDC45 and PCNA. We conclude that SLFN11 is recruited to stressed replication forks carrying extended RPA filaments where it blocks replication by changing chromatin structure across replication sites.

overall_design: Genome-wide mapping of chromatin accessibility was done by following the published method (Buenrostro et al., 2013). Briefly, fifty thousand CCRF-CEM parental and SLFN11-del cells were treated with DMSO or CPT (100 nM) for 2 and 4 hours. K562-Vector, -WT and –E669Q cells were treated with DMSO or CPT (1 μM) for 4 hours. To prepare nuclei, cells were lysed using cold lysis buffer (10 mM Tris-Cl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630). Immediately following the nuclei preparation, the pellet was re-suspended in the transposase reaction mix [25 μl 2x TD buffer, 2.5 μl Transposase (Illumina) and 22.5 μl of nuclease free water], and incubated for 30 minutes at 37 °C. Directly following transposition, the sample was purified using a DNA Clean and Concentrator-5 (ZYMO RESEARCH) and eluted with 25 μl of DNA elution buffer. Following purification, we amplified library fragments using 1x NEBnext PCR master mix and 1.25 μM of custom Nextera PCR primers 1 and 2, using the following PCR conditions: 72°C for 5 minutes, 98°C for 30 seconds, followed by thermo-cycling at 98°C for 10 seconds, 63°C for 30 seconds and 72°C for 1 minute. We amplified the full libraries for 5 cycles, after 5 cycles we took an aliquot of the PCR reaction and added 10 μl of the PCR cocktail with Syber Green at a final concentration of 0.6x. We ran this reaction for 20 cycles, to determine the additional number of cycles needed for the remaining 45 μL reaction. Libraries were amplified for a total of 10–12 cycles and purified using a PCRClean DX yielding a final library concentration of ~30 nM in 20 μl.

first: Yves

last: Pommier

first: Hongliang

last: Zhang


overall_design:
Nascent strand abundance assay (Nascent strand DNA-seq; NS-seq)
Replication origins were identified using the nascent-strand sequencing and abundance assay (Fu et al., 2014; Martin et al., 2011). In brief, twenty million of CCRF-CEM parental and SLFN11-del cells were treated with DMSO, CPT (100 nM), or CPT (100 nM) and VE-821 (2 μM) for four hours. DNA fractions (0.5-2 kb) were isolated using DNA fractionation on a 5-30% neutral sucrose gradient. 5’ single strand DNA ends were phosphorylated by T4 polynucleotide kinase (NEB) and then treated with lambda-exonuclease (NEB) to remove genomic DNA fragments that lacked the phosphorylated RNA primer. After RNase treatment, purified single stranded nascent DNA were random-primed using the Klenow and DNA Prime Labeling System (Invitrogen). Double-stranded nascent DNA (1 μg) was sequenced using the Illumina genome analyzer II (Solexa).
Assay for Transposase-Accessible Chromatin with high throughput sequencing (ATAC-seq)
Genome-wide mapping of chromatin accessibility was done by following the published method (Buenrostro et al., 2015). Briefly, fifty thousand CCRF-CEM parental and SLFN11-del cells were treated with DMSO or CPT (100 nM) for 2 and 4 hours. K562-Vector, -WT and –E669Q cells were treated with DMSO or CPT (1 μM) for 4 hours. To prepare nuclei, cells were lysed using cold lysis buffer (10 mM Tris-Cl, pH 7.4, 10 mM NaCl, 3 mM MgCl2 and 0.1% IGEPAL CA-630). Immediately following the nuclei preparation, the pellet was re-suspended in the transposase reaction mix [25 μl 2x TD buffer, 2.5 μl Transposase (Illumina) and 22.5 μl of nuclease free water], and incubated for 30 minutes at 37 °C. Directly following transposition, the sample was purified using a DNA Clean and Concentrator-5 (ZYMO RESEARCH) and eluted with 25 μl of DNA elution buffer. Following purification, we amplified library fragments using 1x NEBnext PCR master mix and 1.25 μM of custom Nextera PCR primers 1 and 2, using the following PCR conditions: 72°C for 5 minutes, 98°C for 30 seconds, followed by thermo-cycling at 98°C for 10 seconds, 63°C for 30 seconds and 72°C for 1 minute. We amplified the full libraries for 5 cycles, after 5 cycles we took an aliquot of the PCR reaction and added 10 μl of the PCR cocktail with Syber Green at a final concentration of 0.6x. We ran this reaction for 20 cycles, to determine the additional number of cycles needed for the remaining 45 μL reaction. Libraries were amplified for a total of 10–12 cycles and purified using a PCRClean DX yielding a final library concentration of ~30 nM in 20 μl.


