title: Effects of NIK activation on the self-renewal of hematopoietic stem cells

status: Public on Oct 19 2017

city: Iowa City

first: Thomas

middle: B

last: Bair

state: IA

address: 335 EMRB

institute: University of Iowa

summary: Comparison of transcriptome between control and NIK-activated hematopoietic stem cells

overall_design: Mutant NIK, which can not be targeted by TRAF3, was activated in hematopoietic system by crossing Rosa-loxp-STOP-loxp-NIK mice with Vav-iCre mice (caNIK mice). Hematopoietic stem/progenitor cells from control or caNIK mice were sorted by FACS and used for RNAseq analysis.

first: Chen

last: Zhao


title: A viral protein promotes host SAMS1 activity and ethylene production for the benefit of virus infection

status: Public on Oct 20 2017

city: Beijing

first: Shanshan

address: No.5, Yiheyuan Road, Haidian District

country: China

institute: Peking University

type: Expression profiling by high throughput sequencing

summary: Ethylene plays critical roles in plant development and biotic stress response.  However, the mechanism of ethylene in host antiviral response remains unclear. Rice dwarf virus (RDV) triggers the ethylene production resulting in host susceptibility by enhancing the activity of S-adenosyl-L-methionine synthase (SAMS) in ethylene synthesis pathway. Our molecular and genetic data demonstrated that Pns11 specifically interacted with OsSAMS1 to enhance its enzymatic activity, resulting in increased production of ethylene in both Pns11 overexpressing and RDV-infected rice. Treatment of rice plants with ethylene precursor increased their susceptibility to RDV infection. More importantly, knockout of OsSAMS1 (CRISPR/Cas9) or blocking ethylene signaling in an ethylene-insensitive mutant (osein2) decreased susceptibility to RDV infection, and overexpression of OsSAMS1 or Pns11 increased susceptibility. In this study, we have analyzed the relationship between ethylene pathway response and RDV infection or OsSAMS1 and S11 transgene by the gene expression analysis through the RNA-seq.

overall_design: Comparison between Mock and RDV infected rice, 3 control, 3 RDV-infected, Comparison between WT and OsSAMS1 and S11 transgenic rice , 2 control, 3 OsSAMS1 OX, 3 Ossams1 KO, 3 S11 OX, independently grown and harvested, 1 sample derived from 3 plants grown under same conditions.

first: Wei

last: Hong

first: Jianguo

last: Wu

first: Yu

last: Wang

first: Chunhong

last: Wei

first: Yi

last: Li


title: Runx1 Circular Chromosome Conformation Capture Sequencing in Mouse Hematopoietic Progenitor Cells

status: Public on Oct 22 2017

city: Dunedin

first: Judith

last: Marsman

state: Otago

address: 201 Great King Street

department: Surgical Sciences

country: New Zealand

institute: University of Otago

summary: We have performed circular chromosome conformation capture sequencing (4C-seq) with six baits located at the Runx1 locus in the mouse hematopoietic progenitor cell line HPC-7. 4C baits were designed to the P1 and P2 promoters and to the previously characterised +24 hematopoietic enhancer (P1, P2 and 24), with secondary baits located at nearby cohesin/CTCF (cc) binding sites (P1cc, 24cc, P2cc).

overall_design: For each 4C-seq bait, two replicates and one control library (in which the first ligation step was omitted) were sequenced.

first: Amarni

last: Thomas

first: Motomi

last: Osato

first: Justin

middle: M

last: O'Sullivan

first: Julia

middle: A

last: Horsfield


title: A novel role for the EWS portion of EWS/FLI in binding GGAA-microsatellites required for oncogenic transformation in Ewing sarcoma [ChIP-Seq]

city: Columbus

first: Kirsten

last: Johnson

state: Ohio

address: The Research Institute, WA5110, 700 Childrens Drive

department: Childhood Cancer and Blood Disorders

country: USA

institute: Nationwide Childrens Hospital

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Ewing sarcoma usually expresses the EWS/FLI fusion transcription factor oncoprotein. EWS/FLI regulates myriad genes required for Ewing sarcoma development.  EWS/FLI binds GGAA-microsatellite sequences in vivo and in vitro, and these sequences provide EWS/FLI-mediated activation to reporter constructs, suggesting that they function as EWS/FLI-response elements.  Genomic GGAA-microsatellites are highly variable and polymorphic.  Current data suggest that there is an optimal “sweet-spot” GGAA-microsatellite length (of 18-26 GGAA repeats) that confers maximal EWS/FLI-responsiveness to target genes, but the mechanistic basis for this was not known.  We now demonstrate the absolute necessity of an EWS/FLI-bound GGAA-microsatellite in regulation of the NR0B1 gene, as well as for Ewing sarcoma proliferation and oncogenic transformation.  Biochemical studies, using recombinant Δ22 (a version of EWS/FLI containing only the FLI portion) demonstrated a stoichiometry of one Δ22-monomer binding to every two consecutive GGAA-repeats on shorter GGAA-microsatellite sequences.  Surprisingly, the affinity for Δ22 binding to GGAA-microsatellites significantly decreased, and ultimately became unmeasureable, when the size of the GGAA-microsatellite was increased to the “sweet-spot” length.  In contrast, a fully-functional EWS/FLI mutant (Mut9) that retains approximately half of the EWS portion of the fusion showed low affinity for smaller GGAA-microsatellites, but instead significantly increased its affinity at “sweet-spot” microsatellite lengths.  Single-gene ChIP and genome-wide ChIP-seq and RNA-seq studies extended these findings to the in vivo setting.  Taken together, these data demonstrate the absolute requirement of GGAA-microsatellites as EWS/FLI activating response elements in vivo and reveal an unsuspected novel role for the EWS portion of the EWS/FLI fusion in binding to optimal-length GGAA-microsatellites.

overall_design: Examination of EWS/FLI binding in A673 cells subjected to various treatments

first: Stephen

middle: L

last: Lessnick


title: Short Poly(A) Tails are a Conserved Feature of Highly Expressed Genes

city: La Jolla

first: Amy

last: Pasquinelli

state: CA

address: 9500 Gilman Drive

department: Biological Sciences

institute: University of California, San Diego

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

overall_design: Refer to individual Series


title: Transcriptome analysis reveals determinant stages controlling human embryonic stem cell commitment to neuronal cells

status: Public on Oct 18 2017

city: Shanghai

first: yuanyuan

last: li

state: Shanghai

address: Yue yang 320

institute: sibcb

summary: Proper neural commitment is essential for ensuring the appropriate development of the human brain and for preventing neurodevelopmental diseases such as autism spectrum disorders, schizophrenia, and intellectual disorders. However, the molecular mechanisms underlying the neural commitment in humans remain elusive. Here, we report the establishment of a neural differentiation system based on human embryonic stem cells (hESCs) and on comprehensive RNA-Seq analysis of transcriptome dynamics during early hESC differentiation. Using weighted gene co-expression network analysis, we reveal that the hESC neurodevelopmental trajectory has five stages: pluripotency (day 0), differentiation initiation (days 2, 4, and 6), neural commitment (days 8–10), neural progenitor cell proliferation (days 12, 14, and 16), and neuronal differentiation (days 18, 20, and 22). These stages were characterized by unique module genes, which may recapitulate the early human cortical development. Moreover, a comparison of our RNA-Seq data with several other transcriptome profiling datasets from mice and humans indicated that module 3 associated with the day 8–10 stage is a critical window of fate switch from the pluripotency to the neural lineage. Interestingly, at this stage, no key extrinsic signals were activated. In contrast, using CRISPR-Cas9–mediated gene knockouts, we also found that intrinsic hub transcription factors, including the schizophrenia-associated SIX3 gene and septo-optic dysplasia–related HESX1 gene, are required to program hESC neural determination. Our results improve the understanding of the mechanism of neural commitment in the human brain and may help elucidate the etiology of human mental disorders and advance therapies for managing these conditions.

overall_design: RNA-sequencing of twelve samples prepared every other day between hESC neuronal differentiation day 0 to day 22 .

first: Yuanyuan

first: Ran

first: Guangdun

last: Peng

first: Naihe

last: Jing


title: RNA-Seq of L4 C. elegans

summary: RNA-seq libraries were generated for L4 wildtype worms to analyze RNA abundance at this point in development.

overall_design: Three independent replicates of wildtype C. elegans were collected at the L4 stage for RNA.

first: Sarah

last: Azoubel Lima

first: Laura

last: Chipman

first: Angela

last: Nicholson


title: KDM2B is a Histone H3K79 Demethylase and Induces Transcriptional Repression via SIRT1-mediated Chromatin Silencing

status: Public on Oct 21 2017

city: Seoul

first: Joo-Young

last: Kang

address: Heukseok-dong, Dongjak-gu

country: South Korea

institute: Chung-Ang University

summary: The methylation of histone H3 lysine 79 (H3K79) acts as an active chromatin marker and is enriched in actively transcribed regions of the genome. However, demethylase of this mark remains unknown despite intensive research. Here, we show that KDM2B (FBXL10), a member of the Jumonji C (JmjC) family of proteins and previously known for its histone H3K36 demethylase activity, is a di- and trimethyl H3K79 demethylase. We demonstrate that KDM2B induces transcriptional repression of HOXA7 and MEIS1 via occupancy of promoters and demethylation of H3K79. Furthermore, genome-wide analysis suggests that H3K79 methylation levels are increased when KDM2B is depleted, suggesting that KDM2B functions as an H3K79 demethylase in vivo. Finally, stable KDM2B-knockdown cell lines exhibited displacement of SIRT1, with concomitant increases in H3K79 methylation and H4K16 acetylation. Our findings identify KDM2B as an H3K79 demethylase and link its function to transcriptional repression via SIRT1-mediated chromatin silencing.

overall_design: We profiled genomic occupancy of H3K79me2 and H3K79me3 via ChIP-seq for KDM2B KD and control in human 293T stable cell lines.

first: Joo

middle: Y

first: Sang

last: Seo


title: Degradome sequencing of Arabidopsis ago1-27 mutant

status: Public on Oct 23 2017

city: Tianjin

first: Xuan

last: Ma

address: Binshui West Road

department: College of Life Sciences

institute: Tianjin Normal University

type: Non-coding RNA profiling by high throughput sequencing

summary: The goal of this study is to globally identify miRNA/siRNA cleavage sites on mRNAs in Arabidopsis wild type and ago1-27 mutant.

overall_design: Degradome-seq was performed using Arabidopsis wild type and ago1-27 mutant seedlings. Three biological replicates were conducted for each genotype.

