title: The histone demethylase JMJD2B regulates endothelial-to-mesenchymal transition [HTS]

status: Public on Jan 08 2020

city: Frankfurt

last: John

state: Hessen

address: Theodor-Stern-Kai 7

institute: Institute for Cardiovascular Regeneration

summary: Endothelial cells play an important role in maintenance of the vascular system and the repair after injury. Under pro-inflammatory conditions, endothelial cells can acquire a mesenchymal phenotype by a process named endothelial-to-mesenchymal transition (EndMT), which affects the functional properties of endothelial cells. Here, we investigated the epigenetic control of EndMT. We show that the histone demethylase JMJD2B is induced by EndMT promoting pro-inflammatory and hypoxic conditions. Silencing of JMJD2B reduced TGF-β2-induced expression of mesenchymal genes and prevented the alterations in endothelial morphology and impaired endothelial barrier function. Endothelial-specific deletion of JMJD2B in vivo confirmed a reduction of EndMT after myocardial infarction. EndMT did not affect global H3K9me3 levels but induced a site-specific reduction of repressive H3K9me3 marks at promoters of mesenchymal genes, such as Calponin (CNN1), and genes involved in TGF-β signaling, such as AKT Serine/Threonine Kinase 3 (AKT3) and sulfatase 1 (SULF1). Silencing of JMJD2B prevented the EndMT-induced reduction of H3K9me3 marks at these promotors and further repressed these EndMT-related genes. Our study reveals that endothelial identity and function is critically controlled by the histone demethylase JMJD2B, which is induced by EndMT-promoting pro-inflammatory and hypoxic conditions and support the acquirement of a mesenchymal phenotype.

overall_design:
1.) CHIP-SEQ of H3K9me3 in HUVECS in full-(FM) and differentiation medium (DM)
2.) Whole Transcriptome RNA-SEQ of HUVECS in full-(FM) and differentiation medium (DM)
3.) Single Cell RNA-SEQ of whole heart of Jmjd2b flox Cdh5-iCre and wildtype mice 3 days after AMI
4.) Single Cell RNA-SEQ of HUVECS full-(FM) and differentiation medium (DM)
5.) Single Cell RNA-SEQ of HUVECS in full medium (FM) under scrambled knockdown and differentiation medium (DM) under scrambled knockdown and JMJD2B knockdown

first: Simone

last: Glaser

first: Stefanie

last: Dimmeler


title: Gene expression data of Peyer's patch LysoDC differentiation states  and macrophages at steady state and under TLR7 ligand stimulation

city: Marseille

first: Hugues

last: Lelouard

address: parc Scientifique de Luminy case 906

country: France

institute: CIML CNRS UMR 7280

summary: The specialized monocyte-derived phagocytes termed LysoDC are hallmarks of Peyer’s patches where their main function is to sample pathogens. However, their differentiation pathway and migratory properties remain uncharacterized. From single-cell RNA sequencing, we built LysoDC differentiation trajectories and fate diversity in correlation with their location and functions. All LysoDC differentiation states display similar phagocytic activity. One of them is located in follicles in a CXCR5-independent manner whereas the others reside in subepithelial domes (SED) and mature as they get closer to the epithelium. Immature LysoDC proliferate. Mature LysoDC acquire a gene signature shared with SIRP alpha-expressing conventional DC and prime naïve T cells in vitro but, at steady state, do not migrate in naïve T cells-enriched interfollicular regions (IFR). However, upon stimulation, they express the chemokine receptor CCR7 and migrate from the SED to the IFR periphery where they strongly interact with proliferative T cells. Finally, we show that LysoDC populates human Peyer’s patches.

overall_design: Three to five independent replicates of the 4 LysoDC differentiation states (TN, SP, DP, TP) were sorted from Peyer's patch of C57BL/6 mice at steady state. In addition, three to four independant replicates of 2 LysoDC differentiation states (DP and TP) and of TIM-4- and TIM-4+ macrophages (LysoMac) were sorted from Peyer's patch of C57BL/6 mice 9 hours after R848 gavage. Total RNA of Peyer's patch sorted cells was extracted with a Qiagen RNeasy Plus Micro Kit. Quantity, quality and absence of genomic DNA contamination were assessed with a Bioanalyser (Agilent). Microarray experiments were performed by the Plateforme Biopuces of Strasbourg (France) using the Affymetrix GeneChip® Mouse Gene 1.0 ST Array.

first: Johnny

last: Bonnardel

first: Clément

last: Da Silva


title: Astrocyte molecular signatures during Huntington’s disease progression and following huntingtin lowering with zinc finger protein transcriptional repressors

city: Los Angeles

first: Giovanni

last: Coppola

state: CA

address: 1524 Gonda, 695 Charles Young Drive South

department: Psychiatry and Neurology

country: USA

institute: UCLA

type: Expression profiling by high throughput sequencing

summary:
The Khakh laboratory used gfaABC1D-RiboTag AAVs to purify and sequence astrocyte actively translated mRNAs from Huntington's disease mouse models at several stages of the disease.  Two weeks after AAV injection, striata were homogenized and RNA was purified from (i) cleared lysate as the input and control, and (ii) astrocyte-specific ribosome-associated RNA precipitated via a hemagglutinin (HA) tag.

R6/2 mice are a fast developing model of Huntington's disease. Striatal cells show mutant HTT inclusions as early as 4 weeks of age. These mice show the first motor and cognitive symptoms around that age, but they become more evident around 8 week old and progressively impair until the mouse death that occurs around 13 weeks of age. Motor symptoms include increased paw clasping and grooming, impaired grip strength and rotarod performance, gait alterations, involuntary movements, etc. Cognitive impairment includes defects in learning and memory.

NCAR mice are the healthy "non-carrier" controls for R6/2. They don't have unexpected phenotypes.

overall_design: Q175 transcriptomes were compared to WT at 2, 6 and 12 month old. R6/2 transcriptomes were compared to NCAR at 1, 2 and 3 month old.

first: Blanca

last: Diaz-Castro

first: Mohitkumar

last: Gangwani

first: Baljit

middle: S

last: Khakh


title: A Second Heart Field-derived vasculogenic niche contributes to cardiac lymphatics

status: Public on Jan 09 2020

city: Madrid

first: Manuel

middle: J

last: Gomez

state: Madrid

address: Melchor Fernández Almagro, 3

country: Spain

institute: CNIC

summary: The mammalian heart contains multiple cell types that appear progressively during embryonic development. Advances in determining cardiac lineage diversification has often been limited by the unreliability of genetic tracers. Here we combine clonal analysis, genetic lineage tracing, tissue transplantation and mutant characterization to investigate the lineage relationships between epicardium, arterial mesothelial cells (AMCs) and the coronary vasculature. We report a contribution of the second heart field (SHF) to a vasculogenic niche composed of AMCs and sub-mesothelial cells at the base of the pulmonary artery. Sub-mesothelial cells from this niche differentiate into lymphatic endothelial cells and, in close association with AMC-derived cells, contribute to, and are essential for the development of ventral cardiac lymphatics. In addition, regionalized epicardial/mesothelial retinoic acid signaling regulates lymphangiogenesis, contributing to the niche properties. These results uncover a SHF vasculogenic contribution to coronary lymphatic development through a local niche at the base of the great arteries.

overall_design: Transcriptomic analysis comparing mRNA from dissected mesothelial + sub-mesothelial layers of the base of the great arteries of the heart with that of the dissected epicardium + sub-epicardiom of the heart ventricles

first: Lioux

last: Ghislaine

first: Torres

last: Miguel


title: Transcriptome profiling of influenza-specific memory CD8 T cells in the lung airways (BAL), lung interstitium (LR), lung vasculature (LV), and spleen

city: Atlanta

first: Chris

last: Scharer

state: GA

address: 1510 Clifton Rd, Suite 3086A

department: Microbiology and Immunology

institute: Emory University

summary: To examine differences in the transcriptome of resident memory CD8 T cells in the lung, influenza nucleoprotein-specific CD8 T cells from the airways , lung interstitium, and spleen were FACS sorted and RNA isolated for RNA-seq.

overall_design: C57Bl/6 mice were infected with influenza x31 and rested for 35 days to establish T cell memory. Five minutes prior to euthanasia mice were injected with anti-CD45.2-PE to discriminate between tissue resident and circulating cells, and cells were isolated from the airways by bronchoalveolar lavage, from the lung parechyma by enzymatic digestion, and from the spleen by mechanical disruption. Cells were sorted on CD8+ CD44Hi FluNP 366-374 Db tetramer+ populations, and cells from the airways and parenchyma were further sorted on intravital label negative (i.e. tissue resident) cells.  This cohort was used as a replicate cohort.

first: C

middle: D

first: J

middle: E

last: Kohlmeier


title: Single cell mRNA sequencing of Peyer's patch monocyte-derived dendritic cells (LysoDC)

status: Public on Jan 07 2020

summary: Single-cell RNA-sequencing was used to reveal LysoDC transcriptional plasticity and dynamics of differentiation

overall_design: 23,000 LysoDC were isolated from 246 Peyer's patches from 35 C57BL/6Rj mice and processed using the Chromium Single Cell 3’ kit from 10X genomics (California, USA) for the library preparation and the NextSeq 500/550 High Output Kit v2 (75 cycles) cartridges from Illumina for the sequencing. FASTQ raw files were processed using Cell Ranger software (v2.1.1, default parameters), which performs alignment, filtering, barcode counting and unique molecular identifier (UMI) counting. Reads were aligned to the mouse mm10 genome.

first: Camille

last: Wagner

first: Lionel

last: Spinelli


title: Transcriptome profiling of influenza-specific memory CD8 T cells following transfer into the airways or in vitro culture

summary: To examine differences in the transcriptome of resident memory CD8 T cells in the lung, influenza nucleoprotein-specific CD8 T cells from the airways , lung interstitium, lung vasculature, and spleen were FACS sorted and RNA isolated for RNA-seq. In some experiments, FACS sorted cells were transferred into the airways, peritoneum, or in vitro culture before subsequent isolation and RNA isolation.

