title: The chromatin landscape at the HIV-1 provirus integration site determines viral expression

city: Barcelona

first: Guillaume

last: Filion

address: Dr. Aiguader 88

department: GRSCC

institute: CRG

summary: We investigated the link between HIV-1 integration by using B-HIVE and LEDGINs. B-HIVE tracks insert-specific HIV expression by tagging a unique barcode in the HIV genome. LEDGINs are antivirals that inhibit the interaction between HIV-IN and its chromatin tethering cofactor LEDGF/p75. They are known to retarget HIV-1 integration sites. Here, we confirmed that LEDGIN treatment retargets integration out of transcriptionally active regions and reduce HIV expression. Silent provirus was located at increased distance to H3K36me3, the recognition marker of LEDGF/p75, after treatment with LEDGINs. Viral RNA expression was also influenced by the proximity of enhancers, regardless of the presence of LEDGINs.

overall_design: Genomic DNA and RNA was extracted from SupT1 and Jurkat cells 14 days after transduction with barcoded HIV in the presence of varying concentrations of LEDGIN CX014442 (6.25, 15.62 or 31.25µM).

first: Gerlinde

last: Vansant

first: Heng-Chang

last: Chen

first: Eduard

last: Zorita

first: Zeger

last: Debyser


title: Low-grade glioma microglia

status: Public on Jul 03 2020

city: St Louis

first: Winnie

middle: W

last: Pong

state: MO

address: 660 S. Euclid Avenue

department: Department of Neurology

country: USA

institute: Washington University School of Medicine

type: Expression profiling by high throughput sequencing

summary: To examine the effect of Nf1 inactivation in astrocytes on normal microglia in the optic nerve, microglia from Nf1 flox/flox (FF) and Nf1 flox/flox; GFAP-Cre (FFC) were flow sorted. High throughput RNA-seq was employed to compare the expression profiles, and upregulated genes in microglia supportive to Nf1 deficient astrocytes were verified.

overall_design: Microglia from mice lacking Nf1 in astrocytes were flow sorted. Upregulated genes were verified and further examined.

first: Jason

last: Walker

first: Todd

last: Wylie

first: Vincent

last: Magrini

first: Malachi

last: Griffith

first: Obi

middle: L

first: Elaine

middle: R

last: Mardis

first: David

middle: H

last: Gutmann


title: Human junior pilocytic astrocytoma

summary: Microglia in the microenvironment of low-grade tumors have been shown to allow and support tumor growth, but the expression profile of these microglia are unknown. To examine the role of microglia in junior pilocytic astrocytomas, macrophages were flow sorted from cyroprotected surgical excisions of tumors and normal brain tissues. Upregulated markers of tumor-associated microglia were verified and compared to mouse tumor-associated microglia.

overall_design: Junior pilocytic astrocytoma macrophages were flow sorted. Upregulated genes of glioma associated microglia were verified and further examined.

first: Patrice

last: Bittner

first: Jeffrey

last: Leonard


title: Next Generation Sequencing of Wild Type and Zeb1-/- CLP and HSCs

status: Public on Jul 02 2020

city: CARDIFF

first: Peter

last: Giles

address: Heath Park

country: United Kingdom

institute: Cardiff University

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

overall_design: Refer to individual Series


title: Next Generation Sequencing of Wild Type and Zeb1-/- CLP

summary: Zeb1, a zinc finger E-box binding homeobox epithelial-mesenchymal (EMT) transcription factor, confers properties of ‘stemness’, such as self-renewal, in cancer. Yet little is known about the function of Zeb1 in adult stem cells. Here, we used the hematopoietic system, as a well-established paradigm of stem cell biology, to evaluate Zeb1 mediated regulation of adult stem cells. We employed a conditional genetic approach using the Mx1-Cre system to specifically knockout (KO) Zeb1 in adult hematopoietic stem cells (HSCs) and their downstream progeny. Acute genetic deletion of Zeb1 led to rapid onset thymic atrophy and apoptosis driven loss of thymocytes and T cells. A profound cell-autonomous self-renewal defect and multi-lineage differentiation block was observed in Zeb1 KO HSCs. Loss of Zeb1 in HSCs activated transcriptional programs of deregulated HSC maintenance and multi-lineage differentiation genes, and of cell polarity, consisting of cytoskeleton, lipid metabolism/lipid membrane and cell adhesion related genes. Notably, Epithelial cell adhesion molecule (EpCAM) expression was prodigiously upregulated in Zeb1 KO HSCs, which correlated with their enhanced cell survival capacity and diminished differentiation in transplantation. Thus, Zeb1 acts as a crucial transcriptional regulator in hematopoiesis, co-ordinating HSC self-renewal and multi-lineage differentiation fates, in part, via EpCAM repression.

overall_design: Wild Type and Zeb1-/- CLP were isolated from 8-to-12-week old mice (4 mice per genotype)

first: Alhomidi

last: Almotiri

first: Hamed

last: Alzahrani

first: Neil

last: Rodrigues


title: Single-nuclei transcriptomic analysis of the subthalamic nucleus reveals different Pitx2- positive subpopulations

city: Uppsala

first: Åsa

last: Wallén-Mackenzie

address: Norbyvägen 18A

country: Sweden

institute: Uppsala University

summary: Single nuclei from the subthalamic nucleus from Pitx2-Cre mice (Skidmore et al.Mol. Cell. Neurosci. 37, 2008) crossed with mCherryTRAP reporter (Gt(ROSA)26Sor-mCherry-Rpl10a)  (Hupe et al. Nucleic Acid Res. 2014) that was used to FACS sort Pitx2mCherry positive nuclei at postnatal day 28 to identify subpopulations.

overall_design: Sequencing libraries were prepared from FACS sorted individual nuclei with the Smart-Seq2 protocol  (Picelli et al. Nature Methods 2013)

first: Maria

last: Papathanou

first: Asa

middle: K

last: Bjorklund


title: Antihistamine Drug Ebastine inhibits cancer growth by targeting Polycomb Group Protein EZH2

city: Houston

first: Guangyu

last: Wang

address: 6670 Bertner Ave

institute: Houston methodist research institute

overall_design: Examination of mRNA profiles level in 2 cell types

first: Killia

middle: Y

last: Liu

first: Qi

last: Cao

first: Kaifu

last: Chen


title: Variability in cerebrospinal fluid microRNAs through life

city: Derio

first: Ana

middle: Maria

last: Aransay

state: BIZKAIA

address: Parque tecnologico de Bizkaia, Building 801-A

department: Genome Analysis Platform

country: Spain

institute: CIC bioGUNE

type: Non-coding RNA profiling by high throughput sequencing

summary: The development of the human brain starts in the first weeks of embryo differentiation. However, there are many relevant neurodevelopmental processes that take place after birth and during lifespan. Such a fine and changing scenario requires the coordinated expression of thousands of genes to achieve the proper specialization and inter-connectivity. In this context, microRNAs (miRNAs), which can modulate mRNA stability and translation, are gaining recognition for their involvement in both brain development and neurodevelopmental disorders. Therefore, cerebrospinal fluid (CSF) miRNAs should be perfectly differentiated in relevant age periods. In this study, we aimed to highlight the biological variability of miRNA expression in the CSF throughout life, which is also crucial for biomarker discovery in CNS pathologies, especially in children, where they are desperately needed. We analyzed the CSF microRNAome of 14 healthy children (aged 0–7.4 years) by smallRNA-Seq and compared it with previously published data in adults (N = 7) and elders (N = 11). miR-423-5p and miR-22-3p were overexpressed in the < 1 and > 3 years groups, respectively. Additionally, we detected 18 miRNAs that reached their highest peak of expression at different time-points during the lifespan and sets of miRNAs that were exclusively expressed in a specific age group. On the contrary, miR-191-5p showed stable expression in CSF from the first year of life. Our results remark the complex differential miRNA expression profile that can be observed through life, which underlines the need for including appropriate age-matched controls when the expression of CSF miRNAs is analyzed in different pathological contexts.

overall_design: The total RNA was extracted from 200 μL of cerebrospinal fluid and then small RNA sequencing was performed.

first: Endika

last: Prieto-Fernández

first: Elixabet

last: Lopez-Lopez

first: Idoia

last: Martin-Guerrero

first: Laura

last: Bárcena

first: Monika

last: Gonzalez-Lopez

middle: M

first: Juan

middle: J

last: Lozano

first: Javier

last: Benito

last: Falcón-Pérez

first: Africa

last: Garcia-Orad


title: An Integrative Transcriptome – Proteome Approach Reveals a Robust Expression Pattern of Small Intestinal Epithelial Organoids from Germ-Free and Microbiota Associated Donors

city: Zurich

first: giancarlo

last: russo

state: Schweiz

address: winterthurerstrasse 190

department: Functional Genomics Center Zurich

country: Switzerland

institute: ETH/University of Zurich

summary: The goal of this study is to analyze the influence of different donor microbiota on small intestinal epithelial organoid cultures

overall_design: Organoid cultures from mice with two different microbiota or from germ-free mice (3 mice per group) were generated. Organoids were cultured in parallel to passage 5-8 and then frozen for transcriptome analysis. M-ICc12 cells and MEFs were included as control/reference groups. m-ICc12 and Organoid samples were treated with 5 ng/ml TNF for 8 h.

first: Annika

last: Hausmann

first: Giancarlo

last: Russo

first: Mirjam

last: Zuend

first: Miguel

last: Cuenca

first: Schinichi

last: Sunagawa

first: Mikael

last: Sellin

first: Wolf-Dietrich

last: Hardt


title: DNA methylation variations in familial female and male breast cancer

city: Brescia

first: Edoardo

last: Abeni

address: Viale Europa n. 11

department: Department of Molecular and Translational Medicine, Division of Biology and Genetics, University of Brescia, Brescia, Italy.

country: Italy

institute: University of Brescia

type: Methylation profiling by genome tiling array

summary:
About 25% of familial breast cancer (BC) is attributed to germline mutations of BRCA1 and BRCA2 genes while the rest of patients are included in the BRCAX group. BC also affects men with a worldwide incidence of 1%. The epigenetic alterations, including those DNA methylation,  have been rarely studied in the male breast cancer (MBC) on a genome-wide level. The aim of the current work was to study the global DNA methylation profiles of BC patients to look for differences between familial female breast cancer (FBC) and MBC and according to BRCA1, BRCA2 and BRCAX mutation status. The genomic DNA from FFPE tissues of 17 female and 7 male patients with BC was subjected to methylated DNA immunoprecipitation (MeDIP) and hybridized on human promoter microarrays.
The comparison between FBC and MBC showed 2846 differentially methylated regions (DMRs) corresponding to 2486 distinct annotated genes. The gene ontology enrichment analysis revealdrelevant molecular function terms such as the GTPase superfamily genes (in particular the GTPase Rho GAP/GEF and GTPase RAB) and cellular component terms associated to cytoskeletal architecture such as “cytoskeletal part”, “keratin filament”, “intermediate filament". By considering only FBC, several cancer-associated pathways were the most enriched KEGG pathways of differentially methylated genes between BRCA2 and BRCAX or BRCA1+BRCAX groups. The comparison between BRCA1 group vs BRCA2+BRCAX group displayed the enriched molecular function term “cytoskeletal protein binding”. Finally, the functional annotation of differentially methylated genes between BRCAX and BRCA1+BRCA2 groups indicated that the most enriched molecular function terms were related to GTPase activity.
In summary, this is the first study that compares the global DNA methylation profile of familial FBC and MBC and the results may provide useful insights into the epigenomic subtyping of breast cancer and shed light on a possible new molecular mechanisms underlying BC carcinogenesis.