first: Jie

last: Cui

first: Xuemei

last: Chen


title: Post-transcriptional Gene Silencing Mediated by microRNAs is Controlled by Nucleoplasmic Sfpq [RIP-Seq]

city: Nice

first: Silvia

last: Bottini

address: 299, chemin de la ginestiere

country: France

summary: There is a growing body of evidence about the presence and the activity of the miRISC in the nucleus of mammalian cells. Here, we show by quantitative proteomic analysis that Ago2 interacts with nucleoplasmic Sfpq in a RNA-dependent fashion. By HITS-CLIP and transcriptomic analyses, we demonstrated that Sfpq directly controls the miRNA targeting of a subset of crucial miRNA-target mRNAs when it binds locally. Sfpq modulates miRNA targeting in both nucleoplasm and cytoplasm, indicating a nucleoplasmic imprinting of Sfpq-target mRNAs that influence miRNA targeting in both cellular compartments. Mechanistically, Sfpq binds to a sizeable set of long 3’UTR forming long aggregates to optimize miRNA position/recruitment to selected binding sites, as we show for Lin28A mRNA. These results extend the miRNA-mediated post-transcriptional gene silencing into the nucleoplasm and indicate that an unique Sfpq-dependent post-transcriptional strategy for controlling both nuclear and cytoplasmic gene expression takes place in cells during physio-pathological events.

overall_design: Ago2 and SFPQ RIP followed by small-RNA-seq. All experiments were performed in P19 cell and in triplicates.

first: Nedra

last: Hamouda-Tekaya

first: Raphael

last: Mategot

first: Laure-Emmanuelle

last: Zaragosi

first: Stephane

last: Audebert

first: Valerie

last: Grandjean

first: Claire

last: Mauduit

first: Mohamed

last: Benahmed

first: Pascal

last: Barbry

first: Emanuela

last: Repetto

first: Michele

last: Trabucchi


title: Age-dependent human β cell proliferation induced by GLP-1 and calcineurin signaling

city: HUNTSVILLE

first: NRIPESH

last: PRASAD

state: AL

address: 601 GENOME WAY

department: GENOMIC SERVICES LABORATORY

institute: HUDSONALPHA INSTITUTE FOR BIOTECHNOLOGY

summary: We developed an engraftment strategy to examine age-associated human islet cell replication and found that Ex-4, an agonist of GLP-1R, stimulates human β cell proliferation in juvenile, but not adult islets. We showed that this β cell proliferative response to Ex-4 requires Cn/NFAT signaling.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: We transplanted human islets under renal capsule of NSG mice and removed the human islet grafts 4 weeks after Ex-4 or PBS treatment.  The total RNAs were extracted from the grafts and used for RNAseq.

first: Alvin

middle: C

last: Powers

first: Chunhua

last: Dai


title: Excretory/secretory microRNAs from the gastrointestinal nematode Heligmosomoides polygyrus bakeri in culture.

city: Sainte-Anne-de-Bellevue

first: Lucienne

last: Tritten

state: QC

address: 21111 Lakeshore Road

department: Institute of Parasitology

country: Canada

institute: McGill University

summary: We report analyses of the types and amounts of microRNAs found in culture media recovered after incubating approximately 3300 H. p. bakeri adult worms for 48 hours.

sample_organism:
Mus musculus
Heligmosomoides polygyrus bakeri

overall_design: Examination of excretory/secretory miRNAs of the mouse gastrointestinal nematode parasite H. p. bakeri released in culture media.

first: L

first: T

middle: G

last: Geary


title: Comparative analysis of miRNA expression during the development of insects of different metamorphosis modes and germ-band types

city: Barcelona

first: Guillem

last: Ylla

state: Catalonia

address: Passeig Marítim de la Barceloneta, 37

country: Spain

institute: Institute of Evolutionary Biology

summary:
Do miRNAs contribute to specify the germ-band type and the body structure in the insect embryo?  Our goal was to address that issue by studying the changes in miRNA expression along the ontogeny of the German cockroach Blattella germanica, which is a short germ-band and hemimetabolan species.

 We sequenced small RNA libraries representing 11 developmental stages of B.  germanica ontogeny (with especial emphasis on embryogenesis) and the changes in miRNA expression were examined. Data were compared with equivalent data for two long germ-band holometabolan species Drosophila melanogaster and Drosophila virilis, and the short germ-band holometabolan species Tribolium castaneum. The identification of B. germanica embryo small RNA sequences unveiled miRNAs not detected in previous studies, such as those of the MIR-309 family and 54 novel miRNAs. Four main waves of miRNA expression were recognized (with most miRNA changes occurring during the embryonic stages): the first from day 0 to day 1 of embryogenesis, the second during mid-embryogenesis (days 0-6), the third (with an acute expression peak) on day 2 of embryonic development, and the fourth during post-embryonic development. The second wave defined the boundaries of maternal-to-zygotic transition, with maternal mRNAs being cleared - presumably by Mir-309 and associated scavenger miRNAs.

 miRNAs follow well-defined patterns of expression over hemimetabolan ontogeny - patterns that are more diverse during embryonic development than during the nymphal stages. The results suggest that miRNAs play important roles in the developmental transitions between the embryonic stages of development (starting with maternal loading), during which they might influence the germ-band type and metamorphosis mode.

overall_design: Eleven developmental stages were selected for analysis. For each stage, two small RNA libraries were prepared from a pool of specimens using the Illumina NEBNext small RNA libraries kit.

first: Xavier

last: Belles

first: M

middle: D

last: Piulachs


title: Daily oscillation and enrichment patterns in the transcriptome of the behavior-manipulating parasite Ophiocordyceps kimflemingiae

city: Orlando

first: Charissa

last: de Bekker

state: FL

address: UCF Biology Dept, 4110 Libra Dr

department: Biology

institute: UCF

summary: The natural histories of various parasite-host interactions that involve adaptive manipulation of host behavior describe a time-of-day synchronization of certain events. One example is the manipulated biting behavior observed in Carpenter ants infected with Ophiocordyceps unilateralis sensu lato. Field observations in Thailand demonstrated that manipulated biting takes place around solar noon. Laboratory work with species from South Carolina, demonstrated a shift of this synchronized biting event to the early morning. We therefore hypothesize that biological clocks play an important role in these and other parasite-host interactions. To set the stage for our future research into this, we, here, aimed to explore if fungal species of the complex O. unilateralis s.l. have a functional molecular clock. We, therefore, performed RNA-Seq on time course samples of the recently sequenced species Ophiocordyceps kimflemingiae. Fungal blastospores grown in liquid media were harvested under cycling light-dark and continuous darkness conditions. Rhythmic mRNA levels were assessed with JTK_CYCLE. We found that O. kimflemingiae is not hugely rhythmic under our sampling conditions, but rhythmicity can be observed. Moreover, we identified functional homologs of known clock genes. In addition, our data indicated that regulation of transcription appears to take place during the light phase (day time). In contrast, the expression levels of a significant number of secreted enzymes, proteases, toxins and small bioactive compounds peaked during the dark phase (night time). This suggests that this fungal parasite might indeed rely on biological clocks for the precise daily timing of certain events and additionally validates further research into this.

overall_design: Samples were taken of O. kimflemingiae cultures at 4h intervals both under light cycling (LD) and constant darkness (DD) conditions. These samples were used for RNA-Seq and gene expression was analyzed for rhythmicity with a circa 24h period. For each sampled time point one replicate was generated. Part of the data was verified with RT-QPCR.

first: Ian

last: Will

first: Andreas

last: Brachmann

first: Martha

last: Merrow


title: The Effect of Copper and Cerium on the Transcriptome of Methylosinus trichosporium OB3b

city: Ann Arbor

first: Wenyu

last: Gu

state: MI

address: 2350 Hayward St, EWRE 283

institute: University of Michigan

summary: Using RNA-Seq analysis we compared the transcriptome of Methylosinus trichosporium OB3b grown in the presence of varying amounts of copper and cerium.  When copper was added in the absence of cerium, expression of genes encoding for both soluble and particulate methane monooxygenases varied as expected.  Genes encoding for copper uptake, storage, and efflux also increased, indicating that methanotrophs must carefully control copper homeostasis.  When cerium was added in the absence of copper, expression of genes encoding for alternative methanol dehydrogenases varied as expected, but few other genes were found have differential expression.  When cerium concentrations were varied in the presence of copper, few genes were found to be either up or downregulated, indicating that copper over rules any regulation by cerium.  When copper was added in the presence of cerium, however, many genes were upregulated, most notably multiple steps of the central methane oxidation pathway, the serine cycle, and the ethylmalonyl-CoA pathway.  Many genes were also downregulated, including those encoding for nitrogenase and hydrogenase.

overall_design: Examination of transcriptome of Methylosinus trichosporium OB3b when growing at presence or absence of copper and cerium.

first: Jeremy

last: Semrau


title: CD117, CD200 and CD371 identifies seven populations of γδ thymocytes programmed towards three distinct effector subsets in adult mice

city: Copenhagen

first: Terkild

middle: Brink

last: Buus

address: Blegdamsvej 3

department: ISIM

country: Denmark

institute: University of Copenhagen

summary: Murine γδ T cells include several effector subsets that fulfil distinct functional roles. In pathological conditions, such as cancer, some γδ T cell subsets are highly protective whereas other subsets may exacerbate disease. γδ T cells are programmed for their effector function during their development in the thymus. Investigations of γδ T cell development have been hampered by the scarcity of surface markers distinguishing different development stages. In this study, we found that addition of CD117, CD200 and CD371 to existing markers, allowed identification of seven distinct development stages (named A, B, C, D, E, F and G) present in both the Vγ1.1+ and Vγ2+ thymocyte subsets. We provide evidence for the existence of three distinct pathways leading to export of γδ T cells, two of which express high levels of CD24. These pathways are dominated by different TCRδ repertoires shaped by TCR signalling or lack thereof. Each pathway express distinct cytokine and transcription factor profiles associated with γδT1, γδNKT and adaptive γδ T cells. Thus, the identification of additional γδ T cell development stages in this study connects three distinct development pathways to the programming of three γδ T cell effector subsets.

overall_design: FACS sorted TCRVγ1.1+ and TCRVγ2+ γδ thymocytes each divided into 7 populations based on expression of CD24, CD25, CD73, CD117, CD200 and CD371 pooled from 3x8 (rep1) or 2x8 (rep2) mice. Two biological replicates (rep1 and rep2).

first: Niels

last: Ødum

first: Carsten

last: Geisler

first: Jens

middle: P

last: Lauritsen


title: Activating Mutation in STAT5 Is Sufficient To Drive PTCL peripheral T-Cell Lymphoma (PTCL)

city: Vienna

first: Richard

last: Moriggl

address: Waehringer Strasse 13A

country: Austria

institute: Ludwig Boltzmann Institute for Cancer Research LBI-CR

summary: Recurrent gain-of-function mutations in the transcription factor STAT5 have been detected in PTCL, an aggressive, heterogeneous disease, for which currently no targeted therapy exists. Here we investigated whether constitutive activation of STAT5 suffices to drive PTCL and whether inhibition of the JAK/STAT pathway offers a novel therapeutic opportunity in this disease. We found pronounced STAT5 expression and activity in patients from different PTCL subsets. To mimic high STAT5 activity we expressed a hyperactive STAT5A variant (termed vcS5) in the hematopoietic lineage in transgenic mice. vcS5-transgenic animals developed a lethal PTCL-like disease with full penetrance, characterized by massive expansion of CD8+ T-cells and destructive organ-infiltration. Adoptive transfer of vcS5-expressing CD8+ T-cell rapidly induced the disease in immunocompetent recipient mice. Neoplastic vcS5-T-cells displayed cytokine-hypersensitivity and showed activated, memory CD8+ T-lymphocyte characteristics. Histo-pathological analysis as well as mRNA expression profiles of vcS5 mice was closely correlated with distinct human lymphoma subtypes, including PTCL. Treatment of murine and human PTCL cell lines with the clinical JAK inhibitor Ruxolitinib or a selective STAT5 SH2 domain inhibitor induced cell death. Thus, our results demonstrate that enhanced STAT5 signaling drives PTCL development and suggest inhibition of the JAK/STAT5 pathway as a valuable therapeutic option for patients suffering from these aggressive lymphomas.