title: AICDA-induced epigenetic plasticity accelerates germinal center-derived lymphomagenesis

city: New York

first: Matt

last: Teater

state: NY

address: 445 E 69th St

institute: Weill Cornell Medical College

type: Genome variation profiling by high throughput sequencing

summary: Epigenetic heterogeneity is emerging as a significant phenotypic feature of tumors. In diffuse large B-cell lymphomas (DLBCLs), increased cytosine methylation heterogeneity is associated with poor clinical outcome, yet the biological mechanisms driving epigenetic heterogeneity remain unclear. Activation-induced cytidine deaminase (AICDA), an enzyme that deaminates and facilitates demethylation of DNA methyl-cytosines in germinal center (GC) B-cells, is required for DLBCL pathogenesis and linked to inferior clinical outcomes. Here, we show that overexpression of AICDA causes more aggressive disease and worse outcome in BCL2-driven murine lymphomas. This phenotype was associated with significantly increased focal inter-tumor and intra-tumor cytosine methylation heterogeneity as compared to control lymphomas, but not with increased mutational burden. AICDA-mediated epigenetic heterogeneity was accompanied by DNA hypomethylation. Reciprocally, we observed that the focal inter-individual and intra-individual cytosine methylation heterogeneity characteristic of normal GC B-cells was lost upon depletion of AICDA. There was significant overlap of AICDA-induced methylation heterogeneity foci in GC B-cells and murine lymphomas. These observations are relevant to human patients, since DLBCLs with high AICDA expression likewise manifest extensive increases in focal inter-patient and intra-patient heterogeneity as compared to DLBCLs with low AICDA expression. Affected regions significantly overlapped with those found in murine AICDA-overexpressing lymphomas. Our results identify AICDA as a novel source of epigenetic plasticity and heterogeneity in B-cell lymphomas with potential significance for other tumors that express cytosine deaminases.

overall_design:
Enhanced reduced representation bisulfite sequencing (ERRBS), RNA-seq, and exome sequencing on murine VavP-Bcl2 and VavP-Bcl2+Aicda tumors.
Enhanced reduced representation bisulfite sequencing (ERRBS), RNA-seq and exome sequencing on murine VavP-Bcl2 and VavP-Bcl2+Aicda tumors. Paired ERRBS and RNA-seq on low AICDA-expressing and high AICDA-expression DLBCL.

first: Olivier

last: Elemento

first: Ari

last: Melnick


title: The rate of ageing-associated DNA methylation dynamics is a molecular readout of lifespan variation amongst mammalian species [Bis-PCR: naked mole-rat]

summary: We used liver samples from Naked Mole Rat to determine rate of change of methylation with respect to age.

overall_design: Bis-PCR profiling of Naked Mole Rats of various age.