overall_design: The global DNA methylation profiles of 24 breast cancer cases were obtained by methylated DNA immunoprecipitation coupled with Affymetrix Human Promoter 1.0R Tiling Arrays (MeDip-chip). The genomic DNA from 24 breast cancer FFPE tissues was extracted using QIAamp DNA FFPE Tissue Kit supplied by Qiagen- The genomic DNA was digested using micrococcal nuclease (New England Biolabs) to obtain DNA fragments ranging from 200 to 500 nucleotides and used them as input (IN) DNA. Agilent Bioanalyzer with the RNA 6000 Nano LabChip Kit was used to check the size of fragmented DNA. Part of IN DNA (4 µg) was immunoprecipitated (IP) with 10 µl of anti-5-MethylCytosine Antibody (Eurogentec, BI-MECY-0100) using the MeDip protocol. The antibody-DNA complexes were immunoprecipitated using Dynabeads® Protein G immunoprecipitation kit (Life Technologies) and the methylated enriched DNA was purified by standard phenol/chloroform procedure and precipitated with isopropanol. A total of 200 ng of IN and IP DNA was amplified with the Affymetrix Chromatin Immunoprecipitation Assay Protocol.  Totalling 2 arrays per each breast cancer case were performed (1 IP DNA and 1 IN DNA).  The CEL files were imported into Partek® Genomics Suite® (PGS). We performed 9 different comparisons according to gender and mutation status (BRCA1, BRCA2 and BRCAX). For each comparison performed the significantly differentially methylated regions (DMR) were obtained using ANOVA followed by MAT algorithm (model-based analysis of tiling arrays algorithm). Using PGS function, each DMR was associated to its nearest gene of human genome. The genomic coordinates of each DMR were calculated based on UCSC human genomic assembly version 18.

first: Ilaria

last: Grossi

first: Eleonora

last: Marchina

first: Arianna

last: Coniglio

first: Fabio

last: Facchetti

first: Pietro

last: Cavalli

first: Fausto

last: Zorzi

first: Pier Luigi

last: Chiodera

first: Carlo Terenzio

last: Paties

first: Maria Luisa

last: Crosatti

first: Giuseppina

last: De Petro

first: Alessandro

last: Salvi


title: Evolutionarily Conserved Regulation of Immunity by the Splicing Factor RNP-6/PUF60

city: Koln

first: Jorge

last: Boucas

address: Joseph-Stelzmann-Str. 9b

department: Bioinformatics

country: Germany

institute: Max Planck Institute for Biology of Ageing

summary: Splicing is a vital cellular process that modulates important aspects of animal physiology, including metabolism and lifespan. From a cold stress screen, we identified a novel mutant of the splicing factor RNP-6/PUF60, which evokes resistance to various abiotic stresses, yet is highly sensitive to pathogenic bacterial challenge. Molecular and genetic experiments reveal that RNP-6 suppresses immunity, but promotes animal longevity, suggesting a tradeoff between these processes. Pathogen exposure affects gene expression and splicing in a rnp-6 dependent manner. Another longevity promoting splicing factor, SFA-1, similarly exerts an immuno-suppressive effect, working downstream or parallel to RNP-6. Genetic epistasis and biochemical experiments reveal that RNP-6 acts through PMK-1 MAPK signaling to modulate immunity. The mammalian homolog, PUF60, also displays anti-inflammatory properties, and its levels swiftly decrease after bacterial infection in mammalian cells. Our findings demonstrate an evolutionarily conserved modulation of immunity by specific components of the splicing machinery.

overall_design: Transcriptome of C. elegans wild type (N2) and rnp-6(dh1127) (Day 1 adults) under control and S. aureus infection conditions

first: Chun

last: Kew

first: Adam

last: Antebi


title: Combined Inhibition of JAK/STAT Pathway and Lysine Demethylase 1 as a Novel Therapeutic Strategy in CSF3R/CEBPA Mutant Acute Myeloid Leukemia [RUX_GSK RNA-seq]

city: Portland

first: Theodore

middle: Paul

last: Braun

state: OR

address: 3181 SW Sam Jackson Park Road

department: Knight Cancer Institute

institute: OHSU

summary: The presence of CSF3R mutations are associated with adverse outcomes in CEBPA mutant AML. Here we report that inhibitors of LSD1 reverse the mutant CEBPA-mediated differentiation blockade and exhibit synergy with JAK/STAT inhibitors.

overall_design: Comparison of gene expression in mouse bone marrow cells retrovirally expressing mutant CSF3R and CEBPA and treated with Ruxolitinib, GSK-LSD1, or the combination for 24 hours.

middle: P

first: Julia

middle: E

last: Maxson


title: Combined Inhibition of JAK/STAT Pathway and Lysine Demethylase 1 as a Novel Therapeutic Strategy in CSF3R/CEBPA Mutant Acute Myeloid Leukemia [LSD1i ChIP-seq]

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: Comparison of chromatin landscapes in mouse bone marrow cells retrovirally expressing mutant CSF3R and CEBPA and treated with 4 nM GSK-LSD1 for 48 hours.


title: Transcriptomic profiling of normal plasma cells and MM models using bulk RNA-Seq

city: Milwaukee

first: Robert

last: Burns

state: Wisconsin

address: 8727 W Watertown Plank Rd

institute: Blood Research Institute, Versiti

summary: NRAS Q61 mutations are prevalent in advanced/relapsed multiple myeloma (MM) and correlate with poor patient outcomes. Thus, we generated a novel MM model by conditionally activating expression of endogenous NrasQ61R and a MYC transgene in germinal center B cells (VQ mice). VQ mice developed a highly malignant MM characterized by high proliferation index, hyperactivation of ERK and AKT signaling, impaired hematopoiesis, widespread extramedullary disease, bone lesions, kidney abnormalities, preserved PD1 and TIGIT immune checkpoint pathways, and expression of human high-risk MM gene signatures. VQ MM mice recapitulate most of the biological and clinical features of human advanced/high-risk MM. These MM phenotypes are serially transplantable in syngeneic recipients. Two MM cell lines were also derived to facilitate future genetic manipulations.  Combination therapies based on MEK inhibition significantly prolonged the survival of VQ mice with advanced stage MM. Our study provides a strong rationale to develop MEK inhibition-based therapies for treating advanced/relapsed MM.

overall_design: Bulk RNA-Seq analysis was performed using 50,000 sorted CD138+ B220- CD45.2+ bone marrow cells of WT(n=3), V (n=3), recipients of tVk12653 (n=2), VQ-D1 (n=2), VQ-D4 (n=1), and VQ-D5 (n=2).

first: Adhithi

last: Rajagopalan

first: Zhi

last: Wen

first: Jing

last: Zhang


title: Single-cell RNAseq of VQ myeloma cells reveals unique transcriptional signatures from normal plasma cells

overall_design: CD138+ B220- cells were sorted from age-matched control bone marrow (Con-BM); VQ-D2 bone marrow (BM), lymph node (LN) and processed for scRNA-seq


title: Next Generation Sequencing Facilitates Quantitative Analysis of Transcriptomes in fetal and 2 weeks old rat islets

city: Copenhagen

first: Kyoung

middle: Jae

last: Won

state: Hovedstaden

address: Ole Maaløes Vej 5

department: BRIC

country: Denmark

institute: University of Copenhagen

summary: Unlike adult β-cells, fetal and neonatal islets are functional immature and have blunted glucose responsiveness and decreased insulin secretion in response to stimuli. Pancreatic islets are a mixture of different cell types. Study of transcriptomes from intact islets would identify novel molecular mechanisms controlling islet functional development. At e19 and 2 weeks of age, pancreatic islets were isolated and total RNA were extracted for RNA-Seq study. Gene expression profiles were compared in the study.

overall_design: Examination of transcriptomes in 2 time points (e19 and 2 weeks of age)

first: Yu-Chin

last: Lien

first: Rebecca

middle: A

last: Simmons

first: Kyoung-Jae


title: Expression of NrasQ61R and MYC transgene in germinal center B cells induces a highly malignant multiple myeloma in mice


title: A novel model for chronic rejection in vascularized composite allotransplantation identifies CXCL9-11 as potential biomarkers

city: Bochum

first: Stephan

last: Hahn

address: Universitaetsstrasse 150

department: MGO

institute: Ruhr-University Bochum

summary: Global mRNA expression profiling of transplanted rat hindlimbs (muscle and skin) were collected using Agilent rat whole genome array (Agilent-028282 Whole Rat Genome Microarray 4x44K v3). Hind limb transplantation between Fischer344 (donor) and Lewis (recipient) rats were performed. Isogenic transplantations served as controls. Administration of irregular immunosuppression induced chronic rejection. At the endpoint (post-operativ day 100), hind limbs presented clinicial, histomorphological and genetic changes, known to be highly suspicious for chronic rejection.

overall_design: Each group consisted of n=5 animals. Isogenic hind limb transplantations (Lewis as donor and recipient) served as controls and did not receive any immunosuppression. The chronic rejection group was treated only when signs of an acute skin rejection e.g. erythema or edema (defined as foot-swelling of ≥ 25 %) occured. Both muscle and skin of each group are represented by two biological replicates.


title: Transcriptome upon Mof deletion and Etomoxir treatment in ground-state mouse (m) ESCs

city: Ann Arbor

first: Fengbiao

last: Mao

state: Michigan

address: 1301 Catherine St

department: Department of Pathology

institute: University of Michigan

summary: This study describes the transcriptome profiling of: 1) Mof fl/fl mESCs cultured in 2i/LIF medium with Ethanol (WT) or 4-OHT (Mof-/-); 2) E14 mESCs cultured in 2i/LIF medium treated with H2O (control) or Etomoxir treatment (ETO, FAO inhibition) and ETO-released (Released).

overall_design: RNA-Seq profiling in ground-state mESCs. Biological duplicates of each sample are labled as rep1/2.

first: Yali

last: Dou


title: Combined Inhibition of JAK/STAT Pathway and Lysine Demethylase 1 as a Novel Therapeutic Strategy in CSF3R/CEBPA Mutant Acute Myeloid Leukemia


title: Combined Inhibition of JAK/STAT Pathway and Lysine Demethylase 1 as a Novel Therapeutic Strategy in CSF3R/CEBPA Mutant Acute Myeloid Leukemia [LSD1i RNA-seq]

overall_design: Comparison of gene expression in mouse bone marrow cells retrovirally expressing mutant CSF3R and CEBPA and treated with GSK-LSD1 or DMSO for 48 hours.


title: Comparison of human CD39- and CD39+ thymus-derived regulatory T-cells

city: Vienna

first: Markus

last: Jeitler

address: Lazarettgasse 14

country: Austria

institute: Medical University Vienna

summary:
CD39- and CD39+ human thymus-derived regulatory T-cells (tTreg) present as distinct subsets with specific functional and metabolic response patterns. Both cell types express equal levels of the canonical tTreg transcription factors FOXP3 and Helios and overexpression studies have shown that CD39 expression is independent of these two transcription factors. So far, no transcriptomic analyses to adress these differences have been performed.
We used microarrays to detail the global programme of gene expression between CD39- and CD39+ human tTreg and identified distinct classes of up-regulated and down-regulated genes between these two subsets.

overall_design: Magnetic-beads enriched human CD4+ T-cells from three independent healthy donors were labelled with antibodies to CD4, CD25, CD39 and CD127. Subsequently, the CD4+CD25highCD127low population was identified on a FACS Aria II cell sorter and the CD39- (CD39- tTreg) and CD39+ (CD39+ tTreg) fraction were stringently isolated. Total RNA was then extracted from the cells using the RNeasy micro kit from Qiagen.