overall_design: Total RNA extracted from wt and vcS5 (transgenic) splenocytes (2 biological replicates/sample), RNA-seq 50 bp single-end libraries sequenced on HiSeq 2000 (Illumina), to compare gene expression profiles

first: Barbara

last: Maurer

first: Doris

last: Chen


title: Poly(A) Tail Length of L4 C. elegans

first: Amy

last: Pasquinelli

summary: Poly(A) tails are important elements in mRNA translation and stability. However, recent genome-wide studies concluded that poly(A) tail length was generally not associated with translational efficiency in non-embryonic cells. To investigate if poly(A) tail size might be coupled to gene expression in an intact organism, we used an adapted TAIL-seq protocol to measure poly(A) tails in Caenorhabditis elegans. Surprisingly, we found that well-expressed transcripts contain relatively short, well-defined tails. This attribute appears dependent on translational efficiency, as transcripts enriched for optimal codons and ribosome association had the shortest tail sizes, while non-coding RNAs retained long tails.

overall_design: Global investigation of L4 C. elegans poly(A) tail length by using mTAIL-seq and analysis by the tailseeker2 program. Two independent biological replicates were used.

first: Sarah

last: Azoubel Lima

first: Laura

last: Chipman

first: Angela

last: Nicholson


title: Post-transcriptional Gene Silencing Mediated by microRNAs is Controlled by Nucleoplasmic Sfpq [HITS-CLIP]

first: Silvia

last: Bottini

overall_design: Ago2 HITS-CLIP upon let-7a transfection, Ago2 HITS-CLIP upon SFPQ knockdown in control and upon let-7a transfection, SFPQ HITS-CLIP in control and upon SFPQ knockdown. All experiments were performed in P19 cell and in triplicates.

first: Nedra

last: Hamouda-Tekaya

first: Raphael

last: Mategot

first: Laure-Emmanuelle

last: Zaragosi

first: Stephane

last: Audebert

first: Valerie

last: Grandjean

first: Claire

last: Mauduit

first: Mohamed

last: Benahmed

first: Pascal

last: Barbry

first: Emanuela

last: Repetto

first: Michele

last: Trabucchi


title: Classifying Drosophila Olfactory Projection Neuron Subtypes by Single-cell RNA Sequencing

city: Stanford

first: Felix

last: Horns

address: Clark Center E300, Quake Lab

department: Bioengineering

institute: Stanford University

summary:
How a neuronal cell type is defined and how this relates to its transcriptome are still open
questions. The Drosophila olfactory projection neurons (PNs) are among the bestcharacterized
neuronal types: Different PN classes target dendrites to distinct olfactory
glomeruli and PNs of the same class exhibit indistinguishable anatomical and physiological
properties. Using single-cell RNA-sequencing, we comprehensively characterized the
transcriptomes of 40 PN classes and unequivocally identified transcriptomes for 6 classes.
We found a new lineage-specific transcription factor that instructs PN dendrite targeting.
Transcriptomes of closely-related PN classes exhibit the largest difference during circuit
assembly, but become indistinguishable in adults, suggesting that neuronal subtype
diversity peaks during development. Genes encoding transcription factors and cell-surface
molecules are the most differentially expressed, indicating their central roles in specifying
neuronal identity. Finally, we show that PNs use highly redundant combinatorial molecular
codes to distinguish subtypes, enabling robust specification of cell identity and circuit
assembly.

overall_design: Single-cell RNA-seq


title: Exome sequencing analysis of murine medulloblastoma models identifies Wdr11 as a potential tumor suppressor in Group 3 tumors

status: Public on Oct 17 2017

city: Buffalo

first: Lei

last: Wei

state: NY

address: Elm & Carlton Streets

department: Biostatistics & Bioinformatics

institute: Roswell Park Cancer Institute

summary: We previoiusly identified WDR11 as a potential tumor suppressor in murine medulloblastoma models. To determine additional genes/pathways affected by WDR11 overexpression.To compare somatic mutations of murine models with human medulloblastoma (MB), we performed whole-exome sequencing of mouse tumors representing three distinct MB subgroups: Wnt, Sonic Hedgehog (Shh) and Group 3 (G3). 64 somatic mutations were identified and validated, including 40 predicted to cause amino acid changes. After filtering and cross-species analysis with 366 human MBs from four independent studies, human orthologs for 16 of the 40 mouse genes were found to harbor non-silent mutations in human MB. Loss-of-function Mll2 mutations detected in one mouse tumor were previously reported in 30 of 366 human MBs.  In mice with G3 MB, one mouse that died at least 15 days earlier than the others had four novel candidate genes harboring non-silent somatic mutations, Lrfn2, Smyd1, Ubn2 and Wdr11. To test whether these genes had tumor suppressive activity, we constitutively overexpressed each wild type gene in murine G3 tumorspheres followed by intracranial implantation.  Mice harboring mouse G3 MB overexpressing WDR11 showed extended survival compared to the other three genes.  Genes in the KEGG WNT signaling pathway, including Ccnd1/2/3, Myc and Tcf7l1, were down-regulated in G3 MB tumorspheres overexpressing WDR11, consistent with reduced tumor progression. In conclusion, we demonstrated that common spontaneous mutations were shared between human and murine models of MB suggesting similar molecular mechanisms of tumorigenesis, and identified WDR11 as a protein with tumor suppressive activity in G3 MB.

overall_design: Compare differentially expressed genes in WDR11 overexpression group versus control group.

first: Martine

middle: F

last: Roussel

first: Jinghui

last: Zhang


title: Mouse embryonic stem cells can differentiate via multiple paths to the same state

city: Boston

first: James

middle: Alexander

last: Briggs

state: Massachusetts

address: 200 Longwood Avenue, Harvard Medical School

department: Department of Systems Biology

institute: Harvard Medical School

overall_design: Refer to individual Series


title: Digital counting of nucleic acid molecules using random base molecular barcodes

city: Suita

first: TAISAKU

last: OGAWA

state: Osaka

address: 6-2-3 Furuedai

country: Japan

institute: RIKEN

summary: Several template DNA molecules with random base molecular barcodes were amplified and sequenced, and the efficacy of the random base barcode for digital counting was shown.

overall_design: Examination of DNA template quantfication using two replicates.

first: Taisaku

last: Ogawa

first: Kirill

last: Kryukov

first: Tadashi

last: Imanishi

first: Katsuyuki

last: Shiroguchi


title: METTL16 is a N6-methyladenosine (m6A) methyltransferase that targets pre-mRNAs and various non-coding RNAs

city: Goettingen

first: Markus

middle: T.

last: Bohnsack

address: Humboldtallee 23

department: Institute for Molecular Biology

country: Germany

institute: Goettingen University

summary: N6-methyladenosine (m6A) is a highly dynamic RNA modification that has recently emerged as a key regulator of gene expression. While many m6A modifications are installed by the METTL3-METTL14 complex, others appear to be introduced independently, implying that additional human m6A methyltransferases remain to be identified. Using crosslinking and analysis of cDNA (CRAC), we reveal that the putative human m6A “writer” protein METTL16 binds to the U6 snRNA and other ncRNAs as well as numerous lncRNAs and pre-mRNAs. We demonstrate that METTL16 is responsible for N6-methylation of A43 of the U6 snRNA and identify the early U6 biogenesis factors La, LARP7 and the methylphosphate capping enzyme MEPCE as METTL16-interaction partners. Interestingly, A43 lies within an essential ACAGAGA box of U6 that base pairs with 5’ splice sites of pre-mRNAs during splicing, suggesting that METTL16-mediated modification of this site plays an important role in splicing regulation. The identification of METTL16 as an active m6A methyltransferase in human cells expands our understanding of the mechanisms by which the m6A landscape is installed on cellular RNAs.

overall_design: HEK293 cell lines expressing His-FLAG-tagged human METTL16 or the His-FLAG tag alone were crosslinked using wavelengths of 254nm (UV-CRAC) or after 4-thiouridine incorporation 365nm (PAR-CRAC), and analysed by CRAC

first: Ahmed

middle: S

last: Warda

first: Jens

last: Kretschmer

first: Philipp

last: Hackert

first: Christof

last: Lenz

first: Henning

last: Urlaub

first: Claudia

last: Höbartner

first: Katherine

middle: E

last: Sloan

middle: T


title: Maturing an Enteric Nervous System in Human Intestinal Organoid-derived Tissue-Engineered Small Intestine

city: Los Angeles

first: Brendan

middle: H

last: Grubbs

address: 4661 W. Sunset Blvd.

department: Obstetrics and Gynecology

institute: University of Southern California

summary: Acquired or congenital disruption in enteric nervous system (ENS) development or function can lead to significant mechanical dysmotility. ENS restoration through cellular transplantation may provide a cure for enteric neuropathies. We have previously generated human pluripotent stem cell (hPSC)-derived tissue-engineered small intestine (TESI) from human intestinal organoids (HIO). However, HIO-TESI fails to develop an ENS. In a previous report of combined HIO with additional human enteric neural crest cells (ENCC), an ENS was established but lacked maturity. The purpose of our study is to establish a mature ENS derived exclusively from hPSC in HIO-TESI. hPSC-derived ENCC supplementation of HIO-TESI generates ENCC-HIO-TESI with mature submucosal and myenteric ganglia, repopulates excitatory, inhibitory, and sensory neurons, and restores the neuroepithelial circuit and neuron-dependent contractility and relaxation. Our findings validate a novel approach to restoring a functional hPSC-derived ENS in ENCC-HIO-TESI and implicate their potential for the treatment of enteric neuropathies.

overall_design: Examination of HIO-TESI growth with and without the addition of HESC-derived ENCC.

first: Christopher

middle: R

last: Schlieve

first: Tracy

last: Grikscheit

first: Matthew

last: Thornton


title: Sodium arsenite effects on DNA methylation in zebrafish embryos.

city: Camden

first: Jaroslav

last: Jelinek

state: NJ

address: 403 Haddon Avenue

department: Research Labs

institute: Coriell Institute for Medical Research

type: Methylation profiling by high throughput sequencing

summary: There is a need to develop biomarkers in alternative testing models predictive for chemically induced chronic disease in humans, preferably describing key events and their relationships in an adverse outcome pathway analysis. Epigenetic modifications, and particularly DNA methylation effects, have been implicated as a major event in susceptibility to develop chronic disease. Arsenic exposure affects large populations around the world through drinking water and industrial activities. We sought to identify epigenetic markers of arsenic exposure. We analyzed the effect of sodium arsenite on DNA methylation in Danio rerio (zebrafish) embryos using unbiased methylation profiling by high throughput sequencing.