overall_design: C57Bl/6 mice were infected with influenza x31 and rested for 35 days to establish T cell memory. Five minutes prior to euthanasia mice were injected with anti-CD45.2-PE to discriminate between tissue resident and circulating cells, and cells were isolated from the airways by bronchoalveolar lavage, from the lung parechyma by enzymatic digestion, and from the spleen by mechanical disruption. Cells were sorted on CD8+ CD44Hi FluNP 366-374 Db tetramer+ populations, and cells from the airways and parenchyma were further sorted on intravital label negative (i.e. tissue resident) cells.  Lung vasculature cells were lung cells that stained positive for the intravital label. This cohort was used as a replicate cohort. In some experiments, FACS sorted cells from the airways or spleen were transferred into the airways or peritoneum of congenic (CD45.1+) mice, or placed in an in vitro culture, before subsequent isolation and RNA isolation 48 hours post-transfer.


title: Two separate roles for the transcription coactivator SAGA and a set of genes redundantly regulated by TFIID and SAGA [RNA]

city: Oklahoma City

first: Rafal

last: Donczew

state: OK

address: 825 NE 13th St

department: Cell Cycle and Cancer Biology

institute: Oklahoma Medical Research Foundation

summary: Deletions within genes coding for subunits of the transcription coactivator SAGA caused strong genome-wide defects in transcription and SAGA-mediated chromatin modifications. In contrast, rapid SAGA depletion produced only modest transcription defects at 13% of protein-coding genes – genes that are generally more sensitive to rapid TFIID depletion. However, transcription of these “coactivator-redundant” genes is strongly affected by rapid depletion of both factors, showing the overlapping functions of TFIID and SAGA at this gene set. We suggest that this overlapping function is linked to TBP-DNA recruitment. The remaining 87% of expressed genes that we term “TFIID-dependent” are highly sensitive to rapid TFIID depletion and insensitive to rapid SAGA depletion. Genome-wide mapping of SAGA and TFIID found binding of both factors at many genes independent of gene class. DNA analysis suggests that the distinction between the gene classes is due to multiple components rather than any single regulatory factor or promoter sequence motif.

overall_design: Newly synthesized RNAs were labeled with 4-thioU, purified and analyzed with RNA-seq. Depletion of TFIID and/or SAGA subunits was achieved either thorugh targeted degradation (auxin-degron system) or SAGA subunits deletion. Corresponding control and treatment sample files are labeled DMSO and 3IAA, respectively. For SAGA subunit deletion strain BY4705 was used to construct spt20 and spt7 deletion strains and strain SHY565 was used to construct spt3 deletion strain. All experiments were done in two (A,B) or three replicates (A,B,C).

first: Linda

last: Warfield

first: Derek

last: Pacheco

first: Ariel

last: Erijman

first: Steven

last: Hahn


title: Transcriptome profiling of influenza-specific memory CD8 T cells in the lung airways, lung parenchyma, and spleen

summary: To examine differences in the transcriptome of resident memory CD8 T cells in the lung, influenza nucleoprotein-specific CD8 T cells from the airways , lung parenchyma, and spleen were FACS sorted and RNA isolated for RNA-seq.

overall_design: C57Bl/6 mice were infected with influenza x31 and rested for 35 days to establish T cell memory. Five minutes prior to euthanasia mice were injected with anti-CD45.2-PE to discriminate between tissue resident and circulating cells, and cells were isolated from the airways by bronchoalveolar lavage, from the lung parechyma by enzymatic digestion, and from the spleen by mechanical disruption. Cells were sorted on CD8+ CD44Hi FluNP 366-374 Db tetramer+ populations, and cells from the airways and parenchyma were further sorted on intravital label negative (i.e. tissue resident) cells.


title: Viral Delivery Of GDNF Promotes Functional Integration Of Human Stem Cell Grafts In Parkinson’s Disease

status: Public on Jan 06 2020

city: Melbourne

first: Christopher

last: Bye

state: Victoria

address: University of Melbourne

country: Australia

institute: The Florey Institute of Neuroscience and Mental Health

summary: Dopaminergic neurons (DAn), generated from human pluripotent stem cells (hPSC), are capable of functionally integrating following transplantation and have recently advanced to clinical trials for Parkinson’s disease (PD). However, pre-clinical studies have highlighted the low proportion of DAn within hPSC-derived grafts and their inferior plasticity compared to fetal tissue. Here we examined whether delivery of a developmentally critical protein, glial cell line-derived neurotrophic factor (GDNF), could improve graft outcomes. We tracked the response of DAn implanted into either a GDNF-rich environment, or after a delay in exposure. Early GDNF promoted survival and plasticity of non-DAn, leading to enhanced motor recovery in PD rats. Delayed exposure to GDNF promoted functional recovery through increases in DAn specification, DAn plasticity and dopamine metabolism. Transcriptional profiling revealed a role for MAPK-signalling downstream of GDNF. Collectively these results demonstrate the potential of neurotrophic gene therapy strategies to improve hPSC graft outcomes.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Gene expression profile of human xenografts transplanted into host mouse brains. Mixes species RNAseq with 3 biological replicates of 2 xenograft types (midbrain dopamine neurons differentated from embryonic stem cells transplanted into an Untreated or GDNF treated host).

first: Clare

last: Parish


title: The histone demethylase JMJD2B regulates endothelial-to-mesenchymal transition [microarray]

first: David

last: John

country: Germany

overall_design: Affymetrix Human Exon 1.0 ST Array of HUVEC cells in full- and differentiation medium under control and JMJD2B knockdown.

first: Simone

middle: F

last: Glaser

first: Stefanie

last: Dimmeler


title: Identification of therapeutic targets for treatment of intermittent claudication that mimic the effects of exercise

status: Public on Jan 05 2020

city: Tokyo

first: Momoko

last: Ogitani

address: 1-2-58, Hiromachi, Shinagawa-ku

department: Research and Development Division

country: Japan

institute: Daiichi Sankyo Co., Ltd.

summary: Exercise is considered as the most efficacious intervention for improving walking capacity in patients with intermittent claudication (IC). Despite its benefits, the underlying mechanisms inducing this effect remain unclear. To elucidate the local mechanisms in skeletal muscle, we previously developed an electrical stimulating (ES) model which caused skeletal muscle contraction mimicking exercise. ES showed the improvement in walking ability in rat IC model. In this study, we compared gene expressions among hindlimb ischemia (Isch) group, Isch + exercise group, and Isch + ES group in rat using GeneChip analysis and real-time PCR. By extracting genes which were similarly altered in both exercise and ES groups compared with Isch group, we revealed candidate genes that might be involved in improvement of walking ability.

overall_design: In order to investigate the effects of skeletal muscle contraction against IC symptom, we developed electrical stimulation (ES) system in rat IC model. ES could improve walking performance in the rat claudication model as well as exercise. We postulated that nerve-mediated skeletal muscle contraction, which caused by both ES and exercise would affect ischemic hindlimbs directly and the changes contributed to improve walking distance in rat IC model. In order to obtain a comprehensive understanding of the mechanisms of exercise and ES, changes of expressions by exercise and ES versus ischemic group in rat skeletal muscle were analyzed.


title: Cellular and molecular characterization of multiplex autism in human induced pluripotent stem cell-derived neurons

city: Saint Louis

first: Kristen

middle: L

last: Kroll

state: MO

address: 320 McDonnell Sciences/660 S. Euclid Ave.

department: Developmental Biology

institute: Washington University School of Medicine

summary:
Background: Autism spectrum disorder (ASD) is a neurodevelopmental disorder with pronounced heritability in the general population. This is largely attributable to effects of polygenic susceptibility, with inherited liability exhibiting distinct sex differences in phenotypic expression. Attempts to model ASD in human cellular systems have principally involved rare de novo mutations associated with ASD phenocopies. However, by definition, these models are not representative of polygenic liability, which accounts for the vast share of population-attributable risk.

Methods: Here, we performed what is, to our knowledge, the first attempt to model multiplex autism using patient-derived induced pluripotent stem cells (iPSCs) in a family manifesting incremental degrees of phenotypic expression of inherited liability (absent, intermediate, severe). The family members share an inherited variant of unknown significance in GPD2, a gene that was previously associated with developmental disability but here is insufficient by itself to cause ASD. iPSCs from three first-degree relatives and an unrelated control were differentiated into both cortical excitatory (cExN) and cortical inhibitory (cIN) neurons, and cellular phenotyping and transcriptomic analysis were conducted.

Results: cExN neurospheres from the two affected individuals were reduced in size, compared to those derived from unaffected related and unrelated individuals. This reduction was, at least in part, due to increased apoptosis of cells from affected individuals upon initiation of cExN neural induction. Likewise, cIN neural progenitor cells (NPCs) from affected individuals exhibited increased apoptosis, compared to both unaffected individuals. Transcriptomic analysis of both cExN and cIN neural progenitor cells revealed distinct molecular signatures associated with affectation, including misregulation of suites of genes associated with neural development, neuronal function, and behavior, and altered expression of ASD risk-associated genes.