first: Klaus

last: Schmetterer

first: Marlene

last: Gerner


title: Genome wide RNA sequencing to profile response to IL-1 in control and IFT88 depleted cells

city: Oxford

first: Angus

last: Wann

address: Roosevelt Drive, Headington

department: Kennedy Institute of Rheumatology

institute: University of Oxford

summary:
Purpose: RNA sequencing across whole genome downstream to IL-1 to investigate scope and nature of effect of IFT88 depletion
Method: Control pool treated and IFT88 siRNA pool targeted Fibroblast-like chondrocytes were treated with or without IL-1B 10ng/ml  and RNA collected in triplicate at 0, 1, 2, 4, 8 and 24h. PolyA-selected sequencing libraries were prepared using the TruSeq protocol (Illumina). Libraries were subject to 75bp paired-end sequencing (Illumina HiSeq 4000) to an average depth of 28.5 million read pairs per sample and aligned to mouse genome with Hisat2.
Results: Using time-course analysis (ImpulseDE2) the IL-1 response timecourse was determined (control-only) and intersected against the comparative case vs control analysis. Per-point differential expression analysis IFT88 vs control was performed using DESeq2.
Conclusion: Demonstration of the effect of IFT88 depletion on IL-1 response signature over 24h.

overall_design: Bulk RNA profiles of control siRNA targetted (control) and IFT88 siRNA (IFT88) treated fibroblastic chondrocyte line, both treated with and without IL-1 beta in quiescent state.

first: Megan

last: McFie

first: Stephen

last: Sansom

first: Lada

last: Koneva


title: RNAseq transcriptome analysis of K. pneumoniae MS6671 with PBT2 and colistin

status: Public on Jul 01 2020

city: Brisbane

first: David

middle: Manuel Pereira

last: De Oliveira

state: QLD

address: St Lucia

department: School of Chemistry and Molecular Biosciences

country: Australia

institute: The University of Queensland

summary: The emergence of polymyxin resistance in carbapenem-resistant and extended-spectrum -lactamase (ESBL)-producing bacteria is a critical threat to human health, and new treatment strategies are urgently required. Here, we investigated the ability of the safe-for-human use ionophore PBT2 to restore antibiotic sensitivity in polymyxin-resistant, ESBL-producing, carbapenem-resistant Gram-negative human pathogens. PBT2 was observed to resensitize Klebsiella pneumoniae, Escherichia coli, Acinetobacter baumannii and Pseudomonas aeruginosa to last-resort polymyxin class antibiotics, including the less-toxic next-generation polymyxin derivative, FADDI-287. We were unable to select for mutants resistant to PBT2 + FADDI-287 in polymyxin resistant E. coli containing a plasmid-borne mcr-1 gene or K. pneumoniae carrying a chromosomal mgrB mutation. Using a highly invasive K. pneumoniae strain engineered for polymyxin resistance through mgrB mutation, we successfully demonstrated the efficacy of PBT2 + FADDI-287 in vivo for the treatment of Gram-negative sepsis. These data present a new treatment modality to break antibiotic resistance in high priority polymyxin-resistant Gram-negative pathogens.

overall_design: For K. pneumoniae MS6671, 4 sample conditions were analyzed. Sample conditions included: 1) bacterial treatment with cation-adjusted Muller Hinton broth (CA-MHB; reference sample);  2)  CA-MHB+PBT2 (64 µM); 3) Colistin (0.25 µg/mL); 4) PBT2 + colistin

first: Minh-Duy

last: Phan

first: Mark

last: Walker


title: Clonal multi-omics reveals Bcor as a regulator of dendritic cell development and its action at a clonal level

city: Cambridge

first: Luyi

last: Tian

state: MA

address: 415 main st Cambridge

institute: Broad institute

summary: Despite major developments in single cell multi-omics approaches, a single stem or progenitor cell can only be tested once. Here we develop ‘clonal multi-omics’, where recent daughters of a clone act as surrogates of the parental cell allowing multiple tests of its molecular profile and/or cellular fate under different conditions. We first present SIS-seq – a novel clonal method whereby siblings are examined in parallel for gene expression by RNA-seq, or for fate after culture. We use this method to identify, then validate using CRISPR, the genes expressed in single haematopoietic progenitors that control the development of the different dendritic cell (DC) subtypes. In this way, Bcor is identified as a suppressor of plasmacytoid DC (pDC) and conventional DC subtype 2 (cDC2) numbers during Flt3 ligand-mediated ‘emergency’ DC development. Using a complementary method SIS-skew – where WT and BcorKO siblings of the same clone are examined in parallel for fate – we discover that Bcor restricts clonal expansion, especially for generation of cDC2s, and suppresses clonal fate potential, especially for pDCs. SIS-seq and SIS-skew therefore represent novel and broadly applicable clonal multi-omics approaches that could reveal the molecular and cellular mechanisms governing stem, immune, reprogrammed and cancer cell function at a single cell level, including after genetic or extrinsic factor perturbation.

overall_design: Mouse HSPCs were isolated from bone marrow and sorted into 384 PCR plate, with library generated using the CEL-seq2 protocol.

first: Sara

last: Tomei

first: Daniela

last: Amann-Zalcenstein

first: Jaring

last: Schreuder

first: Shalin

last: Naik


title: Next Generation Sequencing of Wild Type and Zeb1-/- HSCs

first: Peter

last: Giles

overall_design: Wild Type and Zeb1-/- HSCs were isolated from 8-to-12-week old mice (4 mice per genotype)

first: Alhomidi

last: Almotiri

first: Hamed

last: Alzahrani

first: Neil

last: Rodrigues


title: Setd2 deficiency impaired erythroid differentiation and accelerated Myelodysplastic Syndrome(MDS) - associated leukemogenesis through S100A8 and S100A9

city: Shanghai

first: Jiaying

last: Zhang

state: Shanghai

address: 320 Yue Yang Road

department: CAS Key Laboratory of Tissue Microenvironment and Tumor, Shanghai Institute of Nutrition and Health

country: China

institute: Shanghai Institutes for Biological Sciences, University of Chinese Academy of Sciences, Chinese Academy of Sciences

type: Methylation profiling by high throughput sequencing

summary: Setd2, the histone H3 lysine 36 methyltransferase, plays an important role in the pathogenesis of hematologic malignancies. The research on the role of Setd2 in leukemogenesis has made great progress, but its role in MDS is still unknown. Here, we knock out Setd2 in the NUP98-HOXD13 transgenic (NHD13 Tg) mouse, and demonstrate that loss of Setd2 accelerates the transformation of MDS into AML. The conditional deletion of Setd2 also interferes the differentiation of hematopoietic stem and progenitor cells (HSPCs), and results in the decrease of granulocyte monocyte progenitor (GMP) cells, increase of megakaryocyte erythroid progenitor (MEP) cells and common myeloid progenitor (CMP) cells. Loss of Setd2 impairs erythroid differentiation, includes cell cycle arrest in G2-M and enhances the selt-renewal ability of HSPCS. Our RNA-seq,ChIP-seq and WGBS analysis indicats that S100A8 and S100A9 are direct target genes of H3K36me3 and expression of heterodimeric S100 calcium-binding proteins S100A8 and S100A9 are downregulated in HSPCs when Setd2 deficiency. Addition of recombinant S100A8 or S100A9 weakens the clonogenic progenitors of Setd2 deficient HSPCs. Therefore, our results demonstrate that loss of Setd2 promotes the transformation of MDS into AML, which proves Setd2 as a tumor suppressor in MDS, and provides a potential therapeutic target for MDS associated leukemia.

overall_design: We used Mx1-Cre-induced deletion of Setd2 in Nup98-HoxD13 (NHD13) transgenic mice. C-Kit+ cells were sorted from spleen cells in NHD13 and NHD13_Setd2 KO mice at leukemia stage (NHD13 - AML and NHD13/Setd2 KO - AML), and ChIP-seq was performed with an anti-H3K36me3 antibody. Hematopoietic stem and progenitor cells (HSPCs) were sorted from bone marrow (BM) cells in C57BL/6,  NHD13/Setd2 flox/flox and NHD13/Setd2 KO mice, and C-Kit+ cells were sorted from spleen cells in NHD13/Setd2 KO - AML, and these cells were subjected to RNA-seq analysis and WGBS.

first: Bingyi

last: Chen

first: Shubei

first: Chun-Hui

last: Xu

first: Xiao-Jian

last: Sun

first: Lan

last: Wang


title: Clonal multi-omics in DC development

summary:
Despite major developments in single cell multi-omics approaches, a single stem or progenitor cell can only be tested once. Here we develop ‘clonal multi-omics’, where recent daughters of a clone act as surrogates of the parental cell allowing multiple tests of its molecular profile and/or cellular fate under different conditions. We first present SIS-seq – a novel clonal method whereby siblings are examined in parallel for gene expression by RNA-seq, or for fate after culture. We use this method to identify, then validate using CRISPR, the genes expressed in single haematopoietic progenitors that control the development of the different dendritic cell (DC) subtypes. In this way, Bcor is identified as a suppressor of plasmacytoid DC (pDC) and conventional DC subtype 2 (cDC2) numbers during Flt3 ligand-mediated ‘emergency’ DC development. Using a complementary method SIS-skew – where WT and BcorKO siblings of the same clone are examined in parallel for fate – we discover that Bcor restricts clonal expansion, especially for generation of cDC2s, and suppresses clonal fate potential, especially for pDCs. SIS-seq and SIS-skew therefore represent novel and broadly applicable clonal multiomics approaches that can reveal the molecular and cellular mechanisms governing cell function at a single cell level, including after genetic or extrinsic factor perturbation.

This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Staphylococcus aureus tolerance to antimicrobial methylene-blue-mediated photodynamic inactivation.

city: Rochester

first: Steven

middle: R.

last: Gill

state: NY

address: 601 Elmwood Avenue

department: Microbiology & Immunology

institute: University of Rochester

summary: A promising alternative to antibiotics for treatment of Staphylococcus aureus infections is photodynamic inactivation (PDI), which employs a photosensitizer (PS) that produces cytotoxic reactive oxygen species (ROS) when exposed to molecular oxygen and antimicrobial blue light in the spectrum of 400-470 nm.  Although the precise mechanistic basis of PDI has not been defined, the formation of ROS and free radicals that oxidize a number of cellular targets, including membrane lipids, damage to proteins and nucleic acids result in inactivation of essential cellular functions and subsequent cell death.  Because PDI is non-selective and affects multiple cellular targets, development of resistance or tolerance to PDI has been considered to be unlikely and attempts to induce S. aureus resistance or tolerance upon repeated sub-lethal doses of PDI have not succeeded.  However, multiple aspects of PDI suggest that development of tolerance is highly probable, in particular when PDI is used for treatment of infections where the environment at the infection site prevents penetration of PDI at a level sufficient to cause death of all bacteria and with tolerant phenotypes emerging from the surviving bacteria. In this study, we sought to identify the mechanisms that contribute to PDI tolerance in S. aureus. S. aureus HG003 and the isogenic HG003DmutSL strain with defects in DNA mismatch repair were used to evaluate the response of S. aureus and the roles of DNA mismatch repair and gene regulatory networks to repeated sublethal doses of PDI. Global transcriptome and genome analyses were used in an agnostic approach to identify the underlying transcriptional responses and genetic adaptations that occur as a result of repeated PDI and contribute to PDI tolerance. Our results reveal multiple metabolic, transport and cell wall biogenesis pathways that contribute to PDI tolerance, and a S. aureus regulatory gene likely responsible for the adaptive transcriptional response to PDI.

overall_design: Eight total samples (samples 1-8) were analyzed in this study. These represent two replicates from each of four biological samples (1A replicate 1, 1B replicate 2, 3A replicate 1, 3B replicate 2, 5A replicate 1, 5B replicate 2, 9A replicate 1, and 9B replicate 2).  Two control samples (1A replicate 1 and 1B replicate 2) were included as control samples.