overall_design: Pools of zebrafish embryos exposed to 0.1 mM and 1 mM sodium arsenite and non-exposed controls were analyzed in quadruplicates by DNA methylation profiling using DREAM (Digital Restriction Enzyme Analysis of Methylation). In short, genomic DNA was sequentially digested by SmaI and XmaI, which both recognize the sequence CCCGGG. SmaI is methylation sensitive, where XmaI is methylation insensitive. Distinct signatures, 5’-GGG at unmethylated sites or 5’-CCGGG at methylated sites were created by enzyme digestion and ultimately high througput sequencing was used to map these sites to the genome. Methylation ratios for each individual CCCGGG site were calculated as a proportion of methylated counts to the sum of unmethylated and methylated counts, and subsequently adjusted for differences in restriction enzyme efficiency using values obtained from spiked in standards.

first: Leo

middle: M

last: van der Ven

first: Hennie

last: Hodemaekers

first: Edwin

last: Zwart

first: Sander

last: Ruiter

first: Evert-Jan

last: van den Brandhof

first: Frank

last: Bakker

first: Jean-Pierre

middle: J

last: Issa

first: Jeroen

last: Pennings

first: Mirjam

last: Luijten


title: ENCoRE: a lightweight software for CRISPR screens identifies new players in apoptosis

city: Neuherberg

first: Dietrich

last: Trümbach

address: Ingolstädter Landstraße 1

department: Institute for Developmental Genetics

institute: Helmholtz Zentrum München German Research Center for Environmental Health

summary: As CRISPR/Cas9 mediated screens with pooled sgRNA libraries in somatic cells become increasingly established, an unmet need for rapid and accurate companion informatics tools has emerged. We have developed a lightweight standalone software to easily manipulate raw large next generation sequencing (NGS) datasets derived from such screens into informative relational context with graphical support. We demonstrate the capabilities of the software to interrogate meaningful results from an in vitro viability screen using Tumor Necrosis Factor-alpha (TNFa). The results not only identified stereotypical players in extrinsic apoptotic signaling but two as yet uncharacterized members of the apoptotic cascade, Smg7 and Ces2a. We further characterized cell lines containing mutations in these genes against a panel of cell death stimuli. In summary, this software enables bench scientists without access to informatic cores to rapidly access and interpret results from large scale experiments resulting from CRISPR/Cas9 library screens.

overall_design: Comparison of mouse fibroblast control cells with cells treated by TNFalpha

first: Susanne

last: Pfeiffer

first: Manuel

last: Poppe

first: Hagen

last: Scherb

first: Sebastian

last: Doll

first: Wolfgang

last: Wurst

first: Joel

middle: A

last: Schick


title: Small RNA-seq analysis of circulating miRNAs to identify phenotypic variability in Friedreich's ataxia patients

city: Valencia

first: Marta

last: Seco-Cervera

address: Av Blasco Ibañez, 15

institute: University of Valencia

summary: Friedreich’s ataxia (FRDA; OMIM 229300), an autosomal recessive neurodegenerative mitochondrial disease, is the most prevalent hereditary ataxia. In addition, FRDA patients showed additional non-neurological features such as scoliosis, diabetes and cardiac complications. Hypertrophic cardiomyopathy, which is found in two thirds of patients at the time of diagnosis, is the primary cause of death in these patients. In this data set, using small RNA-sequencing of small RNA purified from plasma samples of FRDA patients and controls we identified differential expression of miRNAs (hsa-miR-128-3p, hsa-miR-625-3p, hsa-miR-130b-5p, hsa-miR-151a-5p, hsa-miR-330-3p, hsa-miR-323a-3p, and hsa-miR-142-3p) between both groups. In addition, we found that miR-323a-3p can be used as a biomarker for differentiation of FRDA patients with cardiac problems. Identification of miRNA signatures could therefore provide new molecular explanation for pathological mechanisms occurring during the natural history of the FRDA. Since miRNA levels change with disease progression and pharmacological interventions, miRNAs will contribute to design new therapeutic strategies and improve clinical decisions.

overall_design: Plama miRNA profiles of 25 Friedreich's ataxia patients and 17 healthy subjects were generated by deep sequencing using Illumina HiScan SQ.

first: Dayme

last: González-Rodríguez


title: Mouse embryonic stem cells can differentiate via multiple paths to the same state [RNA-seq]

first: James

last: Briggs

summary: In embryonic development, cells must differentiate through stereotypical sequences of intermediate states to generate mature states of a particular fate. By contrast, direct programming can generate similar fates through alternative routes, by directly expressing terminal transcription factors. Yet the cell state transitions defining these new routes are unclear. We applied single-cell RNA sequencing to compare two mouse motor neuron differentiation protocols: a standard protocol approximating the embryonic lineage, and a direct programming method. Both undergo similar early neural commitment. Then, rather than transitioning through spinal intermediates like the standard protocol, the direct programming path diverges into a novel transitional state. This state has specific and abnormal gene expression. It opens a 'loop' or 'worm hole' in gene expression that converges separately onto the final motor neuron state of the standard path. Despite their different developmental histories, motor neurons from both protocols structurally, functionally, and transcriptionally resemble motor neurons from embryos.

overall_design: 2 protocols were profiled at 3 timepoints each (samples 1-5), and compared to primary embryonic motor neurons (sample 6). Each sample contains 667-1,057 single-cell transcriptomes (total = 5,405).

first: Victor

last: Li

first: Seungkyu

last: Lee

first: Clifford

last: Woolf

first: Allon

last: Klein

first: Marc

middle: W

last: Kirschner


title: Biological sex influences gene expressions in cardiac myocytes

status: Public on Oct 16 2017

city: Boulder

first: Leslie

last: Leinwand

state: CO

address: 596 UCB

department: MCDB

institute: University of Colorado at Boulder

summary:
Purpose:Heart disease is the number one killer of men and women, but not much is known about baseline differences in the heart between males and females
Method: Adult rat ventricular myocytes (ARVMs) were isolated from male and female rats and then RNA was isolated and RNA sequencing was performed.
Results: We identified ~ 600 transcripts that were differentially expressed in cardiac myocytes from either sex. We also observed that enriched pathways from this data set were sexually dimorphic

overall_design: ARVMs were isolated, plated for 45 minutes and then frozen with liquid nitrogen. We had at least 5 biological replicates for each sex; n=6 males and n=5 females

first: Christa

last: Trexler


title: Mtor knockout in PDAC cells

city: Berlin

first: Matthias

last: Wirth

address: Hindenburgdamm 30

institute: Charité Berlin

summary: A pancreatic ductal adenocarcinoma cell line was isolated from a Pdx1-Flp, FSF-KRasG12D/+, Rosa26-CreERT2/tdEG, Mtor-lox/lox mouse. Cultured cells were treated with 4-Hydroxytamoxifen to induce CreERT2 mediated excision of floxed Exon 3 of Mtor.

overall_design: Tamoxifen induced knockout of Mtor in a Pdx1-Flp, FSF-KRasG12D/+, Rosa26-CreERT2/tdEG, Mtor-lox/lox pancreatic cancer cell line

first: Günter

last: Schneider

first: Zonera

last: Hassan


title: RNA transcription profile of different yeast mutants under glucose starvation (0.05% glucose) and comparison of transcriptome of WT and H3D_H3H

city: Shanghai

first: Jin

middle: Qiu

last: Zhou

address: 320 Yueyang Road

department: Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences

institute: Chinese Academy of Sciences

summary: Modifications on histone tails largely affect chromatin associated processes. Previous studies have shown the existence of asymmetrically modified nucleosomes in promoters in multiple cell types. However, whether modifications on both sister histones contribute equally to chromatin dynamics remains elusive. Here we devised a bivalent nucleosome system which allows for the constitutive assembly of asymmetrically modified sister histone H3 in nucleosomes in vivo. We found that the H3K36 methylation on either of sister histones sensitize chromatin structure in gene coding region, while the H3K79 methylation on both sister histone H3s are indispensable for maintaining silent chromatin near telomeres. Additionally, the H3K4 methylation on sister histones plays an equivalent role in suppressing the recruitment of Gal4 activator at GAL1 promoter and GAL1 transcription. Moreover, under starvation stress the expression of a significant proportion of genes in the whole genome is affected by the lack of the H3K4 methylation on one tail of the sister histones, indicating that both sister histones function in fine-tuning transcription through their modifications. We conclude that modifications on sister histones either cooperatively or independently respond to the change of an environmental cue to regulate chromatin structure and function. This work provides a unique experimental strategy to investigate the cross talk between sister histones, and for the first time elaborate their contributions to chromatin in vivo.

overall_design: Examination of the profile of RNA transcription of different mutants under glucose starvation (0.05% glucose) and comparison of transcriptome of WT and H3D_H3H

first: Zhen

first: Li

last: Ma

first: Ya-nan

last: Hu


title: RNA-seq of HBV-infected differentiated HepaRG under RG7834

city: Basel

first: Roland

last: Schmucki

address: Grenzacherstrase

country: Switzerland

institute: F. Hoffmann - La Roche AG

summary: Monitor expression of all viral mRNAs including polyA-deficient viral mRNAs

sample_organism:
Homo sapiens
Hepatitis B virus

platform_organism:
Homo sapiens
Hepatitis B virus

overall_design: 36 HepaRG + HBV infection; treated with RG7834 100 nM or not treated; timepoints 6h, 9h, 12h, and 24h

first: Henrik

last: Mueller

first: Erich

last: Kueng

first: Jean-Christophe

last: Hoflack

first: Tomas

last: Racek

first: Jitao

last: Zhang


title: Time series total RNA sequencing of a differentiation of human embryonic stem cells towards trophoblast lineage

status: Public on Oct 14 2017

first: Dmitry

last: Shaposhnikov

institute: Helmholtz Center Munich

summary: Human embryonic stem cells (hESC) can be differentiated towards progeny resembling trophoblast upon exposure to BMP4. Here we analysed transcriptional changes in cells undergoing commitment to trophoblast at 6 time points using total RNA sequencing and identified early and late regulators of trophoblast fate.

overall_design: hESC cell line H9 was differentiated towards trophoblast lineage using treatment with BMP4. Cells were collected at time points 0 hours (undifferentiated cells), 8 hours, 16 hours, 24 hours, 48 hours and 72 hours and subjected to total RNA sequencing using Illumina platform.


title: Single-cell absolute contact probability detection reveals chromosomes are organized by multiple low-frequency yet specific interactions

city: Taipei

first: Jia-Ming

last: Chang

address: NO.64, Sec. 2, ZhiNan Rd., Wenshan District

department: Department of Computer Science

country: Taiwan

institute: National Chengchi University

summary: The discovery that genomes are partitioned into self-interacting modules or topologically associated domains (TADs) has revolutionized our understanding of the multiscale organization of chromatin. TADs are often considered as stable structures over time, however this view contrasts with the structural dynamics needed for DNA to cope rapidly and accurately with cell-specific and developmental programs regulation. Here, we combine super-resolution microscopy with state-of-the-art DNA labeling methods to reveal the variability in the multiscale organization of chromosomes in Drosophila . We find that TAD borders display very low contact frequency independently o f TAD size, epigenetic state, or cell type. Moreover, long range interactions and assemble of epigenetic domains into active/repressed compartments display different degrees of stochasticity that globally depended on cell-type. Overall, our results show that stochasticity is specifically modulated by sequence and epigenetic state at all levels of chromatin organization, revealing a novel mechanism for the spatial organization and regulation of genomes.