Conclusions: We have provided evidence of morphological, physiologic, and transcriptomic signatures of polygenic liability to ASD from an analysis of cellular models derived from a multiplex autism family. ASD is commonly inherited on the basis of additive genetic liability. Therefore, identifying convergent cellular and molecular phenotypes resulting from polygenic and monogenic susceptibility may provide a critical bridge for determining which of the disparate effects of rare highly deleterious mutations might also apply to common autistic syndromes.

overall_design: RNA-sequencing of human induced pluripotent stem cell-derived cortical inhibitory and excitatory neural progenitors for four cell lines from four different individuals with varying autism affectation; four biological replicates per cell line.

first: Emily

middle: M

last: Lewis

first: Kesavan

last: Meganathan

first: Bo

last: Zhang

first: Paul

last: Gontarz


title: A pathogen-responsive gene cluster for the production of highly modified fatty acids in tomato

city: Stanford

first: Elizabeth

last: Sattely

state: California

address: 443 Via Ortega

department: Shriram Center for Biological and Chemical Engineering

institute: Stanford University

summary: In response to biotic stress, plants produce suites of highly modified fatty acids that bear unusual chemical functionality. Despite their chemical complexity and proposed roles in pathogen defense, little is known about the biosynthesis of these decorated fatty acids in plants. Falcarindiol is a prototypical acetylenic lipid present in carrot, tomato, and celery that inhibits growth of fungi and human cancer cell lines. Using a combination of untargeted metabolomics and RNA sequencing, we discovered a biosynthetic gene cluster in tomato (Solanum lycopersicum) that is required for the production of falcarindiol in response to an adapted fungal pathogen, Cladosporium fulvum. By reconstituting the initial biosynthetic steps in a heterologous host (Nicotiana benthamiana) and generating transgenic pathway mutants in tomato, we demonstrate a direct role for three genes in the cluster in falcarindiol biosynthesis. This work reveals a mechanism by which plants sculpt their lipid pool in response to pathogens, and provides critical insight into the complex biochemistry of alkynyl lipid production.

overall_design: Tomato (Solanum lycopersicum VF36, 4.5 week-old plants) were treated with a panel of fungal elicitors via leaf infiltration: chitin (Ct, 0.5 mg/ml), Malassezia restricta (Mz, OD600 = 0.5), Cladosporium fulvum (Cf, OD600 = 1.0), and a panel of bacterial elicitors: flg22 (Fg, 1 μM), Staphylococcus epidermidis (Sc, OD600 = 1.0), Propionibacterium acnes (Pb, OD600 = 1.0) and Xanthomonas euvesicatoria (Xe, OD600 = 0.2), and water (mock, control). After pathogen inoculation, leaves (one leaf from three individual plants per treatment, n=3) were collected at 12, 24, and 48 hours post-inoculation using razor, flash frozen by liquid nitrogen, and stored at -80 °C for later use in gene profiling and RNA-Sequencing.

first: Ju

last: Jeon

first: Jung-Gun

last: Kim

first: Mary

middle: B

last: Mudgett


title: Lung airways reprogram circulating memory CD8 T cells

first: Chris

last: Scharer

summary: To examine how the transcriptome of circulating memory CD8 T cells isolated from the spleen are reprogrammed in the lung airways, influenza nucleoprotein-specific CD8 T cells from the spleen were FACS sorted and RNA isolated for RNA-seq or the cells were transferred to the lung airways and allowed to rest for 2 days.  Following 2 days, donor influenza nucleoprotein-specific CD8 T cells were FACS sorted and RNA isolated for RNA-seq..

overall_design: C57Bl/6 mice were infected with influenza x31 and rested for 35 days to establish T cell memory. Effector memory (CD44Hi CD62L-) CD8 T cells were isolated from the spleen by cell sorting and transferred intratracheally into naive congenic CD45.1+ mice. 48 hours later, transferred cells were isloated from the airways by bronchoalveolar lavage and purified by sorting on donor (CD45.2+) CD8+ CD44Hi FluNP 366-374 Db tetramer+ populations, and RNA was isolated for downstream analysis.

first: C

first: J

last: Kohlmeier


title: The lung microenvironment results in unique gene expression and epigenomic programs in airway and parenchyma resident memory CD8 T cells

type: Genome binding/occupancy profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: mSeq of total and polysome bound mRNAs in two yeast strains

status: Public on Jan 04 2020

city: Bern

first: Norbert

last: Polacek

address: Freiestrasse 3

department: Chemistry and Biochemistry

country: Switzerland

institute: University of Bern

summary: We report on how the association of mRNA to translating ribosomes changes due to the absence of the expansion segment 27L in S. cerevisiae. We observe specific enrichment and depletion of mRNA sets from rmutant ribosomes which reshapes the proteome.

overall_design: Examination of total and ribosome associated mRNA in wild-type and Δ27Lb yeast yJD1045 strain depleted of genome rDNA. rRNA is exclusively plasmid borne.

first: Vaishnavi

last: Shankar

first: Robert

last: Rauscher

first: Walid

middle: H

last: Gharib

first: Julia

last: Reuther

first: Miriam

last: Koch


title: RNA-seq profiles of  WT and Qk-KO freshly isolated oligodendrocytes

city: Houston

first: Jian

last: Hu

state: Texas

address: 1881 East Rd

department: Department of Cancer Biology, Division of Basic Sciences

institute: The University of Texas MD Anderson Cancer Center

summary: Lipid-rich myelin forms insulating axon-wrapping multilayers essential for neural function, but how mature myelin maintains its structural lipid components remains obscure. Here we identify Quaking (Qki) as a major regulator of myelin lipid homeostasis. Qki depletion in adult myelinating oligodendrocytes disrupted myelin lipid metabolism and caused demyelination, resulting in progressive neurological deficits that could be partially rescued by high-fat diet. Mechanistically, Qki serves as a coactivator of the PPARb-RXRa complex, which controls transcription of lipid-metabolism genes, particularly those for fatty acid desaturation and elongation. Treatment of Qki-depleted mice with PPARb or RXR agonists alleviated neurological disability and significantly extended mouse survival. Furthermore, a subset of lesions from patients with primary progressive multiple sclerosis were characterized by preferential reduction of myelin lipids and activities of lipid-metabolism pathways. Together,  our findings demonstrate that continuous lipid synthesis is indispensable for mature myelin maintenance and highlight an underappreciated role of lipid metabolism in demyelinating diseases.

overall_design:
This study is to analyze the transcriptomic profiles of freshly isolated oligodendrocytes from Plp-CreERT2;QKLoxP/LoxP (Qk-KO) mice and control mice to determine the differentially expressed genes.
Plp-CreERT2;QKLoxP/LoxP (Qk-KO) mice and control mice were injected with tamoxifen at 8-week, and brains were collected 4 days after tamoxifen injection to isolate oligodendrocyte freshly. The total RNAs were extracted by Rnasey kit and sent for sequencing with llumina HiSeq2000 Sequencer.

first: Xin

last: Zhou

first: Jiangong

last: Ren


title: Transcriptome (RNA-Seq), translatome (Ribo-Seq), and YB-1/YB-3-bound mRNAs (RIP-Seq) of HEK293T, HEK293TdeltaYB-1, and HEK293T+HA-YB-1 cells

city: Pushchino

first: Ivan

middle: V

last: Kulakovskiy

address: Institutskaya 4

country: Russia

institute: Institute of Protein Research

summary: We present results of RNA-Seq, Ribo-Seq, and RIP-Seq (YB-1, YB-3) experiments performed in HEK293T cells, as well as in HEK293T cells with YB-1 knockout and overexpression. The data shows YB-1 function as a global translation inhibitor and YB-3 ability to substitute YB-1 in its function in YB-1 knockout mutant.

overall_design: Cells were cultivated in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% fetal calf serum, 2 mM glutamine, 100 U/ml penicillin, and 100 µg streptomycin. The cells were incubated at 37°C in a humidified atmosphere containing 5% CO2 and passaged by standard methods. YB-1 knockout was performed with CRISPR/Cas9. YB-1 overexpression was achieved by transit transfection of pcDNA3.1 HA-YB1 plasmid. RIP-Seq was performed using (4202, CST; A303-230A, Bethyl) antibodies for YB-1 and (A303-070A, Bethyl) antibodies for YB-3. Libraries for Ribo-Seq were prepared with TruSeq Ribo Profile Kit (Illumina). The same kit was used for size-matched RNA-Seq (used as the control for Ribo-Seq). RIP-Seq and its control RNA-Seq were produced with NEBNext Ultra Directional RNA Library Prep Kit for Illumina (NEB). Ribosomal RNA depletion was performed with Ribo-Zero rRNA Removal Kit (Human/Mouse/Rat)(Illumina) for Ribo-Seq and with NEBNext Poly(A) mRNA Magnetic Isolation Module (NEB) or NEBNext rRNA Depletion Kit (Human/Mouse/Rat) (NEB) for RIP-Seq. Libraries were sequenced on Illumina HiSeq 2000 and Illumina NextSeq 500.

first: Dmitry

middle: N

last: Lyabin

first: Irina

middle: A

last: Eliseeva


title: Prenociceptin TRAP

city: St Louis

first: Joseph

last: Dougherty

state: Missouri

address: 4566 Scott Ave, box 8232

department: Genetics and Psychiatry

institute: Washington University

summary: We aimed to profile ribosome-bound transcripts in pnoc neurons in the IPN.

overall_design: Injection of TRAP (eGFP/Rpl10a) into pnoc CRE animals, perform TRAP in microdissected IPN.

first: Kristina

last: Sakers


title: RNA-Sequencing of Wild Type, Lola-F and Lola-Null mutant embryos

city: Lausanne

first: Jean-Yves

last: Roignant

address: genopode building

institute: University of Lausanne

summary: longitudinals lacking (lola) is among the most complex genes in Drosophila melanogaster, encoding up to 20 different protein isoforms and acting as a key transcription factor in axonal pathfinding and neural reprograming. To better characterize Lola function we have generated specific mutations in each isoform using the CRISPR/Cas9 system. Our targeted screen allows us to revisit the previously demonstrated roles for few isoforms, to assign known functions to specific isoforms and to reveal a critical role for a specific variant in the octopaminergic pathway. Thus, our comprehensive study expands the repertoire of Lola functions, and demonstrates that the CRISPR/Cas9 approach is a valuable tool to systematically address the role of complex loci in vivo.

overall_design: Embryos were collected at 25°C for two hours and subsequently developed for 13 hours. lola-F and lola-null embryos were hand-sorted and collected based on the expression of mCherry. Control embryos were collected in parallel and both mutant and control embryos were subsequently transferred into TRIzol reagent (Thermo Fisher Scientific) and RNA was isolated using the manufacturers protocol. RNA was DNase I (NEB) treated according to the manufacturers protocol and subjected to library preparation using the NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina®. 1 µg of total RNA was used as starting material for library preparation.

first: Nadja

last: Dinges

first: Nastasja

last: Kreim

first: Violeta

last: Morin


title: RNA-seq profiles of WT and Qk-KO corpus callosum and oligodendrocytes

summary: This study is to analyze the transcriptomic profiles of Plp-CreERT2;QKLoxP/LoxP (Qk-KO) and Plp-CreERT2;QKLoxP/+ (WT) mice and WT and Qk-KO oligodendrocytes to determine the differentially expressed genes.

overall_design: Plp-CreERT2;QKLoxP/LoxP (Qk-KO) and Plp-CreERT2;QKLoxP/+ (WT) mice were injected with tamoxifen at 8-week, and corpus callosum were collected 4 days after tamoxifen injection. WT or Qk-KO neural stem cells were differentiated in Neurobasal medium containing B-27, 2 mM GlutaMAX-I, 100 ng/ml insulin-like growth factor 1 (IGF-1), and 50 ng/ml 3,3',5-Triiodo-L-thyronine (T3). The total RNAs were extracted by Rnasey kit and sent for sequencing with llumina HiSeq2000 Sequencer.