title: Differential transcriptomes of P. aeruginosa PAO1 upon continuous exposure to antibiotic-impregnated catheters

city: Jefferson

first: Kidon

last: Sung

state: AR

address: 3900 NCTR Road

institute: National Center for Toxicological Research

summary:
Purpose: The goals of this study are to compare RNA-Seq transcriptomic data of P. aeruginosa PAO1 in response to chronic exposure of clindamycin and rifampicin eluted from antibiotic-impregnated catheters in a continuous biofilm culture condition.
Methods: Transcriptomic profiles of P. aeruginosa PAO1 grown with catheters without antibiotic coating or with clindamycin/rifampicin coating were generated by deep sequencing, in quadruplicates, using Illumina NextSeq 550 sequencer with SE-75. FastQC and Trimmomatic (v0.39) were used for sequencing quality assessment. Raw read counts of the annotated genes were calculated using HTSeq-count. The log2 fold change and false discovery rate (FDR) were measured by edgeR. Genes with |log2FC| > 1.0 and FDR < 0.05 were considered as differential expression genes. Transcripts per million (TPM) was measured by Kallisto, which is an alignment free expression estimation tool.
Results: P. aeruginosa PAO1 RNA sequencing using Illumina NextSeq 550 sequencer generated 5,205,190 of average number of reads with average read length of 65 bp and a mean coverage of 54X. Differentially regulated proteins of 877 in the planktonic cells were identified, 55 of which displayed up-regulation and 822 displayed down-regulation. On the other hand, 178 proteins were differentially expressed in the biofilm cells containing 152 up-regulated and 26 down-regulated.
Conclusions: Our study represents the first detailed analysis of transcriptomes generated by RNA-seq technology after P. aeruginosa PAO1 planktonic and biofilm cells were continuously exposed to clindamycin/rifampicin-impregnated catheters. Our results show that RNA sequencing offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content of strain PAO1 cells. We conclude that RNA-seq based transcriptome characterization would help the understanding for the mechanism of antimicrobial resistance and virulence in P. aeruginosa after exposure of antimicrobial-coated catheter.

overall_design: Comparison between planktonic and biofilm cells growng with catheters without antibiotic coating or with clindamycin/rifampicin coating

first: Dan

last: Li

first: Ohgew

last: Kweon

first: Joshua

first: Minje

last: Kim

first: Miseon

last: Park

first: Saeed

last: Khan

first: Carl

last: Cerniglia


title: Non-canonical STAT3 activity sustains pathogenic Th17 proliferation and cytokine response to antigen

city: Pittsburgh

first: Amanda

middle: Catherine

last: Poholek

state: Pennsylvania

address: 4401 Penn Ave., Rangos Research Center 8129

department: Pediatrics

institute: University of Pittsburgh

overall_design: Th17 WT (5 mice total, combined from two separate experiments) and Th17ΔSTAT3 (4 mice total combined from two separate experiments) were immunized with MOG(35-55) in CFA to induce EAE. On day 10 post-immunization dLN were harvested and CD4+YFP+ Th17 cells were sorted on a FACSAria II, directly into SmartSeq low-input RNA kit lysis buffer. Libraries were prepared using Nextera XT DNA library prep kits, RNA-Sequencing was performed on Illumina NextSeq500 by the Health Sciences Sequencing Core at University of Pittsburgh.

first: Mandy

last: McGeachy

first: Itay

last: Raphael


title: RNA-targeting Cas9 corrects molecular and physiological features in pre-clinical model of myotonic dystrophy type 1

city: La Jolla

first: Gene

last: Yeo

state: CA

address: 2880 Torrey Pines Scenic Dr. Room 3805/Yeo Lab

institute: UCSD

summary: In this Study, we used RNA-targeting Cas9 (RCas9) to reverse characteristic Myotonic Dystrophy (DM1) cellular phenotypes such as elimination of RNA foci, MBNL relocalization, and reversal of transcriptome-wide splicing in a mouse model of myotonic Dystrophy (DM1). Furthermore we show that gene expression is not altered with RCas9 treatment in WT mice with or without treatment with immunosuppression

overall_design: We performed paired-end RNA-seq for either non-targeting (NT) and CTG-targeting (RCas9-CTG or PIN ) treated tibialis anterior muscles of myotonic dystrophy (DM1) mice called HSALR. Furthermore, we treated WT tibialis anterior muscle with either sgRNA only (G1), RCas9-CTG (G2) or RCas9-CTG along with immunosuppression with Tacrolimus and CTLA4-Ig. Following this we performed paired-end RNA-Seq

first: Ranjan

last: Batra


title: RNA sequencing of late juvenile whole mouse wildtype and Adad2 mutant testis

city: New Brunswick

first: Elizabeth

last: Snyder

state: NJ

address: 59 Dudley Rd Foran Hall 328

department: Animal Science

institute: Rutgers University

summary: Adenosine-to-inosine RNA editing, a fundamental RNA modification, is regulated by adenosine deaminase (AD) domain containing proteins. Within the testis, RNA editing is catalyzed by ADARB1 and is regulated in a cell-type dependent manner. This study examined the role of two testis-specific AD domain proteins, ADAD1 and ADAD2, on testis RNA editing and male germ cell differentiation. ADAD1, previously shown to localize to round spermatids, and ADAD2 had distinct localization patterns with ADAD2 expressed predominantly in mid- to late-pachytene spermatocytes suggesting a role for both in meiotic and post-meiotic germ cell RNA editing. AD domain analysis showed the AD domain of both ADADs was likely catalytically inactive, similar to known negative regulators of RNA editing. To assess the impact of Adad mutation on male germ cell RNA editing, CRISPR-induced alleles of each were generated in mouse. Mutation of either Adad resulted in complete male sterility with Adad1 mutants displaying severe teratospermia and Adad2 mutant germ cells unable to progress beyond round spermatid. However, mutation of neither Adad1 nor Adad2 impacted RNA editing efficiency or site selection. Taken together, these results demonstrate ADAD1 and ADAD2 are essential regulators of male germ cell differentiation with molecular functions unrelated to A-to-I RNA editing.

overall_design: RNA sequencing from 25 dpp whole testis from wildtype and Adad2 mutant C57Bl6J

first: Robert

last: Braun


title: EDMD-causing emerin mutant myogenic progenitors exhibit impaired differentiation using similar mechanisms

city: Bethlehem

first: Ashvin

last: Iyer

address: 1325B Johnston Drive

department: Pharmaceutical Sciences

institute: University of the Sciences

summary: RNA sequencing was performed on proliferating and differentiating emerin-null myogenic progenitors expressing emerin and EDMD-causing emerin mutants to identify molecular pathways implicated in Emery-Dreifuss Muscular Dystrophy.

overall_design: Wildtype and EMD-/y H2K myogenic progenitors were seeded in 6-well dishes coated with 0.01% gelatin at a density of 232,000 cells/well for isolation of total RNA during differentiation. 297,000 cells /well of vector control and emerin mutant cell lines were plated in 6-well dishes coated with 0.01% gelatin. Cells were incubated overnight in proliferative media. In the morning, cell lines were switched to differentiation media. RNA was isolated after 0 hours (to account for changes due to cell density), 24 hours, 48 hours, and 72 hours of differentiation. RNA was isolated using the mRNeasy Plus Kit (Qiagen, Germantown, MD. USA, #74136) per manufacturer’s instructions. Three technical replicates per sample were performed for each biological replicate (n=2).

first: James

last: Holaska


title: RNA sequencing of murine conventional dendritic cell lineage 1 (cDC1) cells from mesenteric lymph nodes

first: Søren

last: Brunak

address: Blegdamsvej 3B

department: Novo Nordisk Foundation Center for Protein Research

summary: A gene expression profile of the conventional dendritic cell lineage 1  from murine mesenteric lymph nodes (MLN) were constructed based on RNA-seq from nine biological biopsies. The gene expression profile was afterwards used to infer (in silico) a cDC1-specific protein-protein interactome.

overall_design: Nine mesenteric lymph nodes were taken from healthy/untreated female 7-10 weeks old C57BL6 mice and CD11c+MHCIIhiXCR-1+CD103+ cDC1 cells were flow cytometry cell sorted. 50bp single-read mRNA sequencing was performed on the cDC1 cells from nine MLNs on illumina machines, resulting in nine FASTQ samples.

first: Kristoffer

last: Niss

first: Cristina

last: Gomez-Casado

first: Jessica

middle: X

last: Hjaltelin

first: Thorsten

last: Joeris

first: William

middle: W

last: Agace

first: Kirstine

middle: G

last: Belling


title: BF_LN_OVS1_NS

city: Seattle

first: Matthew

middle: Price

last: Wood

state: WA

address: 307 Westlake Ave North

department: Global Infectious Diseases

institute: Seattle Children's Research Institute

summary: To assess changes in immune gene expression following HIV-env vaccination through 3 delivery systems,  rhesus macaques were vaccinated with SOSIP trimer with adjuvants using either oral, intraepithelial delivery through a syringe (IEP), a needle free pressurized oral delivery (NF), or an intramuscular injection in the right deltoid (IM). Submandibular lymph nodes were biopsied and stored in RPMI supplemented with 10% fetal bovine serum. RNA was extracted using Macherey Nagel nucleospin columns. RNA from both timepoints was then used for gene expression analysis using the Nanostring nCounter platform.

overall_design: Data from 9 adult macaques was obtained from samples representing time points 2 weeks prior to, and 1 week following, vaccination

first: Donald

last: Sodora

first: Bridget

last: Fisher


title: Brg1 restrains the pro-inflammatory properties of ILC3s and modulates intestinal immunity [ATAC-seq]

first: Ju

last: Qiu

address: 320 Yueyang Road

institute: Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences

summary: To determine the molecular regulation of ILC3s by Brg1, small intestinal ILC3s (Lin-Thy1highCD45low) from Brg1-deficient and control ILC3s were subjected to assay for transposase-accessible chromatin with high throughput sequencing (ATAC-seq).

overall_design: Biological triplicates of 50000 ILC3s (Thy1.2highCD45low cells) sorted by flow cytometry from the small intestine of 7-9 week old Rag1-/-;Smarca4f/f or Rag1-/-;Rorc-cre;Smarca4f/f mice were subjected to ATAC-seq analysis.

first: Xinyi

last: Qi

first: Jinxin

first: Jun

last: Qin


title: miRNA and transcriptome profiling of oropharyngeal tissue from chronically SIV infected rhesus macaques

city: San Antonio

first: Mahesh

last: Mohan

state: Texas

address: 8715 West Military Road

department: Host Pathogen Interaction Program

institute: Southwest National Primate Research Center

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

platform_organism:
Macaca mulatta
Homo sapiens


title: Lumbar intervertebral disc mRNA sequencing identifies the regulatory pathway in patients with disc herniation and spondylolisthesis

first: Asha

last: Nair

address: 200 1ST ST SW

institute: Mayo Clinic

summary: Here, we show results from next generation RNA sequencing (RNA-seq) on mRNA isolated from 10 human nucleus pulposus (NP) samples of lumbar degenerated discs (DH and DS; n=5 for each tissue) and other musculoskeletal tissues (Bone, cartilage, growth plate, and muscle; n=7 for each tissue). Pathway and network analyses based on gene ontology (GO) terms were used to identify the biological functions of differentially expressed mRNAs. A total of 701 genes were found to be significantly upregulated in lumbar NP tissue compared to other musculoskeletal tissues. These differentially expressed mRNAs were primarily involved in DNA damage, immunity and G1/S transition of mitotic cell cycle. Interestingly, DH-specific signaling genes showed major network in chemotactic (e.g., CXCL10, CXCL11, IL1RL2 and IL6) and matrix-degrading pathway (e.g., MMP16, ADAMTSL1, 5, 8, 12, and 15), while DS-specific signaling genes were found to be those involved in cell adhesion (e.g., CDH1, EPHA1and EFNA2) and inflammatory cytokines (e.g., CD19, CXCL5, CCL24, 25 and XCL2). Our findings provide new leads for therapeutic drug discovery that would permit optimization of medical or pharmacological intervention for cases of lumbar DDD.