title: Global gene expression in response to X rays in mouse models of DNA repair deficiency

first: Sally

last: Amundson

department: Center for Radiological Research

institute: Columbia University

summary:
There is a perceived need to develop tools for high-throughput radiation biodosimetry.  In the case of an improvised nuclear device or a dirty bomb in a major city, potentially hundreds of thousands of people would have to be screened to assign exposed individuals to appropriate treatment.  Gene expression represents an emerging approach to biodosimetry and could potentially provide an estimate of both absorbed dose and individual radiation-induced injury.  Mice have been used as an in vivo model to elucidate the radiation response and to construct biodosimetric gene expression profiles.  However, while most studies have used mouse strains with “average” radiation sensitivity, 2-4% of humans are thought to be radiosensitive.  Such individuals would suffer greater radiological injury at a given dose than members of the general population, so it is of interest to determine how such sensitivity may impact the biodosimetric gene expression signatures being developed.
Here, we use wild-type mice and genetically engineered mouse models deficient in two DNA repair pathways that can contribute to radiation sensitivity.  Humans with ataxia telangiectasia or Atm-/- mice are deficient in double-strand break repair and show increased radiosensitivity.  Atm-/- mice have a LD50/30 of 4 Gy compared with 8 Gy in the parental wild-type (WT) C57BL/6 mice.  Humans with SCID, or Prkdcscid mice are deficient in non-homologous end joining, and are also radiosensitive.  SCID mice have a LD50/30 of 3 Gy.  These animal models represent a “worse-case” scenario, and should be viewed as an extreme of radiation sensitivity.  Most radiosensitive members of the population will not have full-blown DNA repair deficiency syndromes, but these extreme models should help to define the maximum impact of DNA repair deficiency on gene expression following radiation exposure.

overall_design: Three genotypes (wild-type, ATM-/-, and Scid) were either exposed to a LD50/30 dose of x rays, or were sham-irradiated as controls, and then sacrificed 24h later.  5 replicates, representing individual mice, were used for each condition, with the exception of un-exposed (0 Gy) wild types and irradiated Atm-/- (4 Gy), for which six replicate mice were used.

first: Suresh

last: Kumar

first: Nils

last: Rudqvist

first: Evagelia

last: Laiakis

first: Albert

last: Fornace Jr


title: CNV analysis of 140 Filipinos with Cleft Lip and/or Palate

city: Iowa City

first: J.

middle: Robert

last: Manak

state: Iowa

address: 455 Biology Building, 129 East Jefferson Street

department: Biology and Pediatrics

institute: The University of Iowa

type: Genome variation profiling by genome tiling array

summary: Orofacial clefts are one of the most common birth defects, affecting 1-2 per 1000 births, and have a complex etiology. High-resolution array-based comparative genomic hybridization has increased the ability to detect copy number variants that can be causative for complex diseases such as cleft lip and/or palate. Utilizing this technique on 97 non-syndromic cleft lip and palate cases and 43 cases with cleft palate only, we identified a heterozygous deletion of Isthmin 1 in one affected case, as well as a deletion in a second case which removes putative 3' regulatory information. Isthmin 1 is a strong candidate for clefting as it is expressed in orofacial structures derived from the first branchial arch and is also in the same synexpression group as fibroblast growth factor 8 and sprouty RTK signaling antagonist 1a and 2, all of which have been associated with clefting.  Copy number variants affecting Isthmin 1 are exceedingly rare in control populations, and Isthmin 1 scores as a likely haploinsufficiency locus.  Confirming its role in craniofacial development, knockdown or CRISPR/Cas9-generated mutation of isthmin 1 in Xenopus laevis resulted in mild to severe craniofacial dysmorphologies, with several individuals presenting with median clefts. Moreover, knockdown of isthmin 1 produced decreased expression of LIM homeobox 8, itself a gene associated with clefting, in regions of the face that pattern the maxilla.  Our study demonstrates a successful pipeline from copy number variant identification of a candidate gene to functional validation in a vertebrate model system and reveals Isthmin 1 as both a new human clefting locus as well as a key craniofacial patterning gene.