first: Hu

last: Jian

first: Shingu

last: Takashi

first: Zhou

last: Xin


title: Genome-wide maps of Qki-5 and PPARb in mouse oligodendrocytes

city: Shanghai

first: Chenxi

last: He

address: 131 Dong'an Rd, Mingdao Bldg, Rm 512

country: China

institute: Fudan University

summary: We found that there is a big overlap between the Qki5- and PPARb-binding events by ChIP-seq. Interestingly, the genomic distribution analyses of Qki-5- and PPARβ-binding events showed significant enrichment at the regions of the promoter/transcription start site (TSS) which were indicated by Pol II-binding events. The strong preference of the promoter/TSS co-occupancy suggested that Qki-5 might play an important role in PPARβ-mediated transcriptional regulation of its downstream genes.

overall_design: Examination of genome-wide maps Qki-5 and PPARb in oligodendrocytes which are differentiated from neural stem cells or oligodendrocytes freshly isolated from mouse brains by ChIP-seq. Neural stem cells were differentiated in Neurobasal medium containing B-27, 2 mM GlutaMAX-I, 100 ng/mL insulin-like growth factor 1 (IGF-1), and 50 ng/mL 3,3',5-Triiodo-L-thyronine (T3). Oligodendrocytes were freshly isolated from mouse brains using O4 magnetic beads.

first: Fei

last: Lan


title: Transcriptome Analyses of Pulmonary Interstitial Macrophages derived from Hypoxic Cx3cr1cre;Hif1α (Δ/Δ) vs. Hif1fα (flox/flox) Mice

city: Durham

first: Yen-Rei

middle: Andrea

last: Yu

state: NC

address: 40 Duke Medicine Circle

department: Medicine

institute: Duke University School of Medicine

summary: To determine roles by which infiltrating pulmonary intersitial macrophages regulate development and progression of pulmonary vascular remodeling and pulmonary hypertension.

overall_design: RNA sequencing (RNAseq) of non-classical monocytes and IMØ purified from the lungs of Cx3cr1cre;Hif1αΔ/Δ and control Hif1αflox/flox mice after 3 weeks of exposure to hypoxia plus SU5416 (Fig 5A).


title: Impact of high fat diet on Mus musculus domesticus secreted milk large RNA transcriptome

city: West Lafayette

first: Theresa

last: Casey

state: Indiana

address: 175 S. University Street

department: Animal Sciences

institute: Purdue University

summary: The impact of high fat diet on secreted milk small RNA transcriptome was studied by isolating total RNA from milk fat fraction collected on lactation day 10 from control diet fed (C; n=5; 10% fat; 7% sucrose; Research Diets #D12450J, Brunswick, NJ) and high fat diet fed (HF; n=4; Research Diets #D12492, 60% of total kcal energy is fat and match 7% of total kcal is sucrose; Brunswick, NJ) mice.

overall_design: Milk RNA-large transcriptome profiles from fat fraction collected on lactation day 10 from control diet fed (n=5) and high fat diet fed (n=4) mice.

first: Junhui

last: Wang


title: A CRISPR-Cas9 screen identifies essential CTCF anchor sites for Estrogen Receptor-driven breast cancer cell proliferation

city: Amsterdam

first: Yongsoo

last: Kim

address: De Boelelaan 1118

country: Netherlands

institute: University Medical Center Amsterdam, Location VUmc

summary: Estrogen receptor α (ERα) is an enhancer activating transcription factor, a key driver of breast cancer, and a main target for cancer therapy. ER-mediated gene regulation requires proper chromatin-conformation to facilitate interactions between ER-bound enhancers and their target promoters. A major determinant of chromatin structure is the CCCTC-binding factor (CTCF), that dimerizes and together with cohesin stabilizes chromatin loops and forms the boundaries of topologically associated domains. However, whether CTCF-binding elements (CBEs) are essential for ER-driven cell proliferation is unknown. To address this question in a global manner, we implemented a CRISPR-based functional genetic screening approach targeting CBEs located in the vicinity of ERα-bound enhancers. We identified four functional CBEs and demonstrated the role of one in inducing chromatin conformation changes in favor of activation of PREX1, a key ERα target gene in breast cancer. Indeed, high PREX1 expression is a bona-fide marker of ER-dependency in cell lines, and associated with good outcome after anti-hormonal treatment. Altogether, our data show that distinct CTCF-mediated chromatin structures are required for ER- driven breast cancer cell proliferation.

overall_design: The analysis of CTCF, ERalpha, and FOXA1 in MCF-7 cells was done by ChIP-seq data infected either with indicated sgRNA or non-targeting as a control.

first: Reuven

last: Agami

first: Gozde

last: Korkmaz

first: Wilbert

last: Zwart


title: Maternal weight affects placental DNA methylation of genes invovled in metabolic pathways in the common marmoset monkey (Callithrix jacchus)

city: Tampa

first: Derek

last: Wildman

state: FL

address: 3720 Spectrum Blvd

institute: University of South Florida

type: Methylation profiling by high throughput sequencing

summary: Accumulating evidence suggests that dysregulation of placental DNA methylation (DNAm) is a mechanism linking maternal weight during pregnancy to metabolic programming outcomes. The common marmoset, Callithrix jaccus, is a platyrrhine primate species that has provided much insight into studies of the primate placenta, maternal condition, and metabolic programming, yet the relationships between maternal weight and placental DNAm are unknown. Here, we report genome-wide DNAm from term marmoset placentas using reduced representation bisulfite sequencing. We identified 74 genes whose DNAm pattern is associated with maternal weight during gestation. These genes are predominantly involved in energy metabolism and homeostasis, including regulation of glycolytic and lipid metabolic processes pathways.

overall_design: 17 samples ran in duplicate. The sample description indicates the location and the placental disc that each sample was obtained from

first: Laren

last: Narapareddy

first: Don

last: Armstrong

first: Amy

last: Weckle

first: Aleeca

last: Bell

first: Crystal

last: Patil

first: Suzette

last: Tardif

first: Corinna

last: Ross

first: Julienne

last: Rutherford


title: RNAseq of parental and neratinib resistant 5637 and OVCAR8 cells with or withour neratinib treatment

city: Nashville

first: Dhivya

last: Sudhan

state: TN

address: 777 Preston Research Building

department: Hematology/Oncology Division

institute: Vanderbilt University

summary: Pan-HER TKI neratinib has demonstrated clinical activity in patients with HER2-mutant cancers. However responses are heterogenoeus and not generally prolonged, suggesting de-novo and acquired resistance to neratinib. To study mechanisms of resistance to neratinib we generated neratinib resistant cells by gradual dose escalation until resistance was achieved and performed various analyses including RNAseq.

overall_design: 5637NR and OVCAR8NR cells were maintained under drug-free conditions for 1 week prior to seeding. Parental and neratinib-resistant cells were seeded in triplicate in 10-cm dishes and then treated with or without neratinib (5637 - 600 nM; OVCAR8 - 1 μM); 24h later, cells were harvested and RNA was purified using ReliaPrep RNA Cell Miniprep system (Promega).

first: Carlos

last: Arteaga


title: ChIP-seq of phytophthora sojae P6497

city: Nanjing

first: Haidong

last: Shu

state: Jiangsu

address: Weigang NO.1

department: Department of Plant Pathology

institute: Nanjing Agricultural University

summary: Centromeres are chromosomal regions that serve as platforms for kinetochore assembly and spindle attachments, ensuring accurate chromosome segregation during cell division. Despite functional conservation, centromeric sequences are diverse and usually repetitive across species, making them challenging to assemble and identify. Here, we describe centromeres in the model oomycete Phytophthora sojae by combining long-read sequencing-based genome assembly and chromatin immunoprecipitation for the centromeric histone CENP-A followed by high-throughput sequencing (ChIP-seq). P. sojae centromeres cluster at a single focus in the nucleus at different life stages and during nuclear division. We report a highly contiguous genome assembly of the P. sojae reference strain, which enabled identification of 15 highly enriched CENP-A binding regions as putative centromeres. By focusing on 10 intact regions, we demonstrate that centromeres in P. sojae are regional, spanning 211 to 356 kb. Most of these regions are transposon-rich, poorly transcribed, and lack the euchromatin mark H3K4me2 but are embedded within regions with the heterochromatin marks H3K9me3 and H3K27me3.

overall_design: Examination of H3K9me3 and H3K4me2 on centromeres locus in phytophthora sojae

first: Han

last: Chen

first: Suomeng

last: Dong

first: Yufeng

last: Fang


title: Neonatal Skeletal Progenitors (Affymetrix)

status: Public on Jan 03 2020

first: Charles

middle: Chan

last: KF

address: 265 Campus Drive

institute: Stanford

summary: Prospectively isolated and characerized skeletal progenitor lineages

overall_design: 3 repeats per FACS sorted population

middle: K

last: Chan

first: Michael

middle: T

last: Longaker

first: Irving

last: Weissman


title: Identification and specification of the mouse skeletal stem cell.

summary: How are skeletal tissues derived from skeletal stem cells? Here, we map bone, cartilage, and stromal development from a population of highly pure, postnatal skeletal stem cells (mouse skeletal stem cells, mSSCs) to their downstream progenitors of bone, cartilage, and stromal tissue. We then investigated the transcriptome of the stem/progenitor cells for unique gene-expression patterns that would indicate potential regulators of mSSC lineage commitment. We demonstrate that mSSC niche factors can be potent inducers of osteogenesis, and several specific combinations of recombinant mSSC niche factors can activate mSSC genetic programs in situ, even in nonskeletal tissues, resulting in de novo formation of cartilage or bone and bone marrow stroma. Inducing mSSC formation with soluble factors and subsequently regulating the mSSC niche to specify its differentiation toward bone, cartilage, or stromal cells could represent a paradigm shift in the therapeutic regeneration of skeletal tissues.

overall_design: Single-cell RNA-sequencing (C1 platform) of FACS-sorted mouse skeletal stem, progenitor, and stromal populations  (mSSC, CSP, Thy+, and 6C3+).