overall_design: A total of 10 intervertebral disc specimens were collected for research use from 10 adult patients undergoing lumbar discectomy. Patients in this study underwent surgery for the treatment of symptomatic degenerative disk disease presenting with disc herniation or with spondylolisthesis.The specimens used in this investigation were collected under institutional review board approved protocols (IRB# 14-008177).

first: Mohamad

last: Bydon

first: Andre

last: van Wijnen


title: PAI-1 derived cancer-associated fibroblasts in the esophageal squamous cell carcinoma promotes the migration and invasion of cancer cells and macrophages by interaction with LRP1

city: Kamakura

first: Satoshi

last: Kondo

state: Kanagawa

address: 10-1 tebiro 6-chome

country: Japan

institute: Toray Industries,Inc.

summary:
Cancer-associated fibroblasts (CAFs) are known to be involved in the progression of various cancers. However, the roles of CAFs in the esophageal squamous cell carcinoma (ESCC) microenvironment remain unclear. We previously co-cultured human bone marrow-derived mesenchymal stem cells (MSCs) with ESCC cells and confirmed the induction of fibroblast activation protein (FAP) in MSCs, which we defined as CAF-like cells. To investigate the roles of CAFs, we performed cDNA microarray analysis between MSCs and CAF-like cells and found that SERPINE1 was highly expressed in CAF-like cells when compared to MSCs.
Plasminogen activator inhibitor-1 (PAI-1), encoded by SERPINE1, is generally accepted to play tumor promoting roles and act as a poor prognostic indicator in several cancers. PAI-1 derived from CAF-like cells promoted the cell migration and invasion of ESCC cells by activating Akt and Erk1/2 signaling pathways via low-density lipoprotein receptor related protein 1 (LRP1), the receptor of PAI-1. The higher expression levels of PAI-1 and LRP1 were correlated with the poor prognosis in ESCC patients. These results suggest that PAI-1/LRP1 axis contributes to the progression of ESCC.

overall_design: In order to induce the CAF-like cells, we established co-culture assay using mesenchymal stem cells (MSCs) and ESCC cell lines (TE-8, TE-9 and TE-15) and  defined MSCs co-cultured with TE-8, TE-9 and TE-15 cells as CAF8, CAF9 and CAF15 cells (CAF-like cells), respectively.  MSCs were seeded in a same manner without co-culture as control. We performed cDNA microarray analysis between MSCs and CAF-like cells.

first: H

last: Sakamoto

first: Y

middle: I

last: Koma


title: Microglial remodeling of the extracellular matrix promotes synapse plasticity

city: San Francisco

first: Leah

last: Dorman

address: 499 Illinois St

department: Data Science

institute: CZBiohub


title: Exosomal microRNAs secreted by induced pluripotent stem cell-derived cardiac cells

city: Aurora

first: Edward

last: Lau

state: CO

address: 12700 E19th Av RCII Rm 8004

department: Medicine/Cardiology

institute: University of Colorado

summary: We report the secreted microRNAs from extracellular vesicles released from (i) cardiomyocytes, (ii) cardiac fibroblasts, (iii) endothelial cells differentiated from identical lines of normal human induced pluripotent stem cells and (iv) undifferentiated induced pluripotent stem cells.

overall_design: Comparison of microRNA expression secreted from four isogenic cell types; 2 technical replicates each of 2 biological replicates each.

first: Mark

last: Chandy

first: June

last: Rhee

first: Joseph

middle: C

last: Wu


title: Microglial remodeling of the extracellular matrix promotes synapse plasticity [Microglia]

summary: This study defines a molecular interaction between neurons and microglia that drives experience-dependent synapse remodeling in the hippocampus.

overall_design: Microglia isolated from mouse hippocampi 4 hours after injection of either PBS (control) or IL-33 were RNA sequenced.

first: Phi

middle: T

last: Nguyen

first: Ilia

middle: D

last: Vainchtein

first: Anna

middle: V

last: Molofsky


title: Transcriptome profiling of Oropharyngeal tissue from chronically SIV infected rhesus macaques

summary: HIV/SIV associated oral mucosal disease/dysfunction (HAOMD) (gingivitis/periodontitis/salivary adenitis) represents a major comorbidity affecting HIV patients on anti-retroviral therapy. Using a systems biology approach, we investigated molecular changes (mRNA/microRNA) underlying HAOMD and its modulation by phytocannabinoids [delta-9-tetrahydrocannabinol (Δ9-THC)] in uninfected (n=5) and SIV-infected rhesus macaques untreated (VEH-untreated/SIV; n=7) or treated with vehicle (VEH/SIV; n=3) or Δ9-THC (THC/SIV; n=3). Relative to controls fewer mRNAs were upregulated in THC/SIV compared to VEH-untreated/SIV macaques. Gene enrichment analysis showed differential enrichment of biological functions involved in anti-viral defense, Type-I interferon, Toll-like receptor, RIG-1 and IL1R signaling in VEH-untreated/SIV macaques. We focused on the anti-ER-stress anterior gradient-2 (AGR2), epithelial barrier protecting and anti-dysbiotic WAP Four-Disulfide Core Domain 2 (WFDC2), and glucocorticoid-induced anti-inflammatory TSC22D3 (TSC22-domain family member 3) that were significantly downregulated in OPM of VEH-untreated/SIV macaques. All three proteins localized to minor salivary gland acini and secretory ducts and showed enhanced and reduced expression in OPM of THC/SIV and VEH/SIV macaques, respectively. Additionally, inflammation associated miR-21, miR-142-3p and miR-29b showed significantly higher expression in OPM of VEH-untreated/SIV macaques. TSC22D3 was validated as a target of miR-29b. These preliminary translational findings suggest that phytocannabinoids may safely and effectively reduce oral inflammatory responses in HIV/SIV and other diseases.

overall_design: Oropharyngeal tissue mRNA profiles of uninfected control (n=5) and chronically SIV-infected rhesus macaques that were vehicle-untreated (n=7) or treated with delta-9-tetrahydrocannabinol were generated by deep sequencing using Illumina.


title: Microglial remodeling of the extracellular matrix promotes synapse plasticity [Neuronal nuclei]

overall_design: Single-cell (Chromium) sequencing of neuronal nuclei from the mouse hippocampus


title: RKIP regulates differentiation-related features in melanocytic cells

city: Leioa

first: cristina

last: penas lago

state: Vizcaya

address: Barrio Sarriena

institute: UPV/EHU

summary: Raf Kinase Inhibitor Protein (RKIP) has been extensively reported as an inhibitor of key signaling pathways involved in the aggressive tumor phenotype and shows decreased expression in several types of cancers. However, little is known about RKIP in melanoma or regarding its function in normal cells. We examined the role of RKIP in both primary melanocytes and malignant melanoma cells and evaluated its diagnostic and prognostic value. IHC analysis revealed a significantly higher expression of RKIP in nevi compared with early-stage (stage I-II, AJCC 8th) melanoma biopsies. Proliferation, wound healing, and collagen-coated transwell assays uncovered the implication of RKIP on the motility but not on the proliferative capacity of melanoma cells as RKIP protein levels were inversely correlated with the migration capacity of both primary and metastatic melanoma cells but did not alter other parameters. As shown by RNA sequencing, endogenous RKIP knockdown in primary melanocytes triggered the deregulation of cellular differentiation-related processes, including genes (i.e., ZEB1, THY-1) closely related to the EMT. Interestingly, NANOG was identified as a putative transcriptional regulator of many of the deregulated genes, and RKIP was able to decrease the activation of the NANOG promoter. As a whole, our data support the utility of RKIP as a diagnostic marker for early-stage melanomas. In addition, these findings indicate its participation in the maintenance of a differentiated state of melanocytic cells by modulating genes intimately linked to the cellular motility and explain the progressive decrease of RKIP often described in tumors.

overall_design: Human melanocytes were transduced with RKIP shRNA Lentiviral Particles (sc-36430-V, 2MOI) or Control shRNA Lentiviral Particles (sc-108080 2MOI).

first: Cristina

last: Penas

first: Aintzane

last: Apraiz

first: Iraia

last: Muñoa

first: Yoana

last: Arroyo-Berdugo

first: Javier

last: Rasero

first: Pilar

last: Ezkurra

first: Veronica

last: Velasco

first: Nerea

last: Subirán

first: Anja

middle: K

last: Bosserhoff

first: Santos

last: Alonso

last: Asumendi

first: María

last: Boyano


title: MicroRNA profiling of Oropharyngeal tissue from chronically SIV infected rhesus macaques

first: Mahesh

last: Mohan

summary: The study describes miRNA expression in Oropharyngeal tissue of chronically SIV-infected rhesus macaques. To identify the underlying molecular mechanisms  we simultaneously profiled miRNA and mRNA expression in oropharyngeal tissues of chronically simian immunodeficiency virus (SIV)-infected rhesus macaques (RMs). Relative to controls, we identified 48 (38-upregulated and 10 downregulated) differentially expressed (DE) miRNAs relative to uninfected controls (n=5). Interestingly, in terms of magnitude, miR-19a, miR-301, miR-142-3p, miR-32 and miR-142-5p were among a select list of miRNAs that showed the highest upregulation in OPM. An important finding is the significant upregulation in OPM of miR-21, a microRNA known to regulate periodontitis, T-cell activation and oral carcinoma. Interestingly, RNA-seq for gene expression profiling also confirmed miR-21 upregulation in OPM of VEH-untreated/SIV rhesus macaques. Using TargetScan 7.2, we identified TSC22D3 (TSC22-domain family member 3), an anti-inflammatory protein induced by glucocorticoids and IL10 that was significantly downregulated in OPM of VEH-untreated/SIV macaques to be a predicted target of miR-29b. TSC22D3 localized to minor salivary gland acini and secretory ducts and showed  reduced expression in OPM of  VEH-untreated chronically SIV macaques. Using luciferase reporter and overexpression assays, we confirmed TSC22D3 to be a direct target of miR-29b. Interestingly, expression of miR-150 was significantly downregulated, a miRNA we previously demonstrated to be downregulated during T cell activation in the intestine. These findings strongly support a role for differential miRNA expression associated with HIV/SIV induced oropharyngeal mucosal dysfunction.

overall_design: Twelve age and weight matched male Indian rhesus macaques were randomly divided into 2 groups. Group 1 (n=5) remained uninfected. Group 2 (VEH-untreated/SIV, n=7) animals were infected intravenously with 100TCID50 of SIVmac251. Oropharyngeal tissue was collected at necropsy, ~180 days post SIV infection. ~100 ng of total RNA was first reverse transcribed and preamplified according to the manufacturer’s recommendation. MicroRNA expression profiling was performed using TaqMan ®OpenArray® Human microRNA panels. Data analysis was performed using ExpressionSuite® software, which utilizes the comparative Cτ (ΔΔCτ) method to rapidly and accurately quantify relative miRNA expression across many genes and samples. miRNA expression data were normalized using a combination of snU6, RNU44 and RNU48.  Comparisons were made between uninfected and VEH-untreated SIV-infected rhesus macaques.