overall_design: array-based Comparative Genomic Hybridization (aCGH) of 140 Filipino probands with cleft lip and palate assessed for rare deletions

first: Lisa

last: Lansdon

first: J


title: Ultraconserved Enhancers Are Required for Normal Development

city: Vienna

first: Iros

last: Barozzi

address: Borschkegasse 8a

country: Austria

institute: Medical University Vienna

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


title: Expression profiling reveals differential expressed genes between bovine IVF and SCNT embryos

status: Public on Jan 16 2018

city: Yangling

first: Xin

last: Liu

state: Shaanxi

address: 3 Taicheng Road

department: College of Veterinary Medicine

institute: Northwest A&F University

summary: Global hypermethylations of histone H3 lysine 9 (H3K9) tri- and di-methylation (H3K9me3/2) have been identified in bovine cloned embryos at the embryonic genome activation (EGA) stage (eight-cell stage), but the intrinsic reason for these anomalies remains elusive. To ascertain key factors responsible for aberrant H3K9 methylation, we performed RNA sequencing of transcripts in eight-cell bovine in vitro fertilized (IVF) and SCNT embryo. From the differentially expressed genes (DEGs) between IVF and SCNT embryos, we identified that the unsuccessful reactivation of two histone demethylases, KDM4D and KDM4E, is responsible for the incomplete H3K9me3/2 demethylation in SCNT embryos at the EGA stage. By mRNA injection, ectopic expression of either KDM4D or KDM4E could erase H3K9me3/2 barriers, improve blastocyst formation, and elevate cloning efficiency of bovine SCNT. To examine the detailed genes responsive to KDM4E overexpression, we also performed RNA sequencing of bovine eight-cell SCNT embryos with KDM4E compensation and found an obvious restoration of global transcriptional patterns in SCNT embryos. Our study first provides the transcriptome data set of bovine IVF and SCNT embryos during EGA with or without KDM4E overexpression, which advance the understanding of incomplete nuclear reprogramming, and contribute to the practical implications for genetically modified livestock breeding using SCNT.

overall_design: RNA-Seq profiles from pools of 8 eight-embryos from bovine IVF (3 replicates) and SCNT with (2 replicates) or without (3 replicates) KDM4E overexpression were generated using Illumina HiSeq 4000. SCNT without KDM4E overexpression are defined as the control compared with the other two samples, respectively.

first: Zhang

last: Yong

first: Liu

last: Xin

first: Xing

last: Xupeng

first: Zhou

last: Chuan

first: Wang

last: Yizhi


title: Peroxisome proliferator activated receptor gamma controls mature brown adipocyte inducibility through glycerol kinase

city: Zuerich

first: Lennart

last: Opitz

address: Winterthurerstr. 190

institute: FGCZ Zuerich

summary: Peroxisome proliferator‑activated receptors (PPARs) have been suggested as the master regulators of adipose tissue formation, however their role in regulating brown fat functionality has not been resolved. To address this question, we generated inducible brown fat specific mouse models for PPARa, b/d and g, respectively. Interestingly, we found that both PPARa and b/d are dispensable for brown fat function. In contrast, we could show that ablation of PPARg in vitro as well in vivo led to a reduced thermogenic capacity accompanied by a loss of inducibility by β-adrenergic signaling, as well as a shift from oxidative fatty acid metabolism to glucose utilization. We identified glycerol kinase (Gyk) as a partial mediator of PPARgfunction, and could show that Gyk expression correlates with brown fat thermogenic capacity in human brown fat biopsies. Thus, Gyk might constitute the link between PPARg mediated regulation of brown fat function and activation by β-adrenergic signaling.

overall_design: We used quadriplicates per condition.

first: David

last: Lasar

first: Christian

last: Wolfrum


title: Cellular models of hepatogenesis using embryonic stem cells

city: Düsseldorf

first: Wasco

last: Wruck

address: Moorenstr. 5

institute: Universitätsklinikum Düsseldorf

summary: Induced pluripotent stem cells (iPSCs) and human embryonic stem cells (hESCs) can be differentiated into hepatocyte-like cells (HLCs) and thus provide a defined and renewable source for toxicology and regenerative medicine. Critical comparison of patient-specific iPSCs to the golden standard—hESCs will provide an assessment of their strengths and weaknesses. This dataset consists of data at the transcriptional level during the differentiation process of hESCs into HLCs including the definite endoderm (DE) and hepatic endoderm (HE) stages. Besides for the study of this in-vitro-hepatogenesis model, it can be used as reference for investigating the differences and similarities between iPSC and hESC cellular models during hepatogenesis.