overall_design: Hi-C experiments in drosophila S2 cell

first: Diego

middle: I

last: Cattoni

first: Andrés

last: Cardozo Gizzi

first: Mariya

last: Georgieva

first: Marco

last: Di Stefano

first: Alessandro

last: Valeri

first: Delphine

last: Chamousset

first: Christophe

last: Houbron

first: Stephanie

last: Déjardin

first: Jean-Bernard

last: Fiche

first: Inma

last: González

first: Thomas

last: Sexton

first: Marc

last: Marti-Renom

first: Frédéric

last: Bantignies

first: Giacomo

last: Cavalli

first: Marcelo

last: Nollmann


title: RNA-Seq comparative analysis of human neuroblastoma cells before and after their confrontation to the embryonic microenvironment

city: Lyon

first: joel

last: lachuer

address: 8 avenue rockefeller

institute: university Lyon1

summary: We set up an innovative model for neuroblastoma that consists in using the chick embryo to place human neuroblastoma cells back to their original environment: the trunk neural crest. In this context, human neuroblastoma cells migrate towards sympathetic derivatives and form proliferative tumor masses. To assess the impact of the embryonic microenvironment on neuroblastoma cells gene program, we performed an RNASeq comparative analysis of neuroblastoma naïve cells and tumor masses settled in sympathetic derivatives.

overall_design: We performed independent graft experiments and for each, we collected neuroblastoma naïve cells (before the graft) and tumor masses formed in sympathetic ganglia (50 hours after the graft). 12-13 tumor masses formed in independent chick embryos were microdissected and pooled for each experiment to achieve enough RNA material.

first: Valérie

last: Castellani

first: Céline

last: Delloye-Bourgeois


title: Polysome-seq in zebrafish early embryonic development

city: Warsaw

first: Cecilia

middle: Lanny

last: Winata

state: Mazovia

address: 4 Ks. Trojdena

country: Poland

institute: International Institute of Molecular and Cell Biology in Warsaw

summary: We followed the polysomal association of maternal and early zygotic transcriptome over the first few hours of embryonic development, prior to and after MBT. We isolated polysome-associated (bound) and non-polysome-associated (unbound) mRNAs using sucrose gradient centrifugation followed by size fractionation. Using next generation sequencing (RNA-seq), we profiled the transcriptome in polysome-bound and unbound fractions. Our analysis revealed distinct dynamics of polysome association of cytoplasmically polyadenylated maternal mRNAs.

overall_design: rRNA-depleted RNA was extracted from pooled embryos of desired stages (Egg, 1-cell, 16-cells, 128-cells, 3.5hpf and 5.3hpf) and subjected to polysome profiling to separate between polysome-free and polysome-associated fragments. Unfractionated, rRNA-depleted samples at corresponding stages were used as background control. One RNA-seq library was generated for each sample.

first: C

first: S

last: Mathavan


title: Microvesicle-mediated delivery of miR-1343: impact on markers of fibrosis

city: Cleveland

first: Ann

last: Harris

state: OH

address: 10900 Euclid Ave

department: Genetics and Genome Sciences

institute: Case Western Reserve University

summary: We previously identified miR-1343 as a potent repressor of TGF-b signaling and fibrosis through the direct attenuation of both canonical TGF-b receptors. Here, we build upon our previous findings to better characterize the function of endogenous miR-1343 in normal biology. CRISPR/Cas9 techniques were used to delete the miR-1343 locus in A549 lung epithelial cells. Loss of miR-1343 was found to impact several processes and genes implicated in fibrosis and known to be TGF-b pathway effectors. These responses are opposite to those we observed previously when miR-1343 was overexpressed in the same cell type.

overall_design: mRNA profiles of 3 independent deletion clones and 3 non-targeted (NT) clones from the same experiment, as well as 3 replicates of non-clonal A549 cells.

first: Lindsay

last: Stolzenburg


title: Ribosome-free RNA sequencing of long noncoding RNAs and circular RNAs in lung adenocarcinoma

status: Public on Oct 13 2017

city: Nanjing

first: Mantang

last: Qiu

state: Jiangsu

address: 42 Baiziting

department: Thoracic Surgery

institute: Cancer Institute of Jiangsu Province

summary: Noncoding RNAs play important roles in various biological processes and diseases, including cancer. Expression profile of circular RNAs (circRNA) is largely unknown in lung adenocarcinoma. This study is designed to explore mRNA, long noncoding RNA (lncRNA), and circRNA in lung adenocarcinoma.

overall_design: 9 pairs of lung adenocarcinoma tissues and paired nontumor tissues were collected. Every 3 tumor and every 3 nontumor tissues were pooled together for RNA sequencing, and 6 samples were sequenced. Primary lung adenocarcinoma tissues and paired nontumor tissues were snap frozen with liquid nitrogen immediately after resection and stored in liquid nitrogen till RNA extraction.

first: Rong

last: Yin

first: Ming

last: Li

first: Jun

last: Wang


title: Gene expression profiles of Ofd1 and Kif3a mutant SMB21 cells generated by CRISPR knockout

status: Public on Oct 15 2017

first: Xuesong

last: Zhao

address: 450 Brookline Ave

department: Cancer Biology

institute: Dana-Farber Cancer Institute

summary: Drug resistance poses a great challenge to targeted cancer therapies. In Hedgehog pathway-dependent cancers, the scope of mechanisms enabling resistance to Smo-inhibitors is not known. Here, we performed a transposon mutagenesis screen in medulloblastoma and identified multiple modes of resistance. Surprisingly, mutations in ciliogenesis genes represent a frequent cause of resistance, and patient datasets indicate that cilia loss constitutes a clinically relevant category of resistance. Conventionally, primary cilia are thought to enable oncogenic Hedgehog signaling. Paradoxically, we find that cilia loss protects tumor cells from susceptibility to Smo-inhibitors and maintains a “persister” state that depends on continuous low output of the Hedgehog program. Persister cells can serve as a reservoir for further tumor evolution, as additional alterations synergize with cilia loss to generate aggressive recurrent tumors. Together, our findings reveal novel patterns of resistance and provide mechanistic insights for the role of cilia in tumor evolution and drug resistance.

overall_design: mRNA profiles of CRISPR-GFP (3 biological replicates), CRISPR-Ofd1 (3 biological replicates) and CRISPR-Kif3a (2 biological replicates) cells

first: Rosalind

last: Segal


title: Serum microRNA sequencing for diagnosis of invasive ovarian cancer

first: Kevin

middle: Meyer

last: Elias

state: MA

address: 75 Francis Street

department: OB/GYN

institute: Brigham and Women's Hospital

summary: We constructed a heterogeneous patient cohort of pre-treatment (prior to either surgery or chemotherapy) blood samples from 180 women enrolled in two independent prospective cohort studies of women presenting with an adnexal mass.  One sample (E-1056) was later excluded due to an abnormally high level of mir-122 believed to be due to a recent MI.  The remaining 179 samples were subdivided 3:1 into a 135 patient training set and 44 patient validation set.  Total serum RNA was extracted, converted into miRNA next generation sequencing libraries, and sequenced.  The variables for classification model development were selected using three methods – a significance filter (using a student’s t-test ), a group-stratified fold change filter, and a correlation-based feature selection (CFS). We deployed 11 different machine learning algorithms on the three sets of variables to separate the cases of invasive cancer from the healthy controls or benign/borderline masses.  The tools were graded in terms of receiver operating characteristic area under the curve (ROC AUC).  The model was validated both by qPCR on the study samples as well as by external validation on an independent publicly available dataset of 454 patients with a range of diagnoses, including ovarian cancer.  The qPCR signature was then externally validated on another independent external cohort.

overall_design: miRNA-seq on 180 pre-treatment serum samples

first: Dipanjan

last: Chowdhury


title: Expression Profiling of MEFs and SKOM induced reprogramming cells after modulation of Cell cycle and Dnmt1 expression

city: Guangzhou

first: Hui

last: Zheng

state: Guangdong

address: 190 Kaiyuan Avenue

institute: Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences

summary: To generate association of DNA methylation level and gene expression profile, we applied RNA-sequencing on mouse embryonic fibroblasts (MEFs) after the modulation of cell cycle and Dnmt1 expression level. To characterize how these factors affect somatic reprogramming, samples of MEFs induced by Yamanaka factors (Sox2, Klf4, Oct4, cMyc) was also included. The cell cycle was accelerated by shRNA targeted to p53, and the expression of Dnmt1 was manipulated by either over expression or knock down.

overall_design: RNA sequencing of cell cycle modulated MEFs and SKOM induced reprogramming cells. SKOM-induced cells were grown either in the presence of Vitamin C (Vc) or absence of Vc.

first: Songwei

last: He

first: Hao

last: Sun


title: Dynamics of embryonic stem cell differentiation inferred from single-cell transcriptomics show a series of transitions through discrete cell states

city: Cambridge

first: Sumin

last: Jang

address: 52 Oxford Street, Rm 402

institute: Harvard University

summary: Using a statistical framework to analyze single-cell transcriptomics data, we infer the gene expression dynamics of early mouse embryonic stem (mES) cell differentiation, uncovering discrete transitions across nine cell states. We validate the predicted transitions across discrete states using flow cytometry. Moreover, using live-cell microscopy, we show that individual cells undergo abrupt transitions from a naïve to primed pluripotent state. Using the inferred discrete cell states to build a probabilistic model for the underlying gene regulatory network, we further predict and experimentally verify that these states have unique response to perturbations, thus defining them functionally.

overall_design: mRNA profiles of 288 single mouse embryonic stem cells undergoing various stages of in vitro early germ layer differentiation were generated on an Illumina Hi-Seq platform

first: Vilas

last: Menon

first: Anne-Rachel

last: Krostag

first: Boaz

last: Levi

first: Sharad

last: Ramanathan


title: DNA methylomes at base resolution of mouse embryonic fibroblasts in G1 and G2 phase of cell cycle

summary: Dramatic change in DNA methylation patterns and levels both globally and/or locus-specifically is associated with the process of cell lineage specification and somatic reprogramming. We found the expression of DNA methyltransferase 1 (Dnmt1), the enzyme that maintains 5-methylcytosine (5mC) after DNA replication, is tightly regulated by the progression of cell cycle. Upregulation of Dnmt1 was observed in cells with shortened G1 and G2 phase. We hypothesize that 5mC level is not completely recovered by Dnmt1 immediately after DNA replication. In the attempt to clarify DNA methylation status before and after DNA replication, we performed Whole Genome Bisulfite Sequencing (WGBS) to map genome-wide DNA methylation separately in G1 and G2 phase of mouse embryonic fibroblasts (MEFs). Cells of different cell cycle phases were sorted by flow cytometry after Propidium iodide (PI) staining. We find an average of 2% decrease of global DNA methylation in G2 phase compared with G1, with the trend more magnificent in high CpG density regions. Generally, the results indicate that the G1 phase of cell cycle is an important time window for the stability of DNA methylation inheritance.