first: M


title: Platelet miRNA bio-signature discriminates between dementia with Lewy bodies and Alzheimer’s disease

city: Barcelona

first: Ana

middle: Gámez

last: Valero

state: Barcelona

address: Universitat de Barcelona, Casanova 143, 4th Floor-Nord

institute: University of Barcelona (Neuroscience Institute)

summary: Dementia with Lewy bodies (DLB) is the second most common cause of degenerative dementia after Alzheimer’s disease (AD) and presents several overlapping pathological and clinical features with both AD and Parkinson’s disease (PD). Consequently, only one in three DLB cases is diagnosed correctly. Platelets are anucleate cells containing cellular structures and molecules, such as microRNA (miRNA), the analysis of which provides biomarkers for several disorders. In this cross-sectional study, we profiled the whole platelet miRNA transcriptome from 7 DLB patients and 7 healthy controls using Next-Generation Sequencing (NGS) and found 22 differentially expressed miRNAs. These miRNAs were further validated with miRCURY LNA miRNA Custom PCR panels in three consecutive validation studies carried out from 2017-2019 with a total of 162 individuals including DLB (n=59), AD (n=28), and PD patients (n=24), and healthy controls (n=51). As a result, three different groups of miRNAs were identified as related to DLB, AD and PD, all in comparison with controls. Additionally, our results demonstrated a significant down-regulation of hsa-miR-26b-5p and hsa-miR-150-5p in DLB compared to PD; and a signature composed of seven miRNAs that had lower expression in DLB compared to AD. The predictive diagnostic value of this bio-signature for the differentiation of DLB from AD was assessed calculating the corresponding ROC curve, which yielded an area under the curve of 1.00. Predictive analyses using specialized software (miRTarbase, miRGate) were performed for the disease-specific miRNAs to identify target genes and corresponding pathways. Three disease-specific clusters of pathways and biological processes were identified as a result of platelet-miRNA deregulation. In DLB, pathways related to gene expression and small RNA metabolism; in AD, pathways related to stress response and RNA stress granules; and in PD pathways related to protein phosphorylation, and protein metabolism and degradation were identified. In conclusion, we describe the identification of a novel, highly specific and sensitive platelet-associated miRNA-based bio-signature, which permits us to distinguish between DLB and AD.

overall_design: miRNA-seq of platelets from DLB patients and healthy controls. PRJNA613191

last: Gámez-Valero

first: Jaume

last: Campdelacreu

first: Dolores

last: Vila

first: Lourdes

last: Ispierto

first: Daniela

last: Samaniego

first: Jordi

last: Gascón-Bayarri

first: Ramón

last: Reñé

first: Ramiro

last: Álvarez

first: Maria

middle: P

last: Armengol

first: Francesc

last: Borràs

first: Katrin

last: Beyer


title: BF_BUC_OVS1_NS

first: Matthew

last: Wood

summary: To assess changes in immune gene expression following HIV-env vaccination through 3 delivery systems,  rhesus macaques were vaccinated with SOSIP trimer with adjuvants using either oral, intraepithelial delivery through a syringe (IEP), a needle free pressurized oral delivery (NF), or an intramuscular injection in the right deltoid (IM). Buccal mucosal samples were collected by cytobrush and stored in Nucleospin lysis buffer RA1 with 1% β-mercaptoethanol and RNA was extracted using Macherey Nagel nucleospin columns. RNA from both timepoints was then used for gene expression analysis using the Nanostring nCounter platform.

overall_design: Data from 6 HIV-env vaccinated adult macaques was obtained from samples representing the 1 week post-vaccination time point

first: Donald

last: Sodora

first: Bridget

last: Fisher


title: Differential transcriptional responses to cardiomyocyte-specific Notch activation in right versus left atria of murine cardiac tissue

city: St Louis

first: Tiandao

last: Li

state: MO

address: 4444 Forest Park Ave

institute: Washington University

summary: We report the mRNA sequencing of right and left atria from an adult inducible, cardiomyocyte-specific Notch gain-of-function mouse model (iNICD). Using  the tetracycline-on system, we activated Notch by feeding doxycycline chow for 3 weeks to mice that were at least 8 weeks old. We asked what transcriptional changes occur in right versus left atrial cardiomyocytes in response to the same stimulus (Notch signaling). mRNA sequencing on separated right and left atria revealed that there are more differentially dysregulated transcripts (1,011) than similarly regulated transcripts (447) in the right and left atria, which is a simiar paradigm as what occurs in human atrial cardiomyocytes of patients with atrial fibrillation.

overall_design: Total mRNA profiles for mouse heart samples (RA, LA) after cardiomyocyte-specific Notch activation as adults.

first: Stacey

middle: L

last: Rentschler


title: Inherited epigenetic repression of the Celf2a splicing factor mediates the amelioration of a Duchenne phenotype

first: Enrique

last: Blanco

address: Dr. Aiguader 88

department: Gene Regulation, Stem Cells and Cancer

institute: Center for Genomic Regulation (CRG)

summary: Exon skipping is an effective strategy for the treatment of Duchenne Muscular Dystrophy (DMD). Natural exon skipping identified in several DMD cases can help with identifying novel therapeutic tools. Here, we demonstrate that CRISPR/Cas9 inactivation of the splicing factor Celf2a in a DMD-D44 background induces skipping of exon-45 and dystrophin rescue. Celf2a ablation could be compatible with life and curative since a DMD case with a milder symptomatology exists where exon-45 skipping occurs because of a Celf2a deficiency. We show that in this individual, Celf2a absence is due to inherited epigenetic silencing and that its repression is controlled by the lncRNA DUXAP8.

overall_design: ChIP-seq (8 samples) and ATAC-seq (4 samples) of WT (healthy) and GSΔ44 myoblasts (see Pubmed ID 26796035)

first: Julie

last: Martone

first: Valerio

last: Di Carlo


title: Circulatory proteins permeate the brain via selective transport impaired with age

city: Palo Alto

first: Andrew

last: Yang

address: 3801 Miranda Ave

institute: Stanford University

summary: 10x single-cell RNA-sequencing to profile the transcriptomic response of the brain endothelium to pharmacologic ALPL inhibition

overall_design: Transcriptomic response of the brain endothelium to pharmacologic ALPL inhibition

first: Tony

last: Wyss-Coray


title: Mesenchymal subtypes of hepatocellular carcinoma

city: Houston

first: Sangbae

last: Kim

state: TX

address: 1 Baylor plaza

department: HGSC

institute: Baylor Colleage of Medicine

summary: Clinical heterogeneity of hepatocellular carcinoma (HCC)  reflected in unequal outcome of treatment is poorly defined in molecular level, and molecular subtypes and their associated biomarkers have not been established to improve prognostification and treatment of HCC. Using microarray technologies, we analyzed gene expression profiling data from HCC patients, uncovered mesenchymal subtype, and identified gene expression signature associated with mesenchymal phenotype of HCC.

overall_design: We generated gene expression data from tumors of 110 HCC patients who underwent surgical resection as primary treatment

first: Ju-Seog

last: Lee


title: H2BG53D mutation in PDAC

city: Hong Kong

first: Ming

middle: Kui

last: Chan

address: Tat Chee Avenue

department: Department of Biomedical Sciences

country: Hong Kong

institute: City University of Hong Kong

summary: Study was conducted to understand the effect of the cancer associated H2BG53D mutation in PDAC. Using CRISPR/Cas9 Knock-in cell lines expressing FLAG-tagged WT and G53D mutant H2B, we conducted gene expression profilling experiments and studied the effect of the H2BG53D mutation on transcription and PDAC development.

overall_design: CRISPR/Cas9 cell lines expressing FLAG tagged Wildtype (WT) H2B and H2BG53D were generated to study the H2BG53D mutation. Two WT and two mutant G53D cell lines as well as the Parental S2VP10 cell lines were used for experiments. Gene expression of WT and G53D mutant lines was profiled by RNA-seq. Genomic localization of FLAG-tagged WT and G53D H2B proteins were profiled by CUT&RUN sequencing. H2A CUT&RUN experiment showed H2A occupancy. Pol 2 occupancy in the CRISPR cell lines were profiled by PRO-seq.

first: Yi Ching Esther

last: Wan

first: Lina

last: Zhu

first: Kui Ming


title: Next Generation Sequencing Facilitates Quantitative Analysis of NC and miR-200ba KD Mice MOE Transcriptomes

city: Baoding

first: Dong

state: Hebei

address: No. 180, Wusi Road

institute: Hebei university

summary: In order to explore the underlying molecular mechanisms of the impaired olfactory behaviors demonstrated in mice with knockdown of miR-200b/a in the MOE, we dissected the MOE tissues of the miR-200b/a knockdown mice and their NC controls for RNA transcriptome analysis.

overall_design: MOE mRNA profiles of 5-month old male NC and miR-200b/a knockdown mice were generated by deep sequencing, in triplicate, using Illumina platform PE150.

first: Zhenshan

last: Wang


title: Identification of BBX proteins as rate-limiting co-factors of HY5

city: Berlin

first: Henrik

last: Johansson

address: Albrecht-Thaer-Weg 6

institute: Freie Universität Berlin

summary: As a source of both energy and environmental information, the importance for plants to monitor the incoming light is evidenced by the highly integrated nature of the light signal transduction pathway with numerous other pathways throughout plant development. One of these signal integrators is the bZIP transcription factor HY5 which holds a key role as a positive regulator of light signalling in plants. Although HY5 is described to act as a DNA-binding transcriptional regulator, the lack of any apparent transactivation domain makes it unclear how HY5 is able to accomplish its many functions. We describe the identification of three B-box containing proteins (BBX20, 21 and 22) as essential partners for HY5 dependent modulation of hypocotyl elongation, anthocyanin accumulation and transcriptional regulation. The bbx202122 triple mutant mimics the phenotypes of hy5 in the light and its strong suppression of the cop1 phenotype in darkness. Furthermore, RNA-seq experiments show that 84% of genes differentially regulated in bbx202122 are also HY5 regulated, consistent with the B-box proteins and HY5 acting interdependently.

overall_design: Investigation of transcriptomic changes in bbx202122 and hy5 Arabidopsis seedlings grown for 4 days in red light, in triplicate, using RNA-seq analysis.

first: Katharina

last: Bursch


title: Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and AC3-/- Mice MOE miRNAs Expression

summary: To explore the correlation between olfactory function and the miRNAs expression, we sequenced small RNAs from MOE tissues dissected from 3 month old male AC3-/- and AC3+/+ mice.

overall_design: MOE miRNA profiles of 3-month old male wild type (WT) and AC3-/- mice were generated by deep sequencing, in twice, using HiSeq 2500.


title: Brg1 restrains the pro-inflammatory properties of ILC3s and modulates intestinal immunity [RNA-seq]

city: Shanghai

first: Ju

last: Qiu

summary: In order to search for cell-intrinsic targets for Brg1 in ILC3s, Brg1-deficient and control ILC3s were sorted from bone marrow chimeric mice, generated by transfering mixed bone marrow cells of Thy1.1/Thy1.2Rag1-/-Smarca4f/f and Thy1.2Rag1-/-Smarca4f/fRorc-cre donors at 1:1  ratio to sub-lethally irriadiated Rag2-/-Il2rg-/- recipients.

overall_design: 2.5 million bone marrow cells from Thy1.2Rag1-/-Smarca4f/f and Thy1.1Thy1.2Rag1-/-Smarca4f/fRorc-cre mice respectively were mixed and transferred to sub-lethally irradiated Rag2-/-Il2rg-/- mice. Small intestinal lamina propria lymphocytes were isolated from the chimeric Rag2-/-Il2rg-/- mice 6-7 weeks later. ILC3s from the Thy1.2Rag1-/-Smarca4f/f origin (Thy1.2highThy1.1-CD45low) and the Thy1.1Thy1.2Rag1-/-Smarca4f/fRorc-cre origin (Thy1.2highThy1.1+CD45low) were sorted by flow cytometry. mRNA was extracted by Trizol reagent and RNA-seq analyses were performed.

first: Xinyi

last: Qi

first: Yan

last: Ji

first: Jun

last: Qin


title: A universal, benchtop, gel-free method for the rapid and simultaneous isolation of all known classes of functional small RNAs

first: Stefan

last: Oberlin

address: 505 Parnassus Ave

department: Department of Microbiology and Immunology

institute: University of California, San Francisco

summary: All known silencing small (s)RNAs operate via ARGONAUTE(AGO)-family proteins within RNA-induced-silencing-complexes (RISCs). Based on AGOs conserved biochemical properties, we have developed a universal, 15-min benchtop extraction procedure allowing simultaneous purification of all classes of RISC-associated sRNAs known to date, without prior knowledge of the samples-intrinsic AGO repertoires. Optimized into a user-friendly kit, the method –coined “TraPR” for Trans-kingdom, rapid, affordable Purification of RISCs– operates irrespectively of the organism, tissue, cell type or bio-fluid of interest, including from minute amounts of input material. The method is highly suited for direct sRNA deep-sequencing, with TrAPR-generated libraries being qualitatively and quantitatively at least on-par with those obtained via gold-standard procedures involving tedious polyacrylamide gel excisions. TraPR considerably improves the quality and consistency of sRNA sample preparation including from notoriously difficult-to-handle tissues/bio-fluids such as starchy storage roots and mammalian plasma, and regardless of RNA contaminants or samples’ RNA-degradation status.