overall_design: Total RNA obtained from embryonic stem cells in several differentiation stages to hepatocyte-like cells.

first: Justyna

last: Jozefczuk

first: James

last: Adjaye


title: SLFN11 blocks stressed replication forks independently of ATR (NS-Seq)

first: Junko

last: Murai

overall_design: Nascent strands were purified with the lambda exonuclease methods from CCRF-CEM parental and SLFN11-KO (knockout) cell lines, which were treated with DMSO,  100 nM camptothecine or 100 nM camptothecin plus 2 uM VE-821 (ATR inhibitor) for 4 hrs.

first: Yves

last: Pommier

first: Christophe

last: Redon


title: WOX11 recruits a histone H3K27me3 demethylase to promote gene expression during shoot development in rice

city: Wuhan

first: Saifeng

last: Cheng

state: Hubei

address: No.1,Shizishan Street

department: National Key Laboratory of Crop Genetic Improvement

institute: Huazhong Agricultural University

summary: WUSCHEL-related homeobox (WOX) genes are key regulators of meristem activity and plant development, the chromatin mechanism of which to reprogram gene expression remains unclear. Histone H3K27me3 is a chromatin mark of developmentally repressed genes. How the repressive mark is removed from specific genes during plant development is largely unknown. Here, we show that WOX11 interacts with the H3K27me3 demethylase JMJ705 to activate gene expression during shoot development in rice. Genetic analysis indicates that WOX11 and JMJ705 cooperatively control shoot growth and commonly regulate the expression of a set of genes involved in meristem identity, chloroplast biogenesis, and energy metabolism in the shoot apex. Loss of WOX11 led to increased H3K27me3 and overexpression of JMJ705 decreased the methylation levels at a subset of common targets. JMJ705 is associated with most of the WOX11-binding sites found in the tested common targets in vivo, regardless of presence or absence of the JMJ705-binding motif. Furthermore, wox11 mutation reduced JMJ705-binding to many targets genome-wide. The results suggest that recruitment of JMJ705 to specific developmental pathway genes is promoted by DNA-binding transcription factors and that WOX11 functions to stimulate shoot growth through epigenetic reprogramming of genes involved in meristem development and energy-generating pathways.

overall_design:
Shoot apex were taken from 5-day-old wild type (HY), wox11 mutant and jmj705 mutant. mRNA profiles of each genotype with two biologicol repeats  were performed using Illumina HiSeq 2000.

ChIP-seq was performed in 5-day-old rice shoot apex isolated from JMJ705 over-expressing lines in wild type or wox11 mutant background with anti-FLAG antibody to detect the JMJ705 enrichment genome-wide.

first: S

first: Y

last: Lu

first: W

last: Yuan


title: The rate of ageing-associated DNA methylation dynamics is a molecular readout of lifespan variation amongst mammalian species [TC1_450k]

first: Robert

last: Lowe

summary: We used liver samples from TC1 Mice to determine rate of change of methylation with respect to age.

overall_design: Illumina 450K profiling of TC1 Mouse at various age.