first: Eun

middle: Y

last: Seo

first: Rahul

last: Sinha


title: microRNA expression analysis in fly ovaries from wuho and mei-p26 mutants

city: Taipei

first: Hweijan

last: Hsu

address: no 128, Sec 2,  Academia Rd. Nankang

department: Institute of Cellular Biology

country: Taiwan

institute: Academia Research

type: Non-coding RNA profiling by high throughput sequencing

summary:
Purpose: The goal of this study is to analyze microRNA affected by wuho and mei-p26 mutations in Drosophila ovary
Ovarian microRNA prolife of 7 day old control flies and flies bearing wuho and mei-p26 mutations, in duplicate.Sample libraries were prepared using the QIAseq miRNA Library Kit (QIAGEN) according to the manufacturer's protocols. Adaptors are ligated sequentially to the 3' and 5' ends of miRNAs. Subsequently, universal cDNA synthesis with UMI assignment, cDNA cleanup, library amplification and library cleanup are performed. Libraries were sequenced on an Illumina instrument (75-cycle single-end read, 75SE). Sequencing data was processed using the Illumina software BCL2FASTQ v2.20.

overall_design: Ovarian microRNA prolife of 7 day-old adult control flies and flies bearing wuho and mei-p26 mutanst in duplicate using Illumina instrument (75-cycle single-end read, 75SE).

first: HweiJan

first: Elham

last: Rastegari


title: Gene expression analysis in fly ovaries from wuho mutant

summary:
Purpose: The goal of this study is to analyze genes affected by wuho mutation in Drosophila ovary
Ovarian mRNA prolife of 7 day old control flies and flies bearing wuho mutation, in duplicate, using Illumina NextSeq 500. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: Burrows–Wheeler Aligner (BWA) followed by ANOVA (ANOVA) and TopHat followed by Cufflinks.

overall_design: Ovarian mRNA prolife of  7 day-old adult control w1118 flies and flies bearing wuho mutant in duplicate, using Illumina NextSeq 500 system.


title: Learning causal networks using inducible transcription factors and transcriptome-wide time series

city: South San Francisco

first: Scott

last: McIsaac

address: 1170 Veterans Blvd.

institute: Calico Life Sciences LLC

summary: A small molecule transcriptional induction system is used to induce transcription factors (TFs) in Saccharomyces cerevisiae. Subsequent gene expression changes are used to train parametric models and make predictions of regulator-gene connections.

overall_design: The majority of experiments were sampled at t = 0, 5, 10, 15, 20, 30, 45, and 90 minutes following beta-estradiol addition under phosphate limitation. Induction of a target TF is detectable in <5 minutes and reaches saturation within ~10 minutes following β-estradiol addition at a median level 53-fold higher than at t = 0 min. We estimate that at 90 min. following beta-estradiol addition, the median TF is 28.4-fold more highly expressed than the native-promoter driven TF allele. Gene expression patterns are captured over time using Agilent 2-color microarrays.

first: Sean

middle: R

last: Hackett

first: Edward

last: Baltz

first: Marc

last: Coram


title: miRNA-based liquid biopsy provides early prediction of lethal acetaminophene poisoning: a case study

city: Maastricht

first: Julian

last: Krauskopf

address: Universiteitssingel 50

institute: Maastricht University

summary: The study comprises of 24 samples that have been taken over a time course during a lethal APAP overdose

overall_design: Samples have been taken after an accidental acetaminophen overdose from a single case. Several blood samples were taken per day during the treatment until the subject deceased. Samples were recovered from whole blood following a centrifugation of whole blood at 3000g for 10 minutes at room temperature and subsequently stored at 80°C. Circulating miRNAs were isolated using the miRNeasy Serum/Plasma kit (Qiagen) from 50ul serum; the quality of the RNA was evaluated using a Bioanalyzer 2100 using the small RNA Kit (Agilent). Sequencing libraries were prepared using the TruSeq Small RNA-Seq Preparation Kit (Illumina), and sequenced on a HiSeq 2000 (Illumina) according to the manufacture's protocol.

first: Jos

last: Kleinjans

first: Jiri

last: Aubrecht


title: The mTOR pathway is necessary for survival of mice with short telomeres.

status: Public on Jan 02 2020

city: Madrid

first: Osvaldo

last: Graña

state: Comunidad de Madrid

address: C/ Melchor Fernández Almagro, 3

department: Structural Biology

institute: CNIO

summary: Gene expression profiles of Terc+/+ and generation 2 (G2) Terc-/- male mice fed rapamycin or control diet during 2 months. Rapamycin diet contains encapsulated rapamycin at 42 ppm (mg of drug per kg of food); control diet contains coating material (Eudragit S100). Rapamycin was microencapsulated by Rapamycin Holdings Inc. (San Antonio, Texas) and was then incorporated into 5LG6 mouse chow (TestDiet, London, UK).

overall_design: 2-3 months old Terc+/+ and G2 Terc-/- male mice were fed rapamycin or control diet during 2 months. Before sacrifice they were fasted overnight and refed during 3 hours.

first: Iole

last: Ferrara

last: Graña-Castro

first: Paula

last: Martínez

first: María

last: Blasco


title: Reprogramming Hutchinson-Gilford Progeria Syndrome fibroblasts resets epigenomic landscape in patient-derived induced pluripotent stem cells [ChIP-Seq]

city: Ottawa

first: Theodore

last: Perkins

state: ON

address: 501 Smyth Rd.

department: Cellular and Molecular Medicine

country: Canada

institute: Ottawa Hospital Research Institute

summary: Hutchinson-Gilford Progeria Syndrome (HGPS) is a segmental premature aging disorder caused by the accumulation of the truncated form of Lamin A known as Progerin within the nuclear lamina. Cellular hallmarks of HGPS include nuclear blebbing, loss of peripheral heterochromatin, defective epigenetic inheritance, altered gene expression, and senescence. To model HGPS using iPSCs, detailed genome-wide and structural analysis of the epigenetic landscape is required to assess the initiation and progression of the disease.

overall_design: Before addition of antibodies, an aliquot of each sample was reserved for input control.  After reverse cross-linking and cleanup, approximately 20ng of each sample was prepared for sequencing using the ChIP-seq sample rep kit (Illumina) according to the manufacturer’s instructions.

first: William

last: Stanford

first: Zhaoyi

last: Chen


title: Discovery and characterization of the antimetabolite action of thioacetamide-linked 1,2,3-triazoles as disruptors of cysteine biosynthesis in Gram-negative bacteria

status: Public on Jan 01 2020

city: Memphis

first: Richard

last: Lee

state: Tennessee

address: 262 Danny Thomas Place

department: Chemical Biology and Therapeutics

institute: St. Jude Children's Research Hospital

summary: Increasing rates of drug-resistant Gram-negative (GN) infections combined with a lack of new GN-effective antibiotic classes is driving the need for the discovery of new agents. As bacterial metabolism represents an underutilized mechanism targeted in current antimicrobial therapies, we sought to identify novel antimetabolites and explore the specific impacts of these inhibitors on  key pathways in bacterial metabolism. This study describes the successful application of this approach to discover N-(phenyl) thioacetamide-linked 1,2,3-triazoles (TAT) that target cysteine synthase A (CysK), an enzyme unique to bacteria that is positioned at a key juncture between several fundamental pathways. The TAT class was identified using a high-throughput screen against Escherichia coli designed to identify modulators of pathways related to folate biosynthesis. TAT analog synthesis revealed a clear structure-activity relationship, and activity was confirmed against GN antifolate-resistant clinical isolates. Spontaneous TAT resistance mutations were tracked to CysK, and mode of action studies led to the identification of a false product formation mechanism between the CysK substrate O-acetyl-L-serine and the TATs. Global transcriptional responses to TAT treatment revealed that these antimetabolites impose substantial disruption of key metabolic networks beyond cysteine biosynthesis. This study highlights the potential of antimetabolite drug discovery as a promising approach to the discovery of novel GN antibiotics.

overall_design: Mid-log preparations of E. coli K12 MG1655 were treated with an antimicrobial thioacetamide-linked 1,2,3-triazole inhibitor to investigate the associated transcriptional response.  The testing was performed in two M9-based medias with either magnesium sulfate or taurine representing the sulfur source.  The experiment was performed in triplicate.

first: Miranda

middle: J

last: Wallace


title: Transcriptome study of MYBPC3 loss of function mutations

city: Ann Arbor

first: Adam

last: Helms

state: MI

address: 7220 MSRB3, 1150 W. Medical Center Dr., SPC 5644

department: Internal Medicine

institute: University of Michigan

summary: Induced pluripotent stem cells were gene edited to introduce heterozygous (promoter deletion, start site deletion, and exon 27 frameshift) and homozygous (exon 27 frameshift) mutations.  iPSCs were differentiated into cardiomyocytes for transcriptome analysis.

overall_design: iPSCs from each genotype were differentiated into cardiomyocytes, purified by SIRPA magnetic bead sorting, and samples collected at day ~30. 5 samples (including at least 3 differentiation batches) were collected for each genotype.


title: Comparative analysis of CreER transgenic mice for the study of brain macrophages – a case study

city: Rehovot

first: Louise

last: Chappell-Maor

address: 234 Hertzl Street

department: Immunology Department

country: Israel

institute: Weizmann Institute of Science

summary: Conditional mutagenesis and fate mapping have contributed considerably to our understanding of physiology and pathology. Specifically, Cre recombinase-based approaches allow the definition of cell type-specific contributions to disease development and of inter-cellular communication circuits in respective animal models. Here we compared Cx3cr1CreER and Sall1CreER transgenic mice and their use to decipher the brain macrophage compartment as a showcase to discuss recent technological advances. Specifically, we highlight the need to define the accuracy of Cre recombinase expression, as well as strengths and pitfalls of these particular systems that should be taken into consideration when applying these models.

overall_design: We examined 4 individual mice from 2 different genotypes, the whole brain from each mouse was further divided into 4 samples: Sort, Sort+anti-HA IP, direct anti-HA IP and direct IgG IP.