sample_organism:
Arabidopsis thaliana
Drosophila melanogaster
Mus musculus

overall_design: Trans-kingdom, rapid, affordable Purification of RISC (TraPR) protocol for sRNA isolation was compared to standard sRNA library preparation from total RNA. A variety of source tissue, like Arabidopsis flower buds, Drosophila ovaries and mouse livers and plasa were used for these comaparisons.

first: Thomas

last: Grentzinger

first: Dominik

last: Handler

first: Julius

last: Brennecke

first: Olivier

last: Voinnet


title: RNA seq of CD4+ T cells from WT and miPEP31-/- mice

city: shanghai

first: Hong

last: Zhou

address: South Chongqing road

department: Immunology

institute: Shanghai Insititute of immunology

summary: To see the differential expression gene between miPEP31-/- and WT mice, we isolated CD4+ T cells and activated by anti-CD3/28. Then performed high-through RNA seq.

overall_design: We isolated CD4+ T cells from WT and miPEP31-/- mice and activated by anti-CD3/28 for 48 hours. Cells were cellected and extration of RNA to high-throughput profiling  seq. The data were analyzed on the free online platform of Majorbio I-Sanger Cloud Platform (www.i-sanger.com)

first: Wei

last: Cai


title: Gene expression profiling in acute liver failure versus healthy human liver

city: Bronx

first: Sanjeev

last: Gupta

address: 1300 Morris Park Ave.

department: Medicine

institute: Albert Einstein College of Medicine

summary:
To understand key pathways and regulatory genes in acetaminophen induced Acute Liver Failure (ALF) for pathophysiological and therapeutic mechanisms
gene expression was analyzed and significantly up and down regulated genes were identified

overall_design: Archival Formalin Fixed Parafin Embedded (FFPE) samples of human subjects with ALF were compared to FFPE samples of normal liver

first: Sriram

last: Bandi

first: Preeti

last: Viswanathan

first: Tatyana

last: Tchaikovskaya


title: RNAseq analysis on E14.5 cortical material from mouse, Tcf4-ko line vs WT littermates.

city: Amsterdam

first: Marten

middle: Piet

last: Smidt

state: NH

address: Sciencepark 904

department: SILS

country: Netherlands

institute: UVA

summary: Tcf4 encodes cortical differentiation during development

overall_design: examination of 2 groups form TCF4ko and WT littermates, N=3 form both. RNA samples from E14.5 Cortical dissected material.

first: M

first: S

last: Mesman


title: Single-cell RNA and electrophysiological profiling of D1+ and D2+ principal neurons of the prefrontal cortex

city: Zurich

first: Csaba

last: Földy

state: Zurich

address: Winterthurerstrasse 190

department: Brain Research Institute

institute: University of Zurich

summary: Dopaminergic modulation of the prefrontal cortex (PFC) circuits is crucial for cognitive processes such as working memory, planning, attention and dysfunction. This system has been linked to cognitive deficits associated with schizophrenia and autism spectrum disorders. Despite its physiological and pathophysiological importance, we are still lacking a clear understanding of the basic principles of dopamine (DA) actions in the PFC. We set out to provide a detailed classification of layer 5 pyramidal neurons mediating the two main streams of DA signaling, namely via DA receptor group 1 (D1) and DA receptor group 2 (D2). To achieve this, we combined electrophysiological whole-cell recordings in acute slices of the prelimbic part (PL) of the medial PFC with single cell RNA sequencing, morphological and immunohistochemical analysis.

overall_design: 15 D1 cells/ 26 D2 cells for the WT condition and 6 D1 cells and 7 D2 cells for the KO condition (Fmr1-/y mice; this is a mouse model for fragile x syndrome and autism spectrum disorder)

first: Rafael

last: De Sa

first: Jochen

last: Winterer

first: Katy

last: Le Corf

first: David

last: Lukacsovich

last: Foldy

first: Andreas

last: Frick


title: Gene expression after treatment in a 3D lung tumor model in comparison to conventional 2D cell culture

city: Würzburg

first: Claus

middle: Juergen

last: Scholz

address: Josef-Schneider-Str. 2

department: Core Unit SysMed

institute: University Hospital Würzburg

summary: Gene expression of a 3D model for the treatment of EGFR mutated lung tumors (HCC827 cell line, sensitive to gefitinib, and A549 cells, insensitive to gefitinib) was analysed and compared to the same treatment effects in the cell lines when cultured under 2D conditions.

overall_design: Global expression profiling of two culture conditions, two treatment groups in two cell lines differing in EGFR activating mutation status.

first: Matthias

last: Schmitt

middle: J

first: Heike

last: Walles

first: Gudrun

last: Dandekar

first: Sarah

last: Nietzer


title: To determine whether interference with mechanisms of vasoconstriction or inflammation would improve outcomes in ischemia reperfusion injury of the liver

first: Sanjeev

last: Gupta

summary: Study was performed in groups of animals including healthy controls, ischemia reperfusion alone, and administration of drug followed by ischemia reperfusion. Samples from liver and isolated hepatocytes intact and one day after reperfusion were analyzed for gene expression in conjunction with other biological variables. Results indicated substantive alteration after drug intervention in multiple gene ontology groups and cellular events and processes after ischemia reperfusion.

overall_design: Three groups of animals were established: healthy control; ischemia reperfusion alone by temporaly clamping of the portal vein; administration of Etanercept 16-18 hrs before ischemia reperfusion and of nitroglycerin as intravenous infusion before and during ischemia reperfusion. Liver samples were obtained from n=3 per condition. Primary hepatocytes were isolated by two-step collagenase perfusion from additional animals n=3 per condition. One day after ischemia reperfusion, RNA was extracted, pooled for each group and processed for the sequencing on Ion Torrent Proton instrument.

first: Thirunavukkarasu

last: Chinnasamy

first: Fadi

last: Jaber

middle: B

last: Rhee

first: Kevin

middle: R

last: Shieh

first: Wilber

last: Quispe

first: Alexander

middle: Y

last: Maslov

first: Aaron

last: Golden


title: Fibroblasts in Nodular Sclerosing Classical Hodgkin Lymphoma Are Defined by a Specific Phenotype and Protect Tumor Cells From Brentuximab-Vedotin Induced Injury

status: Public on Jun 30 2020

city: Frankfurt

first: Claudia

last: Döring

address: Theodor Stern Kai 7

department: Medical School, Institite of Pathology

institute: Johann Wolfgang-Goethe University Frankfurt a.M.

summary: Several studies have described a crosstalk between the tumour cells of cHL, the Hodgkin- and Reed-Sternberg (HRS) cells, and cancer-associated fibroblasts (CAF). However, to date a deep molecular characterization of these fibroblasts is lacking. Aim of the present study therefore was a comprehensive characterization of these fibroblasts.

overall_design: Gene expression of fibroblasts isolated from primary lymph node suspensions (LA = lymphoadenitis, n=7, cHL_MC = classical Hodgkin lymphoma mixed cellularity subtype, n=5; cHL_NS = classical Hodgkin lymphoma nodular sclerosing subtype, n=8; Fib = primary fibroblasts culture at passage 4-6 in vitro, KB = arbitrary sample ID prefix, FST = arbitrary sample ID prefix). As previously shown for other types of cancer-associated fibroblasts, treatment by luteolin could reverse this fibroblast phenotype. Working concentration of 30 µM luteolin was added to sample KB33 and harvested 48 h post treatment for comparative analysis with Fib_cHL_NS samples. Three biological replicates were applied. The cHL specific adherence-based interaction of both cell fractions revealed an initiation of inflammatory key regulators. The classical Hodgkin lymphoma cell line L-428 and adherent activated fibroblasts cells from primary nodular sclerosis cHL were used. Floating cell fractions were removed by repeated washing. Adherent interacting coculture layer was harvested for comparative gene expression analysis to mock coculture. Mock coculture was set up separated in equivalent ratios and pooled before RNA analysis. Three biological replicates were applied.

first: Katrin

last: Bankov


title: Differential gene expression profiling and functional network analysis of Mycobacterium tuberculosis bioflims and planktonic populations

city: Chang Chun

first: Zhiqiang

last: Yang

state: Jilin

address: No. 5333 Xian Road

department: College of Veterinary Medicine

institute: Jilin University

summary: In order to unearth the marker genes in the formation process of M. tuberculosis biofilm, Illumina sequencing technology was firstly used to catalog global gene expression profiling and functional network analysis of M. tuberculosis bioflim and planktonic populations in the world.

overall_design: RNA profiles of 25-day Mycobacterium tuberculosis mature biofilm and planktonic cells were generated by deep sequencing,in duplicate, using Illumina 2500.

first: Qiaoli

last: Zhang

first: Yang

last: Wang

first: Chao

first: Yingxin

last: Qi

first: Bo

last: Liu

first: Mingyuan

first: Lu

last: Yu


title: A smart polymer for sequence-selective binding, pulldown, and release of DNA targets

city: Dresden

first: Elisha

last: Krieg

state: Saxony

address: Hohe Str. 6

department: Institute of Biofunctional Polymer Materials

institute: The Leibniz Institute of Polymer Research Dresden (IPF)

summary: Selective isolation of DNA is crucial for applications in biology, bionanotechnology, clinical diagnostics and forensics. We herein report a smart methanol-responsive polymer (MeRPy) that can be programmed to bind and separate single- as well as double-stranded DNA targets. Captured targets are quickly isolated and released back into solution by denaturation (sequence-agnostic) or toehold-mediated strand displacement (sequence-selective). The latter mode allows 99.8% efficient removal of unwanted sequences and 79% recovery of highly pure target sequences. We applied MeRPy for the depletion of insulin, glucagon, and transthyretin cDNA from clinical next-generation sequencing (NGS) libraries. This step improved data quality for low-abundance transcripts in expression profiles of pancreatic tissues. Its low cost, scalability, high stability and ease of use make MeRPy suitable for diverse applications in research and clinical laboratories, including enhancement of NGS libraries, extraction of DNA from biological samples, preparative-scale DNA isolations, and sorting of DNA-labeled non-nucleic acid targets.

overall_design: 9 Samples are analyzed: 3 untreated samples that were not depleted with MeRPy; 3 negative control samples that were treated with MeRPy but lacked the target-specific catcher strand library (CSL); 3 experiment samples that were treated with MeRPy and a target-specific CSL.

first: Krishna

first: Andreas

last: Dahl

first: Mathias

last: Lesche

first: Susanne

last: Boye

first: Albena

last: Lederer

first: William

middle: M

last: Shih


title: Genome wide distribution of gammaH2AvD in Wild Type and RNAi dH1 Drosophila melanogaster S2 cells

first: Oscar

last: Reina Garcia

address: C/Baldiri Reixac 10

department: Biostatistics and Bioinformatics

institute: IRB Barcelona

overall_design: ChIP-Seq peak calling of gammaH2AvD against Input sample in Drosophila melanogaster S2 WT and RNAi dH1 cells

first: Aleix

last: Bayona

first: Jordi

last: Bernués

first: Fernando

last: Azorin


title: miRNA atlas of hypertrophy progression in a murine TAC model of experimental Hypertrophy

first: Claus

last: Scholz

summary: miRNA pattern of cardic myocytes, cardiac fibroblasts and cardiac endothelial cells isolated from adult mice hearts at different time points after transaortic constriction (TAC) or sham surgery.