title: Transcriptome of Melampsora larici-populina sexual stages [JGI version 1 genome]

status: Public on Jan 15 2018

city: Champenoux

first: Sebastien

last: Duplessis

address: INRA Centre Grand Est - Nancy

department: UMR 1136 IAM

institute: INRA

summary: Illumina HiSeq2500 technology was used to generate mRNA-sequencing expression profiles of  sexual stages of the poplar rust fungus M. larici-populina:   - Basidia (BSD) were collected from rust-infected dead poplar leaves;  - Pycnia (PYC) were collected from rust-infected larch needles one week after inoculation with basidiospores and corresponded to different haploid fungal cell types such as in planta infection hyphae, receptive hyphae and pycniospores;   - Aecia (AEC) were collected from rust-infected larch needles two weeks after inoculation, and corresponded to freshly produced aecia and dikaryotic aeciospores and remaining pycnia.  Paired-end reads of 100bp were generated for three sets of biological replicates corresponding to each of the targeted stage and aligned to the genome of M. larici-populina isolate 98AG31 (version 1.0; US Department of Energy Joint Genome Institute; http://genome.jgi.doe.gov/Mellp1/Mellp1.home.html; Duplessis et al. 2011a) using CLC Genomics Workbench 8.0.1

overall_design: In total, nine mRNA profiles of M. larici-populina sexual stages were generated by Illumina HiSeq2500 (2x100bp). Three biological replicates were sequenced for each stage BSD, PYC and AEC.

first: Cécile

last: Lorrain

first: Clémence

last: Marchal

first: Stéphane

last: Hacquard

first: Christine

last: Delaruelle

first: Jérémy

last: Petroswky

first: Benjamin

last: Petre

first: Arnaud

last: Hecker

first: Pascal

last: Frey

first: Sébastien


title: The rate of ageing-associated DNA methylation dynamics is a molecular readout of lifespan variation amongst mammalian species

sample_organism:
Macaca mulatta
Canis lupus familiaris
Mus musculus
Heterocephalus glaber

platform_organism:
Homo sapiens
Canis lupus familiaris
Heterocephalus glaber


title: A conserved RNA regulates miRNA turnover and animal behavior through a near-perfect miRNA site

status: Public on Jan 14 2018

city: Paris

first: Yuvia

middle: Alheli

last: Perez Rico

address: 26 rue d'Ulm

institute: Institut Curie

summary: Metazoan high complementarity microRNA (miRNA) binding sites can induce miRNA turnover through 3′ tailing and trimming, however the in vivo importance of this mechanism remains unknown. Here, we show that a transcript harboring a highly complementary and deeply conserved miRNA binding site for miR-29 controls zebrafish and mouse behavior. This brain-enriched transcript originated in distal vertebrates as a long noncoding RNA, which we called libra (lncRNA involved in behavioral alterations), and evolved to the protein-coding gene known as NREP in mammals where the miR-29 binding site is located within the 3′UTR. libra-deficiency results in viable fish that manifest abnormal exploration behavior. To determine if the Nrep transcript retained the regulatory noncoding function and to define the contribution of the highly complementary miR-29 binding site, we generated the NrepmiR-29scr allele in mice where the sequence of the site was scrambled to specifically uncouple Nrep function from miR-29 regulation. We show that the Nrep miR-29 binding site restricts miR-29b expression in the cerebellum. The ectopic miR-29b expression in NrepmiR-29scr mice is associated with altered cerebellum function and behavior. We finally show that the near-perfect miRNA site triggers miR-29b turnover through 3′ trimming. In summary, we demonstrate the first example of an endogenous RNA-directed miRNA degradation event in vivo that is required for normal animal behavior.

sample_organism:
Danio rerio
Mus musculus

platform_organism:
Danio rerio
Mus musculus

overall_design: Total RNA profiles of 72hpf embryos of zebrafish wild type (wt), full deletion of libra locus (libradel) and  inversion of the libra conserved region (librainv) in technical duplicate; Size-selected small RNA profiles of wild type and NrepmiR-29scr mouse NPCs (n=3 for both lines)

first: Alena

last: Shkumatava

first: Angelo

last: Bitetti

first: Allison

last: Mallory


title: Loss of CNFy toxin-induced  inflammation drives Yersinia pseudotuberculosis into persistency