first: Masha

last: Kolesnikov

first: Jung-Seok

last: Kim

first: Anat

last: Shemer

first: Zhana

last: Haimon

first: Jonathan

last: Grozovski

first: Sigalit

last: Boura-Halfon

first: Takahiro

last: Masuda

first: Marco

last: Prinz

first: Steffen

last: Jung


title: Identification of ALDH1A3 as a viable therapeutic target in breast cancer metastasis-initiating cells

city: Toon

first: Daisuke

last: Yamashita

address: Shitsukawa

department: Neurosurgery

institute: Ehime University Graduate School of Medicine

summary: The development of efficacious therapies targeting metastatic spread of breast cancer to the brain represents an unmet clinical need. Accordingly, an improved understanding of the molecular underpinnings of central nervous system spread and progression of breast cancer brain metastases (BCBM) is required. In this study, the clinical burden of disease in BCBM was investigated, as well as the role of aldehyde dehydrogenase 1A3 (ALDH1A3) in the metastatic cascade leading to BCBM development. Initial analysis of clinical survival trends for breast cancer and BCBM determined improvement of breast cancer survival rates, however this has failed to positively impact the prognostic milestones of triple-negative breast cancer (TNBC) brain metastases (BM). ALDH1A3 and a representative epithelial-mesenchymal transition (EMT) gene signature (mesenchymal markers CD44, Vimentin) were compared in tumors derived from BM, lung metastases (LM) or bone metastases (BoM) of patients as well as mice post-injection of TNBC cells. Selective elevation of the EMT signature and ALDH1A3 were observed in BM, unlike LM and BoM, especially in the tumor edge. Furthermore, ALDH1A3 was determined to play a role in BCBM establishment via regulation of circulating tumor cell (CTC) adhesion and migration phases in the BCBM cascade. Validation through genetic and pharmacologic inhibition of ALDH1A3 via lentiviral shRNA knockdown and a novel small molecule inhibitor demonstrated selective inhibition of BCBM formation with prolonged survival of tumor-bearing mice. Given the survival benefits via targeting ALDH1A3, it may prove an effective therapeutic strategy for BCBM prevention and/or treatment.

overall_design: We established a metastatic murine model via the cardiac injection of MDA-MB-231 using immunocompromised mice (SCID Beige) and then produced a series of derivative cell lines from the resultant tumors within the brain (231-BM), lung (231-LM) or bone (231-BoM)

first: Mutsuko

last: Minata

first: Ichiro

last: Nakano


title: Genome wide gene expression profiling and methylation profiling of directly reprogrammed osteoblasts from mouse fibroblasts

city: stanford

first: Hui

last: Zhu

address: 1651 Page Mill Road

institute: Stanford Universtiy

summary: We identified four transcriptional factors, Runx2, Osx, Dlx5, and ATF4, that rapidly and efficiently reprogram mouse fibroblasts derived from 2.3 kb type I collagen promoter-driven green fluorescent protein  (Col2.3GFP) transgenic mice into induced osteoblast cells (iOBs). The global transcriptome profiling validated that iOBs resemble primary osteoblasts. Genome-wide DNA methylation analysis demonstrates that within differentially methylated loci, the methylation status of iOBs more closely resembles primary osteoblasts than mouse fibroblasts. We further demonstrate that Col2.3GFP+ iOBs have transcriptome profiles similar to GFP+ cells derived from Col2.3GFP transgenic mouse bone chips.

overall_design: Examine gene expression profiling of Induced osteoblasts (iOB), fibroblast, calvarial derived osteoblast (calvarial-OB), Col2.3GFP+ sorted from iOB (iOB GFP+), Col2.3GFP+ sorted from bone chip (Bone chip GFP+); Examine mathylation profiling of iOB, fibroblast, calvaril-OB

first: Joy

last: Wu


title: Integrative QTL analysis of gene expression and chromatin accessibility identifies multi-tissue patterns of genetic regulation (ATAC-seq)

city: Chapel Hill

first: Terry

last: Furey

address: 120 Mason Farm Road

department: Genetics

institute: University of North Carolina, Chapel Hill

summary: Gene transcription profiles across tissues are largely defined by the activity of regulatory elements, most of which correspond to regions of accessible chromatin. Regulatory element activity is in turn modulated by genetic variation, resulting in variable transcription rates across individuals. The interplay of these factors, however, is poorly understood. We characterize expression and chromatin state dynamics across three tissues—liver, lung, and kidney—in 47 strains of the Collaborative Cross (CC) mouse population, examining the regulation of these dynamics by expression quantitative trait loci (eQTL) and chromatin QTL (cQTL). QTL whose allelic effects were consistent across tissues were detected for 1,101 genes and 133 chromatin regions. Also detected were eQTL and cQTL whose allelic effects differed across tissues. Leveraging overlapping measurements of gene expression and chromatin accessibility on the same mice from multiple tissues, we used mediation analysis to identify chromatin and gene expression intermediates of eQTL effects. This analysis demonstrates the complexity of transcriptional and chromatin dynamics and their regulation over multiple tissues, as well as the value of the CC and related genetic resource populations for identifying specific regulatory mechanisms within cells and tissues.

overall_design: Investigation of relationships between genetic variation, chromatin accessibility, and gene expression in lung, liver, and kidney tissue from 47 Collaborative Cross mouse strains by differential analysis between tissues, QTL mapping, and mediation analysis

first: Bryan

middle: C

last: Quach

first: Terrence

middle: S


title: Combined Solution and Crystal Methods Reveal the Electrostatic Tethers That Provide a Flexible Platform for Replication Activities in the Bacteriophage T7 Replisome

city: Cullowhee

first: Jamie

last: Wallen

state: North Carolina

address: 111 Memorial Drive

department: Chemistry & Physics

institute: Western Carolina University

type: Genome variation profiling by high throughput sequencing

summary: We use Illumina sequencing to monitor mutations in the bacteriophage T7 genome in the presence of T7 DNA polymerase that has an altered exonuclease active site. These alterations include mutation of key residues in the exonuclease active site.

overall_design: Bacteriophage T7 that is missing the gene 5 DNA polymerase (T7Δ5) was passaged ten times in the presence of either wild-type or exonuclease-deficient T7 DNA polymerase provided on a plasmid DNA. The genomic DNA from the passaged virus was purified, sequenced, then compared to a T7 genome deposited in GenBank (NC_001604.1-2). Mutations that accumulated during the passages were mapped to specific regions of the bacteriophage genome. We also sequenced our starting passage zero virus as a reference control.


title: Integrative QTL analysis of gene expression and chromatin accessibility identifies multi-tissue patterns of genetic regulation (RNA-seq)


title: Enhanced pre-pubertal nutrition upregulates mitochondrial function in the testes and sperm of post-pubertal Holstein bulls

city: Calgary

first: Chinju Therese

last: Johnson

state: Alberta

department: Production Animal Health

institute: University of Calgary

summary: Recent studies have documented the profound impact of pre-pubertal nutrition on reproductive performance in bulls. Previously, a high plane of nutrition during calfhood (2-30 wk) in beef and dairy bulls hastened puberty explained by an increase in the level of reproductive hormones (LH, testosterone and IGF-I) pre-pubertally, compared to bulls fed less than recommended amounts of energy and protein with no apparent effect on sperm function evaluated by traditional techniques. In addition, upregulated steroid biosynthesis and Sertoli cell maturation was identified in the testicular tissue of high diet bulls at 16 and 24 weeks, respectively. The objective of this study was to evaluate the post- pubertal testes for responses to high (20.0% CP, 67.9% TDN), medium (17.0% CP, 66.0% TDN) and low (12.2% CP, 62.9% TDN) diets fed from 2-30 wk of life. Based on RNA sequencing data, 497 genes were differentially expressed in high vs low diet and 2961 genes in high vs medium diet (P<0.1). According to KEGG analysis, oxidative phosphorylation and ribosome pathways were upregulated in the high diet group (vs medium and low) with majority of the upregulated genes encoding for essential subunits of complex I, III, IV and V of OXYPHOS pathway. In addition, mitochondrial translation, mitotic nuclear division and cell division were enriched in the high vs medium diet group. Supporting the above, the percentage of sperm exhibiting loss of mitochondrial function was lower in the high diet compared to the medium diet (P<0.1). Thus, enhanced early life nutrition upregulated mitochondrial function in both the testes and sperm of post-pubertal Holstein bulls.

overall_design: Holstein bull calves (n=24) maintained on either a high, medium or low diet from 2-32 wk of age, were castrated at 72 wk, testicular mRNA extracted and sequenced.

first: Chinju

first: Jacob

last: Thundathil


title: Exploring the integration of innovation in the dorsal head of Onthophagus beetles using head compartment-specific RNAseq

city: Bloomington

first: David

last: Linz

state: IN

address: Myers Hall 3rd Street

institute: Indiana University

summary: The general morphology of the insect head has remained relatively unchanged through more than 400 million years of evolution. Yet throughout this period this same region has also become a hotspot for evolutionary novelty, yielding structures such as the eyestalks of stalk-eyed flies or the cephalic horns of dung beetles. How novelty can be integrated within ancient complex traits without disrupting the function and formation of that trait is a foundational, yet largely unresolved question in developmental and evolutionary biology. Here, we approached this question by performing unique, head compartment specific RNAseq using the heads of Onthophagus taurus beetles, which bear impressive posterior horns in males. We sequenced the transcriptomes in horned males and hornless females from six distinct head compartments covering two major axes of patterning: anterior to posterior (AP) and medial to lateral (ML). Our results provide evidence of differential compartmentalization of the head along both AP and ML axes, and reveal striking parallels between morphological and transcriptomic complexity – that is, head regions with more complex morphologies, such as the posterior region, are more transcriptionally intricate compared to morphologically homogenous regions, such as the anterior of the head. Our findings support the hypothesis that the integration of novel traits within ancestral trait complexes may require the recruitment of additional genes and pathways into the networks instructing within and among compartment development. However, sexual dimorphism in posterior horn development was not paralleled by a corresponding sexual dimorphism in transcriptional complexity, instead hornless females exhibited approximately the same diversity of differentially expressed genes across posterior head compartments than did horn-bearing males.

overall_design: Dorsal head regions of male and female pre-pupae larvae were dissected into six distinct compartments: left and right posterior, medial posterior, left and right anterior, and medial anterior.