overall_design: Two groups: Sham and TAC; 3 time points: 3day, 2weeks and 4weeks post surgery

first: Seema

last: Dangwal

first: Thomas

last: Thum


title: Sensory neurons expressing calcitonin gene-related peptide regulate adaptive thermogenesis and diet-induced obesity

city: Los Angeles

first: Celine

last: Riera

address: 8700 Beverly Blvd

department: Biomedical Sciences

institute: Cedars-Sinai Medical Center

summary: We report role of CGRP+ neurons in the regulation of themogenesis on High fat diet in adipose tissues. We have used RNA Sequencing approach to identify genes that are differentially expressed in BAT and IWAT  in HFD fed animals regulating thermogenic program.

overall_design: mRNA  sequencing was performed on BAT and IWAT adipose tissue from HFD and Normal chow fed animals (n=3 for each group)

first: Kuldeep

last: Makwana

first: Alexandre

last: Lalande

first: Nancy

last: Morones

first: Harshita

last: Chodavarapu

first: Peter

last: Nguyen

first: Predrag

last: Jovanovic

first: Yidan

last: Wang

middle: E


title: Identification of clonal neoantigens derived from driver mutations in an EGFR mutated lung cancer patient benefitting from anti-PD-1

city: Shenzhen

first: Li

last: Xiaoting

address: 9th Floor, Dabaihui Center, Yantian District, Shenzhen

institute: YuceBio Technology Co., Ltd.

summary: Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) have been recommended as the first line therapy for non-small cell lung cancer (NSCLC) with EGFR mutations. However,  acquired resistance to EGFR-TKIs is inevitable. Although  anti- programmed cell death 1 (PD-1)/PD-ligand (PD-L1) immunotherapies have achieved great clinical success as second-line treatment for many cancer types, the clinical efficacy of anti-PD-1/PD-L1 blockades in EGFR mutated NSCLC patients has been demonstrated to be obviously lower than those without EGFR mutations. Here, we reported an advanced NSCLC patient with exon 19 deletion and T790M EGFR mutation benefitting from anti-PD-1 blockade therapy after acquiring resistance to EGFR-TKI. We characterized the mutational landscape of the patient with next-generation sequencing (NGS), and successfully identified neoantigen-specific T cell clones derived from  EGFR exon 19 deletion, TP53 A116T and DENND6B R398Q mutations. Our findings support the potential application of immune checkpoint blockades in NSCLC patients with acquired resistance to EGFR-TKIs in the context of specific clonal neoantigens with high immunogenicity. Personalized immunomodulatory therapy targeting these neoantigens should be explored for better clinical outcomes in EGFR mutant NSCLC patients.

overall_design: In order to assess the patient's mRNA expression level, we performed the RNA-seq of the FFPE tumor tissue.

first: Di

last: Wu


title: The PD-1 pathway regulates development and function of memory CD8+ T cells following respiratory viral infection

city: Boston

first: Kristen

middle: Elaine

last: Pauken

state: MA

address: 77 Avenue Louis Pasteur NRB rm 837

institute: Harvard Medical School

summary:
The PD-1:PD-L co-inhibitory pathway regulates dysfunctional T cells in chronic viral infection and cancer, but the role of this pathway in effector and memory responses following acute infection or vaccination remains less clear. Here we demonstrated that in the absence of signals from the PD-1 pathway, cell intrinsic alterations during initial CD8+ T cell priming resulted in excessive early CD8+ T cell expansion, but increased CD8+ T cell contraction and aberrant effector to memory CD8+ T cell transition. Overall, our studies revealed a critical and previously unappreciated role for PD-1 as an integrator of early CD8+ T cell activation signals that promoted optimal CD8+ T cell memory formation and durability. This novel PD-1 function has therapeutic implications for the generation of T cell memory during PD-1 cancer immunotherapy and modulation of the PD-1 pathway to enhance immune memory following acute infection or prophylactic vaccination.
Microarrays were performed on influenza-specific CD8+ T cells isolated from the lungs of wildtype and PD-L1/L2 double knockout mice that were infected intranasally with X31-GP33.  Influenza-specific CD8+ T cells were harvested at day 8 in order to determine the global program of gene expression at during the effector phase of the response to influenza to begin to dissect the role of PD-1 signaling early during effector and memory differentiation.

overall_design: Wildtype and PD-L1/L2 double knockout mice were infected with X31-GP33 influenza intranasally. 8 days later, influenza-specific (immunodominant NP366-374 tetramer positive and subdominant GP33-41 or PA224-233 tetramer positive) CD8+ T cells were sorted from the lungs of infected animals.

first: Arlene

middle: H

last: Sharpe


title: Expression data from poplar stems

city: Houghton

first: Victor

last: Busov

state: MI

address: 1400 Townsend Drive

department: Forest Resources and Environmental Science

institute: Michigan Technological University

summary:
We study differences in gene expression between Populus P35S::PLGT , RNAi_PLGT and control/WT plants, affecting tropic responses and stem development. We used microarrays to detail the global program of gene expression underlying morphological and developmental changes droved by th etransgenic manipulations of PLGT gene.
By screening activation tagging population we identified the PLAGIOTROPICA (PLGT) mutant. The mutant is severely impaired in gravitropic responses and displays consistent south orientation of the shoot tip in the overexpressing plants. We positioned the tag, localized a putative candidate gene and verified transcription activation. The activated gene encodes an ortholog of the OCTOPUS gene from Arabidopsis.  We fully recapitulated the phenotype by overexpression of the gene into the same genotype under strong constitutive promoter (P35S::PLGT). We further downregulated the gene using RNAi approach.   The gene was most highly expressed in young stems and phloem tissues.

overall_design: Poplar stem segments derived from the 5th internodes were selected  for RNA extraction and hybridization on Affymetrix microarrays. We sought to uncover the affected geens associated with the native function of this gene by looking at the genome-wide expression changes in the overexpression, downregulated and wild type plants. To do that,  we selected stem segments derived from the 5th internodes tissue, becuase the native expression of PLGT was found to be highest in this tissue.


title: Signal-Specific Genomic Regulation of NF-κB by EGFR and IL-1R

city: Cleveland

first: George

last: Stark

state: OH

address: 9500 Euclid Avenue

department: Cancer Biology

institute: Cleveland Clinic Lerner Research Institute

summary: To study how NF-κB-dependent transcription is regulated differentially by EGFR, compared to the Interleukin-1 receptor (IL-1R), a classical NF-κB activator, we integrated transcriptional analyses by profiling by RNA-seq) and DNA binding by ChIP-seq analyses in human mammary epithelial cells (HMECs).

overall_design: Signal-specific NF-κB-dependent genes activated or repressed by EGF or IL-1 were identified by differential gene expression analysis of RNA-seq data of treated versus untreated samples with or without knockdown of the RELA (p65) subunit of NF-κB. Signal-specific NF-κB binding sites were identified by ChIP-seq using an antibody against p65 followed by differential binding analysis of p65 peaks in treated versus untreated samples.

first: Josephine

last: Dermawan


title: Early MicroRNA Indicators of PPARα Pathway Activation in the Liver

city: Research Triangle Park

first: Brian

last: Chorley

state: NC

address: US EPA, 109 TW Alexander Dr

institute: US EPA

summary: In this study, we measured liver miRNA in male B6C3F1 mice exposed to a known chemical activator of the peroxisome proliferator-activated receptor alpha (PPARα) pathway, di(2-ethylhexyl) phthalate (DEHP), for 7 and 28 days at concentrations of 0, 750, 1500, 3000, or 6000 ppm in feed. At the highest dose tested, DEHP altered 61 miRNAs after 7 days and 171 miRNAs after 28 days of exposure, with 48 overlapping miRNAs between timepoints. Analysis of these 48 common miRNAs indicated enrichment in PPARα targets and other pathways related to liver injury and cancer. Four of the 10 miRNAs exhibiting a clear dose trend were linked to PPARα activation: mmu-miRs-125a-5p, -182-5p, -20a-5p, and -378a-3p.

overall_design: We measured liver miRNA in male B6C3F1 mice exposed to a known chemical activator of the peroxisome proliferator-activated receptor alpha (PPARα) pathway, di(2-ethylhexyl) phthalate (DEHP), for 7 and 28 days at concentrations of 0, 750, 1500, 3000, or 6000 ppm in feed.

middle: N

first: Gleta

last: Carswell


title: Processing by RNase 1 forms tRNA halves and distinct Y RNA fragments in the extracellular environment

city: Woodbury

first: Thomas

last: Gingeras

state: NY

address: 500 Sunnyside Blvd

institute: Cold Spring Harbor Laboratory

summary: In this study, we use small RNA sequencing to investigate the effect of a null mutation of RNase 1 on the levels of tRNA halves and Y RNA fragments in the extracellular environment of cultured human cells. Complemented and extended by the use of northern blots, our results demonstrate that tRNAs and Y RNAs in the vesicle-depleted extracellular compartment are released from cells as full-length precursors. Following their release, tRNAs and Y RNAs are processed by RNase 1 into distinct fragments. In addition, our findings show that standard sequencing methods employed to detect tRNA fragments leave many of such fragments undetected and that a combination of end healing, 3’ deacylation and RNA modification removal before library preparation can substantially improve detection of tRNA halves and reduce biases.

overall_design: An RNASE1 null mutation was generated in K562 human chronic myelogenous leukemia cells. RNA from extracellular vesicle-depleted conditioned medium was sequenced using a small RNA sequencing workflow. Four replicates each of RNA from RNASE1 mutant and wild type cells grown in RPMI 1640 were sequenced with pretreatment of the RNA with T4 polynucleotide kinase (PNK). Two replicates of RNA from RNASE1 wild type cells grown in RPMI 1640 were sequenced without RNA pretreatment with PNK. One replicate each of mutant and wild type cells grown in QBSF-55 medium were sequenced with RNA pretreatment with PNK. Two replicates of RNA from RNASE1 wild type cells grown in RPMI 1640 were sequenced with RNA pretreatment including deacylation followed by PNK and demethylation with AlkB.

first: Gal

last: Nechooshtan

first: Dinar

last: Yunusov


title: DNA methylation in the superior temporal gyrus of 44 subjects with schizophrenia and 44 non-psychiatric control subjects

city: Pittsburgh

first: Brandon

middle: Chad

last: McKinney

state: PA

address: 3811 O'Hara Street

department: Psychiatry

institute: University of Pittsburgh

summary: DNA methylation (DNAm) signal from the planum temporale of superior temporal gyrus (STG) of forty-four subjects with schizophrenia (SZ) and forty-four non-psychiatric control (NPC) subjects was measured on the Illumina MethylationEPIC BeadChip Infinium (EPIC) array. Averaged, normalized beta-values at each site were correlated with dendritic spine density (DSD) previously measured per subject to identify site-specific DNAm-DSD correlations that met methylome-wide significance, or a suggestive-level of significance. We tested the hypothesis that DNAm correlates with DSD in human STG at several sites across the methylome and that this relationship is disrupted in SZ. DSD measures were available for 40 of the subjects with SZ and 40 of the NPC subjects. We found DNAm to correlate with DSD at more sites than expected by chance in NPC, but not SZ, subjects. In addition, we show that the slopes of the linear DNAm-DSD correlations differed between SZ and NPC subjects at more sites than expected by chance. Together, these data suggest that alterations in the intercation between DNAm and neurobiology in SZ may be a mechanism for SZ-related DSD reductions.

overall_design:
A total of 96 genomic DNA samples taken from postmortem STG of subjects with SZ (N=44, with 4 subjects replicated) and NPC subjects (N=44, with 4 subjects replicated) were bisulfite converted and hybridized to the probes on the EPIC array.
SZ: Schizophrenia
NPC: Non-psychiatric control
STG: Superior temporal gyrus (of human brain)

first: Christopher

last: Hensler

middle: C