city: Braunschweig

first: Michael

last: Beckstette

state: Lower Saxony

address: Inhoffenstrase 7

department: Molecular Infection Biology

institute: Helmholtz Centre for Infection Research

summary: Gastrointestinal infections caused by enteric yersiniae can become persistent and complicated by relapsing enteritis and severe autoimmune disorders. To establish a persistent infection, the bacteria have to cope with hostile surroundings when they transmigrate through the intestinal epithelium and colonize underlying gut-associated lymphatic tissues. How the bacteria gain a foothold in the face of host immune responses is poorly understood. Here, we show that the CNFY toxin, which enhances translocation of the antiphagocytic Yop effectors, induces IL-6-triggered inflammatory responses. This results in extensive tissue destruction, alteration of the intestinal microbiota and bacterial clearance. An elimination of CNFY, however, increases inter-feron-γ-mediated responses, comprising non-inflammatory antimicrobial activities and Ido1-promoted tolerogenesis. This process is accompanied by a preterm reprogram-ming of the pathogen's transcriptional response towards persistence, which gives the bacteria a fitness edge against host responses. These results have significant implica-tions for our understanding how enteric bacteria establish a commensal-type life style.

overall_design: RNA of Y. pseudotuberculosis infected and uninfected caecal tissue of Mus musculus

first: Wiebke

last: Heine

first: Petra

last: Dersch

first: Ann-Kathrin

last: Heroven


title: The transcriptional repressor Blimp-1/PRDM1 restrains Type III IFN signalling in mammary epithelial cells

status: Public on Jan 13 2018

city: Oxford

first: Arne

middle: Wyndham

last: Mould

state: Oxfordshire

address: Warneford Hospital

department: Psychiatry

institute: Oxford University

summary:
Expression profiling of wild-type and Prdm1 null mouse MECs using Illumina whole genome mouse V2 arrays.
The hypothesis tested was that Prdm1-regulated gene expression in MECs is required for correct epithelial function.

overall_design: Tamoxifen inducible Prdm1 null (Prdm1BEH/CA;ROSA26:CreERT2)  and wild type MEC were treated with 4OH Tamoxifen on D0 of cuture.  Total RNA was obtained from three wild type (C57BL6) and three Prdm1 null MECs each on D3 and D4 of culture and hybridized to Illumina WG6_V2 arrays

first: S

middle: K

last: Elias

first: E

last: Robertson

last: Bikoff

first: A


title: BCR heavy and light chain repertoire sequencing of pertitoneal cavity B-1a cells from wild-type and Igll1 knockout mice

first: Jane

last: Skok

address: 550 First Ave, Smillow 307

institute: NYU

summary: We report that B-1a cells develop in a surrogate light chain independent context. As a consequence, the precursor B-1a cell population avoids a pre-BCR positive selection stage. To confirm that the B-1a cells generated in this manner repersent a bonafide B-1a cell compartment, we did NGS on BCR rearrangements to assess the repertoire diversity. We find that as a whoile, B-1a cell repertoire that develop in Igll1 kncokout mice are similar compared to wild-type. This supports our findings that B-1a cells develop properly in the absence of surrogate light chain.

overall_design: Peritoneal cavity B-1a cells were sorted from 6-8 week old mice, DNA was extracted and rearranged heavy chains were amplified by PCR. These PCR products were then process to form DNA libraries that were compatible with Illumina NGS.

first: Jason

middle: B

last: Wong


title: ICG-001 activates a series of ER stress response genes in monocyte derived macrophages (MDMs)

status: Public on Jan 12 2018

first: Stephen

last: Taylor

address: Headington

institute: CBRG

summary: By screening a target-focused Wnt-sgnaling small compound library we identified the Wnt-modulator ICG-001 as inihibitory for activating the mitochondrial fission protein Drp1. To explore how ICG-001 might act we undertook gene expression profiling in iCG-001 treated primary macrophages.

overall_design: The experiment compares transcriptional profiles of unstimulates control cells, cells that were pretreated with 10 µM ICG-001 or vehicle control and then stimulated with 200 ng/ml L-18 MDP for 2 h. The experiment was performed in biological triplicates.