overall_design: Whole genome bisulfite sequencing of G1 and G2 phase MEFs.

first: Lingyu


title: Mechanisms of HERV-K (HML-2) transcription during human mammary epithelial cell transformation

city: Boston

first: John

last: Coffin

address: 136 Harrison Ave.

department: Molecular Biology & Microbiology

institute: Tufts University

summary: Elevated transcript expression of the endogenous retrovirus family HERV-K (HML-2) is seen in the majority of breast cancers, although the identity of the individual loci contributing to this expression as well as their mechanism of activation is unclear. Using high-throughput next-generation sequencing techniques optimized for the capture of HML-2 expression, we produced a complete profile of the HML-2 transcriptome before and after human mammary epithelial cell transformation.

overall_design: Using Illumina MiSeq sequencing, we profiled the HERV-K (HML-2) mRNA expression in four human mammary epithelial cell lines of varying tumorigenicity.

first: Meagan

last: Montesion


title: Pervasive concerted evolution in gene expression shapes cell type transcriptomes

city: Heidelberg

first: Jacob

last: Musser

address: Meyerhofstrasse 1

institute: European Molecular Biology Laboratory

summary: Complex multicellular organisms have evolved numerous cell types with many different functions. Comparative transcriptomic data yields valuable insights into cell type, tissue, and organ evolution. However, interpreting this data requires understanding how transcriptomes evolve. A particularly difficult problem is that cell type transcriptomes may not evolve independently, a key assumption of most evolutionary analyses. Non-independence of cell types can occur when cell types share regulatory mechanisms. This leads to concerted evolution in gene expression across different cell types, confounding efforts to unravel the history of cell type evolution, and identify cell type-specific patterns of expression. Here we present a statistical model to estimate the level of concerted transcriptome evolution and apply it to published and new data. The results indicate that tissues undergo pervasive concerted evolution in gene expression. Tissues related by morphology or developmental lineage exhibit higher levels of concerted evolution. Concerted evolution also causes tissues from the same species to be more similar in gene expression to each other than to homologous tissues in another species. This result may explain why some tissue transcriptomes cluster by species rather than homology. Our analysis of bird skin appendages data suggests levels of concerted evolution also varies with phylogenetic age of the tissue. Our study illustrates the importance of accounting for concerted evolution when interpreting comparative transcriptome data, and should serve as a foundation for future investigations of cell type evolution.

sample_organism:
Dromaius novaehollandiae
Gallus gallus

platform_organism:
Dromaius novaehollandiae
Gallus gallus

overall_design: We sampled epidermis from 4 different tracts of developing bird skin appendages in chicken and emu embryos: feather placodes, scutate scale placodes, reticulate scale placodes, and claw placodes. In chicken we sampled two biological replicates for each skin appendages. In emu we sampled 2 biological replicates for feathers and scutate scales, and 1 biological replicate for reticulate scales and claw (due to limitations in available material).


title: Genome-wide DNA methylation maps of mouse embryonic fibroblasts and reprogramming cells modulated by cell cycle and Dnmt1 expression

summary: Dramatic change in DNA methylation patterns and levels both globally and/or locus-specifically is associated with the process of cell linage specification and somatic reprogramming. We found the expression of DNA methyltransferase 1 (Dnmt1) is tightly regulated by the features of cell cycle, which we referred as cell cycle-directed Dnmt1 expression adjustment. To further investigate how DNA methylation is affected by this mechanism, we applied reduced representation bisulphate sequencing (RRBS) to map genome-wide DNA methylation on mouse embryonic fibroblasts (MEFs). To characterize how this mechanism affects somatic reprogramming, samples of MEFs induced by Yamanaka factors (Sox2, Klf4, Oct4, cMyc) was included. The cell cycle was accelerated by shRNA targeted to p53, and the expression of Dnmt1 was manipulated genetically. We find 5mC level remains constant in regardless of cell proliferation rate, a result from the adjusted expression of Dnmt1. However, repressed expression of Dnmt1 in fast-proliferating cells results in global DNA demethylation. The patterns of DNA methylation and demethylation affected by this mechanism is sensitive to CpG densities. Generally, the results demonstrate cell cycle-directed Dnmt1 expression adjustment as an mechanism for the insurance of the stability of genomic 5mC inheritance.

overall_design: Reduced representation bisulphate sequencing in cell cycle modulated MEFs and SKOM induced reprogramming cells.

last: Li


title: Sequential inflammatory processes define human progression from M. tuberculosis infection to tuberculosis disease

city: Seattle

first: Daniel

last: Zak

state: WA

address: 188 East Blaine Street, Suite 300

institute: Regeneron

summary: To define mechanisms that underlie progression of Mycobacterium tuberculosis (M.tb) infection to active tuberculosis (TB), we followed M.tb-infected adolescents longitudinally. Those who ultimately developed TB disease (“progressors”) were compared with matched controls, who remained healthy. Whole blood transcriptomic and plasma proteome analyses showed sequential modulation of immunological processes: type I/II interferon signalling and complement cascade were elevated 18 months before TB diagnosis, while changes in myeloid inflammation and lymphoid cell, monocyte and neutrophil gene modules occurred more proximally to TB disease. Analysis of gene expression in purified T cells revealed early suppression of Th17 responses in progressors. This was confirmed in an adult BCG re-vaccination cohort, where expression of interferon response genes in blood was associated with suppression of IL-17 expression by BCG-specific CD4 cells. We concluded that sequential inflammatory dynamics and immune alteration precede TB disease manifestation, with important implications for developing diagnostics, vaccines and host-directed therapies for TB.

overall_design: 6,363 healthy adolescents, aged 12-18 years and enrolled into the Adolescent Cohort Study (ACS) (Zak et al. 2016), were followed for 24 months or more. Blood was collected at 6-monthly intervals. Among those with evidence of M.tbinfection (QuantiFERON and/or tuberculin skin test positive), 41 ultimately developed microbiologically confirmed TB disease >6 months after study enrolment; they were designated “progressors”. Among progressors, the time from blood collection to diagnosis of active TB (“time to diagnosis”) ranged from 1 to 894 days. Realigning blood collection to “time to diagnosis” allowed serial assessment of transition from infection to TB disease. Controls (N=104), who remained asymptomatic, were matched to progressors by age, gender, ethnicity, school and prior history of TB .

first: Ethan

last: Thompson

first: Adam

last: Penn Nicholson

first: Thomas

last: Scriba

first: Smitha

last: Shankar


title: Opposing roles of dendritic cells subsets in experimental GN

status: Public on Oct 12 2017

city: Cologne

first: Sebastian

last: Brähler

address: Joseph-Stelzmann-Str. 26

department: Nephrology

institute: University Hospital of Cologne

summary: Classical dendritic cells (DCs) are key players at the interface between innate and adaptive immunity. In the kidney exist 2 major subsets of cDCs: CD11b+ cDCs and CD103+ cDCs. We investigated their function in the most widely used model of experimental glomerulonephritis (GN) in mice: nephrotoxic nephritis (NTN). Consistent with a role for cDCs in nephrotoxic nephritis, depletion of ZBTB46+ cells (all cDCs) attenuated kidney injury, while deficiency of the CD103+ subset of cDCs accelerated injury via a mechanism that involved increased neutrophils. This RNAseq was performed to analyze transcriptional changes in FACS-sorted renal CD11b+ and CD103+ cDCs under healthy conditions and at day 7 of NTN to reveal why both subsets have different functions in GN.

overall_design: The study was performed with total of 6 mice (wildtype, male, age 8-12 weeks). 3 mice were sacrificed in the healthy situation, 3 mice were sacrificed 7 days after injection of the nephrotoxic nephritis antiserum (NTN). From each mouse CD11b+ and CD103+ DCs were sorted, resulting in 4 experimental conditions with 3 biological replicates each: CD103_healthy, CD11b_healthy, CD103_NTN, CD11b_NTN.

first: Andrey

last: Shaw


title: Melanocyte Stem Cell Activation and Translocation Initiate Cutaneous Melanoma in Response to Ultraviolet Exposure

city: Ithaca

first: Jennifer

middle: K

last: Grenier

address: 526 Campus Rd

department: Biomedical Sciences

institute: Cornell University

summary: We report the gene expression changes during melanoma initiation from tumor-competent melanocyte stem cells

overall_design: Examination of gene expression profilies between 3 different cell types

first: Hyeongsun

last: Moon

first: Andrew

middle: C

last: White


title: Genome-nuclear lamina interactions regulate progenitor cell lineage restriction during cardiogenesis

city: Philadelphia

first: Michael

middle: Patrick

last: Morley

state: PA

address: 3400 Civic Center Blvd, Bldg 421

department: Penn Cardiovascular Institute

institute: Perelman School of Medicine at the University of Pennsylvania

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

overall_design: Refer to individual Series


title: Microbiota-associated intestinal methylation changes confer immune-metabolic gene network of NEC in preterm pig model

city: Shenzhen

first: Desheng

last: Gong

address: No.7 PengFei road

institute: Agricultural Genomes Institute at Shenzhen

summary: Intestinal microbiota colonization is important for intestinal development and health of preterm infants, especially those with extremely low birth weight. Recent studies indicated for a dynamic crosstalk between that gut microbiota and DNA methylome of host intestinal cells. Thereby, we sought to determine the epigenomic and metagenomic consequences of suppression of microbiota colonization in the intestine of preterm neonates to gain insights into biological pathways that shape the interface between the gut microbiota and the preterm intestinal cells. We examined 14 preterm piglets by comparing the conventional preterm neonates with those ones treated with oral antibiotics for genome wide DNA methylation and 16S rDNA microbiome. Our results demonstrated an extensive genome-wide DNA methylation changes in response to the suppression of intestinal microbe colonization, especially genes involved in neonatal immune response signaling and glycol-metabolism pathways were identified. Our study highlights several key genes that might predispose preterm neonates to NEC risk due to their key roles involved in the immune-metabolic networks. Our study not only provided rich omic-data to interpret molecular program in relation with microbiota-associated methylome-proteome network changes, but also confer clinical usage of key gene markers for potential early diagnostics of NEC of preterm neonates.

overall_design: Fourteen piglets from three sows (Danish Landrace x Large White x Duroc) were Caesarean-delivered at preterm (d 106 or 90% of gestation), which was established as a model for the preterm human infant at approximately 70% gestation due to the intestinal maturity. Pigs were randomized according to birth weight and sex into two groups: half were conventionally raised with spontaneous commensal microbiota colonization by receiving saline through orogastric catheter (CON group, n=7) and the other half with delayed colonization by receiving broad-spectrum antibiotics through orogastric catheter (OAB group, n=7).

first: Xiao

middle: Y

last: Pan


title: Stable 5-Hydroxymethylcytosine (5hmC) Acquisition Marks Gene Activation During Chondrogenic Differentiation.

city: Stanford

first: Nidhi

last: Bhutani

state: California

address: 300 Pasteur Drive, R164 Edwards Bldg

department: Orthopaedic Surgery

institute: Stanford University


title: Detecting Activated Cell Populations Using Single-Cell RNA-Seq

status: Public on Oct 11 2017

city: Los Angeles

first: Ye

middle: Emily

last: Wu

state: CA

address: 635 Charles E. Young Dr. South

institute: UCLA

summary: Single-cell RNA sequencing of mouse amygdala using Drop-Seq

overall_design:
Drop-Seq was performed for amygdala cells dissociated from male C57BL/6J mice in different experimental groups using different single-cell dissociation methods.