first: Armin

middle: P

last: Moczek


title: U1 snRNP regulates chromatin retention of noncoding RNAs

city: Beijing

first: Yafei

last: Yin

state: Beijing

address: Zhongguancun North Street

department: Medical School

institute: Tsinghua University

summary: Pervasive transcription in the mammalian genome produces thousands of long noncoding RNAs (lncRNAs) and promoter- or enhancer-associated unstable transcripts. They preferentially locate to chromatin, at which some regulate chromatin structure, transcription and RNA processing. While several RNA sequences responsible for nuclear localization have been identified, such as repeats in the lncRNA Xist and Alu-like elements for long RNAs, how lncRNAs as a class are enriched on chromatin remains elusive. To screen for cis-elements that contribute to RNA-chromatin localization, we developed a high-throughput method named RNA elements for subcellular localization by sequencing (REL-seq), and discovered a U1 small nuclear ribonucleoprotein (snRNP)-recognition motif being critical for chromatin localization of reporter RNAs. Across the genome, chromatin-bound lncRNAs, which are enriched with 5’ splice sites and depleted of 3’ splice sites, exhibit high levels of U1 snRNA binding compared to cytoplasm-localized protein-coding mRNAs. Acute depletion of U1 snRNA, or U1 snRNP protein component SNRNP70, drastically reduces the chromatin association of hundreds of lncRNAs and unstable transcripts without altering the overall transcription rate in cells. In addition, rapid degradation of SNRNP70 reduces the localization of both nascent and polyadenylated lncRNA transcripts to chromatin, and disrupts the nuclear-speckles and genome-wide localization of Malat1, a highly conserved and abundant lncRNA. Moreover, chromatin-bound U1 snRNP interacts with transcriptionally engaged RNA polymerase (Pol) II. Together, these results demonstrate that U1 snRNP acts widely to tether and mobilize lncRNAs to chromatin in a Pol II transcription-dependent manner. Our findings uncover a novel role of U1 snRNP beyond pre-mRNA processing and provide molecular insights into how lncRNAs are recruited to Pol II-transcribed genes and have a propensity for chromatin-associated functions.

overall_design: All RNA-seq were used to identify the subcellular localization changes of lncRNAs upon U1 AMO inhibtion or SNRNP70AID degradation. Total RNA-seq identifies both nascent and mature RNAs; polyA RNA-seq mainly identifies mature RNAs; SLAM-seq was used to discriminate newly-synthesized polyA RNA after SNRNP70AID degradation from pre-existed RNAs. All ChIP-seq and TT-seq were used to chracterize the changes of Ser5 or Ser2 phosphorylated Pol II occupancy and transcription rate upon SNRNP70 acute degradation. All ChIRP-seq were used to characterize the changes of Malat1 genome-wide targeting upon SNRNP70 acute depletion or transcription inhibition.

first: Jinlong

last: Lu

first: Xiaohua

last: Shen


title: Robust cullin-RING ligase function is established by a multiplicity of poly-ubiquitylation pathways

city: Toronto

first: Micheal

middle: D

last: Tyers

state: Ontario

address: 555 University Avenue

institute: The Hospital for Sick Children

summary: The large family of SCF ubiquitin ligases catalyze ubiquitylation by bridging protein substrates and ubiquitin-modifying enzymes. S. cerevisiae SCFs employ a sole, essential enzyme, Cdc34, to build poly-ubiquitin chains required for degradation. However, humans have no less than six chain building enzymes associated with SCFs, including the long assumed to be essential Cdc34 orthologs, UBE2R1 and UBE2R2. Furthermore, uncertainty regarding the physiological concentrations of ubiquitin-modifying enzymes has hindered in vitro mechanistic work. Proteomics was used to estimate enzyme levels in cells, and ubiquitylation assays were employed to quantitatively compare enzyme activities. The results show that UBE2R2 alone has negligible ubiquitylation activity at physiological concentrations, and the ablation of UBE2R1/2 had no effect on the stability of SCF substrates in cells. A genome-wide CRISPR screen revealed that UBE2G1 buffers against the deletion of UBE2R1/2. UBE2G1 had robust in vitro activity with SCF, and UBE2G1 knockdown in cells lacking UBE2R1/2 resulted in stabilization of the SCF substrate p27 as well as the Cul3-RING ligase substrate NRF2. The results reveal the human SCF enzyme system is heavily buffered and suggest that SCF specificity is diversified by association with multiple enzyme partners.

overall_design: Nalm-6 cells were sequentially infected with two sgRNAs, targeting either UBE2R2, CDC34 (UBE2R1), the AAVS1 locus or GFP. These were then transduced with a Cas9-expressing lentiviral vector. Six resulting populations, along with a wild-type Nalm-6 clone with a doxycycline-inducible Cas9, were then infected with the EKO sgRNA library (Bertomeu et al. 2017, PMID:29038160), and allowed to grow for an additional 14 days. sgRNA abundance was then assessed by high-throughput sequencing at the beginning and end of the 14 days.

first: Spencer

last: Hill

first: Kurt

last: Reichermeier

first: Daniel

last: Scott

first: Lorena

last: Samentar

first: Luisa

last: Izzi

first: Jasmin

last: Coulombe-Huntington

first: Xiaojing

last: Tang

first: Connor

first: Rebeca

last: Ibarra

first: Thierry

last: Bertomeu

first: Annie

last: Moradian

first: Michael

last: Sweredoski

first: Nora

last: Caberoy

first: Brenda

last: Schulman

first: Frank

last: Sicheri

first: Mike

first: Gary

last: Kleiger


title: Transcriptome analysis of Wild Type and ΔfadR in Shewanella oneidensis MR-1

city: Hangzhou

first: Haichun

last: Gao

state: Zhejiang Province

address: 866 Yuhangtang Rd

department: Institude of microbiology college of life science

institute: Zhejiang University

summary:
Purpose: The goals of this study are to find out the differential expression genes in the fadR mutant strain(ΔfadR) compared with wild-type (WT) and  to further explore the regulation mechanisms of fadR.
Methods: Shewanella oneidensis MR-1 WT and ΔfadR were collected in log phage(OD~0.6). RNA extraction was performed using the RNeasy minikit (Qiagen) and the RNA was quantified by using a NanoVue spectrophotometer (GE Healthcare). RNA seq was performed using Illumina NextSeq 500, 2×150 bp.
Results: Our study represents that the expression of 146 genes were decreased and 94 genes were increased inΔfadR compared with WT. Branched-chain keto acid dehydrogenase (BKD) produces corresponding branched-chain acyl coenzyme A which further participating branchend-chain fatty acids synthesis. The expression of bkdA2 was also promoted in △fadR compared with WT.
Conclusions: Combined with our expression results, it declared that FadR can suppress bkd operon in some degree,which further increase the synthesis of branched-chain fatty acids in ΔfadR.

overall_design: mRNA profiles of Shewanella oneidensis MR-1 and fadR mutant strain were generated by Illumina NextSeq 500

first: Qiu

last: Meng

first: Yongting

last: Zhang


title: Microfluidic platform for next-generation sequencing library preparation with low-input samples

city: Blacksburg

first: Chang

last: Lu

state: VA

address: 235 Goodwin Hall, 635 Prices Fork Road, Virginia Tech

department: Chemical Engineering

institute: Virginia Tech

summary: Advances in Next Generation Sequencing (NGS) have made available a wealth of information that had previously been inaccessible to researchers and clinicians. NGS has been applied to understand genomic, transcriptomic, and epigenomic changes and gained traction as a significant tool capable of accelerating diagnosis, prognosis, and biomarker discovery. However, these NGS assays have yet to be practical methods for patient stratification or diagnosis because of the gap between the tiny quantities of biomaterials provided by a clinical sample and the large DNA input required by most of these assays. Current library preparation methodologies typically require large input amounts of DNA and a long and complicated manual process. Here we present a microfluidic reactor system for NGS library preparation, capable of reducing the number of pipetting steps significantly, automating much of the process, while supporting extremely low DNA input requirement (10 pg per library). This largely automated technology will allow for low-input preparations of 8 libraries simultaneously while reducing batch to batch variation and operator hands-on time.

overall_design: We examined library preparation efficiency using various ChIP DNA amounts looking at genome wide H3K4me3 profiles of GM12878 cell line. The system was further validated using ChIP DNA samples prepared from prefrontal cortex (PFC) of 10-week old male CD-1 mice, exploring different histone modifications: H3K4me3, H3K9me3, H3K36me3, and H3K79me2.

first: Travis

last: Murphy


title: Genome  occupancy of NIPBL in mouse P19 teratocarcinoma cells.

city: Sevilla

first: Jose

middle: C.

last: Reyes

address: Av. Américo Vespucio

department: Genome biology

institute: CABIMER

summary: Mutations in NIPBL are the major cause of Cornelia de Lange Syndrome (CdLS). NIPBL is the cohesin loading factor and has recently been associated with the BET (Bromodomains and Extra Terminal (ET) domain) proteins BRD2 and BRD4. Related to this, a CdLS-like phenotype has been described associated to BRD4 mutations. We have study the genomic  occupancy of NIPBL in mouse P19 teratocarcinoma cells.

overall_design: Chi-seq of Nipbl and imput

first: Noelia

last: Luna-Peláez

last: Guerrero-Martínez

first: Mario

last: García-Domínguez


title: Transcriptome Analysis of ER Stress-Related Genes in Cyclocarya paliurus

city: guiyang

first: zhengting

last: yang

address: huaxi university town

institute: guizhou normal university

summary: Cyclocarya (Juglandaceae) is a genus comprising only one species (C. paliurus (Batal.) Iljin.), which is considered an endangered tree species in China . It belongs to three types of protected plants and is mainly distributed in Hunan, Jiangxi, Anhui, and Guizhou.we investigated the genetic mechanisms of ER stress in C. paliurus by performing transcriptome analysis to compare the profiles of ER stress-related gene expression via different time-course Tunicamycin (TM) treatment（0 hour, 6 hour,14 hour）. A total 196,207 unigenes were detected, with 1,867 down-regulated and 3,040 up-regulated differentially expressed genes (DEGs) found in both treatment (TM_6h and TM_14h )compare to control(TM_0h).