first: Heidi

last: Zinecker

first: Steve

first: Dilair

last: Baban

first: Alison

last: Simmons


title: Comparative cardiac induction time-courses using wild-type or genetically modified human embryonic stem cells

city: Münster

first: Boris

last: Greber

address: Von-Esmarch-Str. 54

institute: Max Planck Institute for Molecular Biomedicine

summary:
This study compares directed cardiac differentiation time-courses using (i) HuES6 cells with endogenous ISL1 knockout + inducible ISL1 transgene, and (ii) wild-type HuES6 cells.
For the first series, a subset of samples was left untreated (= ISL1 knockout scenario) while another subset received doxycycline treatment to upregulate ISL1 at days 3 and 4 of differentiation (ISL1 rescue). For the second series, a subset of samples was differentiated under default conditions (no RA) while another subset received retinoic acid (RA) on days 3 and 4 of differentiation, which leads to an atrial-like phenotype.

overall_design: Total RNA was isolated in a time course manner from the distinct series of differentiating samples (on days 0, 1, 2, 3, 4, 5, 6, and 8). Note that the two subsets of each series are only diverging from day 4 onwards.

first: Roberto

last: Quaranta


title: Accurate H3K27 methylation patterns can be established de novo by SUZ12-directed PRC2

city: Copenhagen N

first: Jonas

middle: Westergaard

last: Hojfeldt

address: Ole Maaloes Vej 5

department: BRIC

country: Denmark

institute: University of Copenhagen

summary: The Polycomb repressive complex 2 (PRC2) catalyzes H3K27 methylation and is required for maintaining transcriptional patterns and cellular identity, but the specification and maintenance of genomic PRC2 binding and H3K27 methylation patterns remain incompletely understood. Epigenetic mechanisms have been proposed, wherein pre-existing H3K27 methylation directs recruitment and regulates the catalytic activity of PRC2 to support its own maintenance. Here we investigate if such mechanisms are required for specifying H3K27 methylation patterns in mouse embryonic stem cells. Through genetic knockouts of subunits, we find that PRC2 is responsible for all levels of H3K27 methylation, and that methylation patterns can be accurately established de novo, when a subunit is reintroduced in the absence of pre-existing H3K27 methylation and PRC2 chromatin binding. Furthermore, we find that the core-subunit SUZ12 directs genomic binding of PRC2, which is essential for establishing correct methylation patterns.

overall_design:
H3K27me1, H3K27me2, H3K27me3 enrichment profiles (ChIPseq) analyzed in mouse ES cells which are wildtype, knockout for Ezh1 and Ezh2 (Ezh1/Ezh2 dKO), or re-expressing wildtype EZH2 (10 samples including 1 ChIP input sample for Ezh1/Ezh2 dKO).
H3K27me1, H3K27me2, H3K27me3 enrichment profiles (ChIPseq) analyzed in mouse ES cells which are wildtype, knockout for Suz12, or re-expressing either wildtype SUZ12 or one of two SUZ12 fragment (15 samples).
In addition, H3K27me1, H3K27me2, H3K27me3 enrichment profiles (ChIPseq) analyzed in E14 mouse ES cells treated with Ezh2 inhibitor and followed in a timecourse study after inhibitor washout (24 samples).
PRC2 subunit enrichment profiles through Suz12, Ezh2 and Flag ChIPseq in various PRC2 wildtype, knockout or mutant ES cells (9 Suz12 ChIP samples, 2 Flag ChIP samples, 2 Ezh2 ChIP samples).
Differential expression analysis (RNAseq) of mouse ES cells which are wildtype (3 replicates), knockout for Ezh1 and Ezh2 (Ezh1/Ezh2 dKO) (3 replicates), or re-expressing wildtype EZH2 (3 replicates) (9 samples).

middle: W

first: Anne

last: Laugesen

first: Berthe

last: Willumsen

first: Helene

last: Damhofer

first: Lin

last: Hedehus

first: Kristian

last: Helin