Single-cell suspensions were generated using the Act-Seq protocol or conventional dissociation protocol, and processed through Drop-Seq to produce single-cell cDNA libraries attached to microbeads. cDNAs were amplified by PCR and prepared for sequencing through tagmentation.

first: Weizhe

last: Hong


title: mTOR kinase inhibition effectively decreases progression of a subset of neuroendocrine tumors that progress on rapalog therapy and delays cardiac impairment

city: Cincinnati

first: Hala

middle: Elnakat

last: Thomas

state: Ohio

address: 3125 Eden Ave,Vontz bldg, rm 1226, ATTN: Hala Elnakat Thomas

department: Internal Medicine

institute: UNIV OF CINCINNATI

summary: Inhibition of mTOR signaling using the rapalog everolimus is an FDA-approved targeted therapy for patients with lung and gastroenteropancreatic neuroendocrine tumors (NET). However, patients eventually progress on treatment, highlighting the need for additional therapies. We focused on pancreatic NETs (pNETs) and reasoned that treatment of these tumors upon progression on rapalog therapy, with an mTOR kinase inhibitor (mTORKi) such as CC-223 could overcome a number of resistance mechanisms in tumors and delay cardiac carcinoid disease. We performed preclinical studies using human pNET cells in vitro and injected them subcutaneously or orthotopically to determine tumor progression and cardiac function in mice treated with either rapamycin alone or switched to CC-223 upon progression. Detailed signaling and RNA sequencing analyses were performed on tumors that were sensitive or progressed on mTOR treatment. Approximately 57% of mice bearing pNET tumors which progressed on rapalog therapy showed a significant decrease in tumor volume upon a switch to CC-223. Moreover, mice treated with an mTORKi exhibited decreased cardiac dilation and thickening of heart valves than those treated with placebo or rapamycin alone. In conclusion, in the majority of pNETs that progress on rapalogs, it is possible to reduce disease progression using an mTORKi, such as CC-223. Moreover, CC-223 had an additional transient cardiac benefit on valvular fibrosis compared to placebo- or rapalog-treated mice. These results provide the preclinical rationale to further develop mTORKi clinically upon progression on rapalog therapy and to further test their long term cardioprotective benefit in those NET patients prone to carcinoid syndrome.

overall_design: We performed RNA sequencing analyses as an unbiased means to assess changes in gene expression. Our major goal was to identify the differences in tumor mRNAs between the CC-223- and non-CC-223 responders compared to the rapamycin alone treatment arm (Fig 5A in Orr-Asman et al manuscript). The analysis was conducted using 1 tumor each from 13 and 14 mice treated with rapamycin or switched to CC-223 respectively.

middle: E


title: Population RNA sequencing of Ly6D+ and TN EPLM subpopulations as well as Pro-B cells

city: Basel

first: Llucia

last: Alberti-Servera

state: Basel stat

address: Mattenstrasse 28

department: Department of Biomedicine

institute: Uni Basel

summary: Nowadays, there is an active debate regarding the heterogeneity of apparently phenotypically homogenous progenitors having different multiple lineage potentials. It has been previously characterized a B220+ CD117int CD19- NK1.1- uncommitted and multipotent haematopoietic progenitor with combined lymphoid and myeloid developmental potential, the so called Early Progenitor with Lymphoid and Myeloid potential (EPLM). This project is devoted to unravelling whether the EPLM is truly multipotent or whether it is composed by a mixture of cells with distinct lineage biases. Based on the expression of the cell-surface markers Ly6D, SiglecH and CD11c, the EPLM can be subdivided into at least four major subpopulations. By interrogating the transcriptome of the subpopulations (bulk RNA sequencing), we aim to unravel their genetic signatures and to identify the key genes driving their precursor-product relationship.

overall_design: Total RNA was extracted from ex-vivo sorted hematopoietic progenitors (from Flt3Ltg mice) using TRIzol-based method. Transcriptome profiles were generated by sequencing, in quadruplicates, using NextSeq 500.

first: Robert

last: Ivanek

first: Antonius

last: Rolink


title: Next-Generation Sequencing Facilitates Quantitative Analysis of the Effects of Wnt Agonist Treatments on Palate Formation

city: Salt Lake City

first: Shihai

last: Jia

state: ut

address: 20 South 2030 East, Room 320 BPRB

department: Department of Neurobiology

institute: University of Utah

summary: Nonsyndromic clefts of the palate and/or lip are common birth defects arising in about 1/700 live births worldwide. They are caused by multiple genetic and environmental factors, can only be corrected surgically and require complex post-operative care that imposes significant burdens on individuals and society. Our understanding of the molecular networks that control palatogenesis has advanced through studies on mouse genetic models of cleft palate. In particular, the transcription factor Pax9 regulates palatogenesis through the Bmp, Fgf and Shh pathways in mice. But there is still much to learn about Pax9's relationship with other signaling pathways in this process. Here we show alterations of Wnt expression and decreased Wnt activity in Pax9-/- palatal shelves are a likely result of Pax9's ability to directly bind and repress the promoters of Dkk1 and Dkk2, proteins that antagonize Wnt signaling. We exploited this relationship by delivering small-molecule Dkk inhibitors into the tail-veins of pregnant Pax9+/- females from E10.5 to E14.5. Such therapies restored Wnt signaling, promoted cell proliferation, bone formation and fusion of palatal shelves in Pax9-/- embryos. These data uncover a connection between the roles of Pax9 and Wnt genes in palatogenesis and offer a new approach for treating human cleft palates.

overall_design: E14 embryos of Pax9-/- and control littermates with or without WAY-262611 treatment, mouse embryos palate were micro-dissected, control and mutant samples were separated and individually lysed for the RNA extraction.

first: Rena

last: D'Souza


title: Differences in DNA-binding specificity of floral homeotic protein complexes predict organ-specific target genes

city: Berlin

first: Jose

middle: M.

last: Muino

address: Max-Dohrn-Str. 8-10

institute: German Federal Institute for Risk Assessment

summary: Floral organ identities in plants are specified by the combinatorial action of homeotic master regulatory transcription factors (TFs). How these factors achieve their regulatory specificities is however still largely unclear. Genome-wide in vivo DNA binding data show that homeotic MADS-domain proteins recognize partly distinct genomic regions, suggesting that DNA binding specificity contributes to functional differences of homeotic protein complexes. We used in vitro systematic evolution of ligands by exponential enrichment followed by high throughput DNA sequencing (SELEX-seq) on several floral MADS-domain protein homo- and heterodimers to measure their DNA-binding specificities. We show that specification of reproductive organs is associated with distinct binding preferences of a complex formed by SEPALLATA3 (SEP3) and AGAMOUS (AG). Binding specificity is further modulated by different binding site (BS) spacing preferences. Combination of SELEX-seq and genome-wide DNA binding data allows to differentiate between targets in specification of reproductive versus perianth organs in the flower. We validate the importance of DNA-binding specificity for organ-specific gene regulation by modulating promoter activity through targeted mutagenesis. Our study shows that intrafamily protein interactions affect DNA-binding specificity of floral MADS-domain proteins. DNA-binding specificity of individual dimers, as well as DNA-binding preferences of higher-order complexes differ between floral homeotic protein complexes. Differential DNA-binding of MADS-domain protein complexes plays a role in the specificity of target gene regulation.

overall_design: SELEX-seq for MADS-domain protein homo- and heterocomplexes using random pool of DNA and in vitro synthetized proteins.

first: Cezary

last: Smaczniak

last: Muiño

first: Dijun

last: Chen

first: Gerco

last: Angenent

first: Kerstin

last: Kaufmann


title: Stable 5-Hydroxymethylcytosine (5hmC) Acquisition Marks Gene Activation During Chondrogenic Differentiation [seq]

summary: Regulation of chondrogenic differentiation by DNA demethylation is little understood. The ten-eleven-translocation (TET) proteins oxidize methylated cytosines (5mC) to 5hmC, 5fC and 5caC eventually leading to DNA demethylation. However, 5hmC is stable and can potentially act as an epigenetic mark as well. Here, we report that global changes in 5hmC mark chondrogenic differentiation.

overall_design: Examination of changing 5hmC levels in progenitor and differentiated chondrocytes.

first: Sarah

last: Taylor

middle: H

last: Li

first: Wing

last: Wong

first: Piera

last: Smeriglio


title: Molecular characterisation of sorted populations of DRG and TG ganglia neurons

city: London

first: Franziska

last: Denk

address: Guy's Campus

department: Wolfson SPaRC

country: United Kingdom

institute: King's College London

summary: Sensory neurons were extracted from the dorsal root or trigeminal ganglia of AdvGFP transgenic mice and their molecular profile analysed via RNA-seq.

overall_design: RNA-seq of naïve FACS-purified DRG and TG neurons

first: D

last: Lopes

first: S

middle: B

last: McMahon

first: F


title: RNAseq of duodenal tissue from C57BL/6 mice both susceptible and resistant to Heligmosomoides polygyrus

first: Lewis

middle: James

last: Entwistle

state: London

address: 1 Midland Road

institute: The Francis Crick Institute

summary:
Purpose: Identify genes implicated in resistance to H. polygyrus
Method: RNA was extracted from duodenal tissue from C57BL/6 mice during primary and secondary H. polygyrus. Total RNA libraries were created using the Encore® Complete RNA-Seq Library Systems kit (NuGEN), following manufacturer’s instructions. Total RNA libraries were sequenced using the Illumina® HiSeq 2500. The raw Illumina reads were analysed as follows. First, the data quality was analysed using FastQC. Then the low quality bases were trimmed using Trimmomatic. The read pairs which passed the trimming quality filters were then aligned to mm10 (Ensembl version 75) using Tophat2. Counts were determined using htseq_count. Normalisation and statistical analysis was performed using edgeR. Differential gene analysis was calculated from naïve control group (A). Statistically significant genes with FDR < 0.05 are reported.
Results: Following analysis, we identified many transcripts significantly different from the naïve control following primary and secondary H. polygyrus infection.
Conclusions: The intestinal transcriptome of both primary and secondary infected mice differ significantly, identifying qualitative and quantitative differences between susceptible (primary infected) and resistant (secondary infected) mice.

overall_design: Total RNA profiles were generated from duodenal tissue of mice infected with the intestinal helminth H. polygyrus using the Illumina HiSeq 2500. 6 groups, 8 replicates per group. All data compared to naïve group (A).

middle: J

first: Nikolay

last: Nikolov

first: Mark

middle: S

last: Wilson


title: Methyl-Spec-seq control data sets

city: St. Louis

first: Gary

middle: D.

last: Stormo

state: MO

address: 4515 McKinley Ave. Room 4208.

department: Genetics

institute: Washington University School of Medicine

summary: We developed a novel in-vitro experimental method to characterize the protein-DNA interaction specificity and methylation sensitivity, we called Methyl-Spec-seq. In this data set, mouse AP1 was used as a positive control example to show that Methyl-Spec-seq can determine the relative binding energy for variants with different methylation property, i.e., unmethylated, top hemimethylated, bottom hemimethylated, duplex methylated by either chemical synthesis or enzymatic treatment.

overall_design: Except different methylation types of DNA, the overall design is the same as conventional Spec-seq, which we have published work before.

first: Yiming

last: Chang


title: RNA transcription profile of different yeast mutants under glucose starvation (0.05% glucose)

city: Shanghai

first: Jin

middle: Qiu

last: Zhou

address: 320 Yueyang Road

institute: Chinese Academy of Sciences

summary: Histone tail modificationscan greatly influencechromatin-associated processes. Asymmetrically modified nucleosomes exist inmultiple cell types; however,whether modifications on both sister histones contribute equally to chromatin dynamicsremainselusive. Here, we devised a bivalent nucleosome system thatallowedfor the constitutive assembly of asymmetrically modified sister histone H3s in nucleosomes inSaccharomyces cerevisiae. The sister H3K36 methylationsaffectedcryptic transcription in gene coding regions, whereassister H3K79 methylation had cooperative effectsongene silencing near telomeres.H3K4 methylation on sister histonesplayed an independent role in suppressing the recruitmentof the Gal4 activator and inhibitingGAL1transcription. Under starvation stress, the expression of glycometabolism pathway geneswasfinetuned by sisterH3K4 methylation.Thus,different modifications on sister histones act either cooperatively or independentlyto regulate transcription in response to external stimuli.

overall_design: Examination of the profile of RNA transcription of different mutants under glucose starvation (0.05% glucose)

first: Zhen

first: Li

last: Ma

first: Yu-ting

last: Liu

first: Ya-nan

last: Hu


title: RNAseq of IL-36 stimulated primary human keratinocytes

first: Francesca

last: Capon

address: Great Maze Pond

department: Genetics and Molecular Medicine

summary: Keratinocytes isolated from one healthy donor were stimulated in triplicate for 24h with IL-36α, IL-36β or IL-36γ

overall_design: Gene expression profile of IL-36 stimulated keratinocytes


title: Formaldehyde facilitates cell transformation by compromising chromatin assembly

city: Tuxedo

first: Chunyuan

last: Jin

state: NY

address: 57 Old Forge Rd

institute: New York University School of Medicine

summary: Formaldehyde (FA) is an environmental and occupational chemical carcinogen. Recent studies demonstrated that exogenous FA caused only a modest increase in DNA adducts above levels caused by endogenous FA. This raised a possibility that epigenetic mechanisms might contribute to FA-mediated carcinogenicity. Here we report a drastic decrease of acetylation of N-terminal tails of the cytosolic histones following FA exposure, the modifications important for histone nuclear import and assembly into chromatin. In fact, the amount of histone proteins in chromatin fraction decreases following FA exposure. Moreover, the histone depletions are evident at most of the genomic loci we tested, suggesting that FA compromises chromatin assembly.  Notably, FA increases chromatin accessibility and changes expression of hundreds of cancer-related genes. Importantly, the knockdown of histone H3.3 (a H3 variant), which mimics inhibition of chromatin assembly, facilitates FA-mediated cell transformation. We propose that inhibiting chromatin assembly represents a novel mechanism for FA carcinogenesis.

overall_design: mRNA profiles of formaldehyde-responsive genes in BEAS-2B cells were generated by deep sequencing in duplicates using Ilumina HiSeq

last: Chunyuan

first: Mei

last: Shenglin


title: Independent manipulation of histone H3 modifications in individual nucleosomes reveals the contributions of sister histones to transcription

department: Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences


title: Gene expression profiles of parental and mutant SMB21 cells derived from drug resistance screen

city: Boston

first: Xuesong

last: Zhao

state: Massachusetts

address: 450 Brookline Ave

institute: Dana-Farber Cancer Institute

summary: Drug resistance poses a great challenge to targeted cancer therapies. In Hedgehog pathway-dependent cancers, the scope of mechanisms enabling resistance to Smo-inhibitors is not known. Here, we performed a transposon mutagenesis screen in medulloblastoma and identified multiple modes of resistance. Surprisingly, mutations in ciliogenesis genes represent a frequent cause of resistance, and patient datasets indicate that cilia loss constitutes a clinically relevant category of resistance. Conventionally, primary cilia are thought to enable oncogenic Hedgehog signaling. Paradoxically, we find that cilia loss protects tumor cells from susceptibility to Smo-inhibitors and maintains a “persister” state that depends on continuous low output of the Hedgehog program. Persister cells can serve as a reservoir for further tumor evolution, as additional alterations synergize with cilia loss to generate aggressive recurrent tumors. Together, our findings reveal novel patterns of resistance and provide mechanistic insights for the role of cilia in tumor evolution and drug resistance.

overall_design: mRNA profiles of parental (3 replicates), Sonidegib-treated parental (2 replicates), Sufu mutant (2), and Ofd1 mutant (4) SMB21 cells

first: Rosalind

middle: A

last: Segal


title: A transposon screen identifies loss of primary cilia as a mechanism of resistance to Smo inhibitors


title: N6-adenine DNA methylation in Tetrahymena

city: New York

first: Chao

last: Lu

state: New York

address: 1130 Saint Nicholas Ave

department: Genetics & Development

institute: Columbia University Medical Center

summary: In this study, we provided the first genome-wide, base pair-resolution map of 6mA in Tetrahymena by applying single-molecule real-time (SMRT) sequencing.

overall_design: Examination of N6-adenine DNA methylation on Tetrahymena MAC genome.

first: Shan

last: Gao

first: Xiao

last: Chen


title: The compact genome in the clavulanic acid producing Streptomyces  strain unravels a variety of horizontal transfer reservoirs

status: Public on Oct 10 2017

city: Jinan

first: Guangxiang

last: Cao

address: 18877 jingshi Road

institute: Shandong Academy of Medical Sciences

summary: Bacterial genomic plasticity and instability carry multiple functional genetic information in Streptomyces secondary metabolism. Our previously publication has reported an effective industrial Streptomyces strain, with a unique phenotype of the high clavulanic acid yield. The complete genome of strain F163-1 harboring a 136.9-kb giant region of plasticity (RGP) was sequenced. The chromosome and plasmid are densely packed by an exceptionally huge variety of potential secondary metabolic gene clusters, excluding production of putative antibiotics. Intriguingly, architecture and size differences of plasmid pSCL4 between F613-1 and ATCC 27064 suggest the pSCL4 plasmid evolving from pSCL4-like and pSCL2-like extrachromosomal replicons, in addition to the previously proposed ATCC 27064 mega-plasmid formation hypothesis through recombination between the smaller F613-1 pSCL4 plasmid arm regions and the linear chromosome. Comparative genomics systemically investigate secondary metabolism capacitates in this study indicates that frequent exchange of genetic materials between Streptomyces replicons may shape remarkable diversities of secondary metabolite repertoires. Consequently, the F613-1 strain seems to have evolved its specific genomic architectures and genetic patterns to meet the requirement in subsequent industrial processes.

overall_design: mRNA profiles of S. clavuligerus F613-1 and ATCC27064 strains were generated by deep sequencing

first: Chuanqing

last: Zhong


title: RNAseq Analysis of Mouse Stomach and Intestines Following Vertical Sleeve Gastrectomy

city: South San Francisco

first: Joshua

last: Lichtman

address: 333 Oyster Point Boulevard

department: Translational Science

institute: NGM Biopharmaceuticals

summary: Bariatric surgery, an effective treatment for obesity and diabetes, leads to profound remodeling of whole body energy homeostasis. We utilized a mouse model of vertical sleeve gastrectomy (VSG), a common bariatric surgery as a tool to identify novel secreted proteins and peptides that might act as important metabolic regulators. We analyzed gene expression in the stomach and intestines following VSG or sham surgery in diet-induced obese mice and sought to identify differentially regulated genes encoding secreted proteins/peptides.

overall_design: Gene expression analysis of 5 different mouse intestinal tissues after VSG or sham surgery

first: Xuecai

last: Ge

first: Hong

last: Yang

first: Maria

last: Bednarek

first: Hadas

last: Galon-Tilleman

first: Peirong

first: Michael

first: Yan

last: Wang

first: Olivier

last: Dalmas

first: Yiyuan

last: Yin

first: Hui

last: Tian

first: Lutz

last: Jermutus

first: Joseph

last: Grimsby

first: Cristina

middle: M

last: Rondinone

first: Anish

last: Konkar

first: Daniel

middle: D

last: Kaplan


title: Roles of female and male genotype in post-mating responses in Drosophila melanogaster

first: Sofie

last: Delbare

address: 435 East 30th Street

institute: NYU Langone

summary:
Purpose: Mating induces a multitude of changes in female behavior, physiology and gene expression. Interactions between female and male genotype lead to variation in post-mating phenotypes and reproductive success. So far, few female molecules responsible for these interactions have been identified.
Methods: We used Drosophila melanogaster from five geographically dispersed populations to investigate such female x male genotypic interactions at the female transcriptomic and phenotypic levels.
Methods: Females from each line were singly-mated to males from the same five lines, for a total of 25 combinations. To assess whether female x male genotypic interactions affect the female post-mating transcriptome, next-generation RNA sequencing was performed on virgin and mated females at 5 to 6 hours post-mating.
Results: Seventy-seven genes showed strong variation in mating-induced expression changes in a female x male genotype-dependent manner. These genes were enriched for immune response and odorant-binding functions, and for expression exclusively in the head.
Conclusions: The transcriptional variation found in specific functional classes of genes might be a read-out of female x male compatibility at a molecular level. Understanding the roles these genes play in the female post-mating response will be crucial to better understand the evolution of post-mating responses and related conflicts between the sexes.

overall_design:
Five Drosophila melanogaster inbred lines were used. These lines are derived from five geographically dispersed populations (Global Diversity Lines Beijing 04; Ithaca 16; Netherlands 01; Tasmania 01 and Zimbabwe 184 – the latter line was collected in Africa, but turned out to be a recent migrant) (Grenier et al., 2015).
Virgin females from each line were singly-mated to virgin males from each of the five inbred lines, similar to a 5x5 full factorial design.
For RNAseq, mated females were flash frozen 5 to 6h after the start of mating. Age-matched virgin females were flash-frozen in parallel.
Three independent biological replicates were generated for each of the 25 mating combinations and for virgin females of each genotype (90 samples total). Flies from each replicate were collected from separate bottles, and matings for all three replicates were set up simultaneously. RNA was extracted from five to ten pooled females per replicate.

middle: N

first: Clement

last: Chow

first: Mariana

middle: F

last: Wolfner

first: Andrew

middle: G

last: Clark


title: RNAseq and ChIPseq on Mouse Embryonic Stem Cells and Sorted Mesendoderm Progenitor Cells

city: Lausanne

first: Frederic

last: Burdet

address: Quartier Sorge - Batiment Amphipole

institute: Swiss Institute of Bioinformatics

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

overall_design: Refer to individual Series