overall_design: The 0.2 gram leaves from three individual trees were treated with tunicamycin (TM, 5 µg/mL) and Triton (0.5%, to improve leaf uptake) for 0 hours, 6 hours, and 14 hours(TM_0h, TM_6h, and TM_14h. Novogene (Beijing, China) in triplicate,performed Illumina Hiseq apparatus sequencing for this study,

first: ZHENGTING

last: YANG


title: Genome-wide promoter DNA Methylation Profiling of Hepatocellular Carcinomas Arising Either Spontaneously or due to Chronic Exposure to Ginkgo biloba Extract (GBE) in B6C3F1/N Mice [Methylation]

city: Research Triangle Park

first: Deepak

last: Mav

address: 2 Davis Drive

institute: Sciome LLC

type: Methylation profiling by genome tiling array

summary: Epigenetic modifications, such as DNA methylation, play an important role in carcinogenesis. In a recent NTP study, chronic exposure of B6C3F1/N mice to Ginkgo biloba extract (GBE) resulted in a high incidence of hepatocellular carcinomas (HCC). Genome-wide promoter methylation profiling on GBE-exposed HCC (2000 mg/kg group), spontaneous HCC (vehicle-control group) and age-matched vehicle control liver was performed to identify differentially methylated genes in GBE-exposed HCC and spontaneous HCC. DNA methylation alterations were correlated to the corresponding global gene expression changes to identify differentially methylated promoter regions of relevant cancer genes altered in GBE-exposed HCC compared to spontaneous HCC.

overall_design: Male and female B6C3F1/N mice were exposed to 0, 200, 600, and 2000 mg/kg GBE by corn-oil gavage, 5 days a week for 104 weeks . Upon examination of the hematoxylin and eosin stained slides made out of FFPE sections representing the frozen counterpart, five CNTL liver (3 males and 2 females), five SPNT-HCC (4 males and 1 female), and five GBE-HCC from high dose 2000 mg/kg group (4 males and 1 female) frozen tissue samples were selected for DNA isolation for methylation array study. Six control liver (4 males and 2 females), six SPNT-HCC (5 males and 1 female) and six GBE-HCC (5 males and 1 female) from 2000 mg/kg are used for RNA isolation for microarray experiments.

first: Ramesh

last: Kovi

first: Sachin

last: Bhusari

first: Ruchir

last: Shah

first: Mark

last: Hoenerhoff

first: Arun

last: Pandiri


title: Transcriptome analysis upon Nipbl, Brd2 and Brd4 knockdown in P19 cells

summary: Mutations in NIPBL are the major cause of Cornelia de Lange Syndrome (CdLS). NIPBL is the cohesin loading factor and has recently been associated with the BET (Bromodomains and Extra Terminal (ET) domain) proteins BRD2 and BRD4. Related to this, a CdLS-like phenotype has been described associated to BRD4 mutations. To understand the relationship between NIPBL and BET proteins, we have performed RNA-Seq expression analysis following depletion of the different proteins in mouse P19 teratocarcinoma cells. Results indicate that genes regulated by NIPBL largely overlap with those regulated by BRD4 but not with those regulated by BRD2.

overall_design: mRNA profiles of P19 mouse teratocarcinoma cells treated with siControl, siNipbl, esiBrd2 or esiBrd4 were generated by deep sequencing, in duplicates, using Illumina Nexseq.


title: Hepatocellular Carcinomas Arising Either Spontaneously or due to Chronic Exposure to Ginkgo biloba Extract (GBE) in B6C3F1/N Mice

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

overall_design: Refer to individual Series


title: DNA sequencing for multiple rounds of the pMHC-yeast display selection for TCR6, TCR7 and FluTCR [other]

city: Stanford

first: Ricardo

last: Fernandes

state: California

address: 279 Campus Drive

department: Molecular and Cellular Physiology

institute: Stanford University

summary: A peptide library was displayed on Db MHC class I allele expressed on yeast. TCRs of interest were used to select and enrich for cognate peptide-MHC.

overall_design: A naive pMHC-yeast display library was selected with biotinylated TCRs of interest present on magnetic beads (Miltenyi) or fluorescently labelled streptavidin. After 4 rounds of TCR selection DNA was extracted using zymolase for all rounds of selected yeast, enriched for cognate pMHC, as well as the naive library. Isolated DNA/plasmid was amplified by PCR with appropriate barcodes and sequencing was performed using a Miseq.

middle: M

last: Davis

first: Naresha

last: Saligrama


title: RNA sequencing of periheral CD4+ T cells and Mucosal associated invariant T cells (MAIT) cells from periphery blood

city: Albany

first: Qi

last: Yang

address: 43 New Scotland Avenue

institute: Albany Medical College

summary: MAIT cells and CD8+ T cells from periphery blood were analysed and revealed altered transcriptome of MAIT cells compared to CD4+ T cells

overall_design: CD4 T cells and MAIT cells from peripheral blood mononuclear cell (PBMC) from healthy donor were sorted.

first: Longyun

last: Ye


title: Expression Profiling of Hepatocellular Carcinomas Arising Either Spontaneously or due to Chronic Exposure to Ginkgo biloba Extract (GBE) in B6C3F1/N Mice [Expression]

state: NC

summary: In a recent NTP study, chronic exposure of B6C3F1/N mice to Ginkgo biloba extract (GBE) resulted in a high incidence of hepatocellular carcinomas (HCC). Genome-wide promoter methylation profiling and Expression profiling on GBE-exposed HCC (2000 mg/kg group), spontaneous HCC (vehicle-control group) and age-matched vehicle control liver was performed to identify differentially methylated genes and differentially expressed genes in GBE-exposed HCC and spontaneous HCC. DNA methylation alterations were correlated to the corresponding global gene expression changes to identify differentially methylated promoter regions of relevant cancer genes altered in GBE-exposed HCC compared to spontaneous HCC.


title: Treg cells promote the SREBP1-dependent metabolic fitness of suppressive intratumoral macrophages via repression of CD8+ T cell-derived interferon-γ

city: Pittsburgh

first: Wei

last: Chen

state: PA

address: 4401 Penn Ave., Rangos Research Building, Room 9125

department: Division of Pulmonary Medicine, Allergy and Immunology, Department of Pediatrics

institute: Children’s Hospital of Pittsburgh of UPMC

summary: Regulatory T (Treg) cells  are crucial for immune homeostasis but they also contribute to tumor immune evasion by promoting a suppressive tumor microenvironment (TME). Mice with Treg cell-restricted Neuropilin 1 deficiency show tumor resistance while maintaining peripheral immune homeostasis, thereby providing a controlled system to interrogate the impact of intratumoral Treg cells on the TME. Using this and other genetic models, we showed that Treg cells shaped the transcriptional landscape across multiple tumor-infiltrating immune cell types. Treg cells suppressed CD8+ T cell secretion of interferon-gamma(IFN[gamma]), which would otherwise block the activation of sterol regulatory element-binding protein 1 (SREBP1)-mediated fatty acid synthesis in immunosuppressive (M2-like) tumor-associated macrophages (TAMs). Thus, Treg cells indirectly but selectively sustained M2-like TAM metabolic fitness, mitochondrial integrity and survival. SREBP1 inhibition augmented the efficacy of immune checkpoint blockade, suggesting that targeting Treg cells or their modulation of lipid metabolism in M2-like TAMs could improve cancer immunotherapy.

overall_design: Freshly sorted CD45+ cells from B16 tumors implanted in Foxp3Cre-YFP or Nrp1L/LFoxp3Cre-YFP mice (female, 5 week of age) on day 14 post inoculation

first: Chang

last: Liu

first: Dario

last: Vignali


title: RNA-Seq in control and TET2 Knockout cells with IFNγ treatment

first: Yue

last: Xiong

address: 450 West Dr

department: Linebeger Cancer Center

institute: UNC-Chapel Hill

summary: Detect overall RNA level change under TET2 knockout in response to IFNγ. Loss of TET2 alters IFNγ transcriptome, TET2 mediates the IFNγ signaling pathway.

overall_design: The experiment is designed to determine the overall RNA level change between TET2 knockout and control group in response to IFNγ. Briefly, we first generated TET2 KO THP1 cell lines by CRISPR technology. Then we treated control and TET2 KO THP1 cells with 100ng/ml IFNγ for 20hrs, total RNAs from each group were extracted and sequenced, 3 replicates in each group and 2 technical repeat for each sample.


title: Transcriptomic profiling of LP lesions, hypertrophic LP (HLP) lesions, and healthy controls

city: Ann Arbor

first: Lam

last: Tsoi

address: Medical Sciences 1 Bldg. 1301 E. Catherine

institute: University of Michigan

summary: The microarray experiment was employed to evaluate the gene expressions in skin lesions of LP, hypertrophic LP (HLP), and healthy controls.

overall_design: To investigate the specific gene regulations in LP, microarray profiling was performed on RNA extracted from paraffin embedded skin biopsy samples obtained from patients with LP, hypertrophic LP (HLP), and healthy controls.

first: Johann

middle: E

last: Gudjonsson


title: SETDB1 safeguards genome stability against necroptosis and bowel inflammation (ChIP-seq)

city: xiamen

first: hongda

last: li

address: School of Life Sciences, Xiang'an Campus, Xiamen University, Xiang'an District, Xiamen, Fujian Province, China

institute: Xiamen University

summary: The aetiology of bowel inflammatory disease (IBD) is a multifactorial interplay between heredity and environment1-3. Here we report SETDB1, a histone methyltransferase (HMTs) for histone H3 lysine 9 trimethylation (H3K9me3) whose deficiency participates in the pathogenesis of IBD. We found that SETDB1 level decreased in IBD patients and that mice with reduced SETDB1 in intestinal stem cells (ISCs) developed spontaneous terminal ileitis and colitis. SETDB1 safeguards genome stability4, of which the loss in ISCs released repression of endogenous retrovirus (ERVs), the retrovirus-like elements with long repeats that comprise ~8% of genome5. Excessive viral mimicry generated by motivated ERVs triggered Z-DNA-binding protein 1 (ZBP1) dependent necroptosis, which irreversibly disrupted the homeostasis of epithelial barrier and promoted bowel inflammation. Of note, genome instability, reactive ERVs, upregulation of ZBP1 and necroptosis were seen in IBD patients. Pharmaceutic inhibition of RIP3 showed curative effect in SETDB1 deficient mice, suggesting therapy targeting ISC necroptosis may represent a new approach for severe IBD treatment.

overall_design: Examination of H3K9me3 in intestinal stem cell.

first: Hongda

last: Li

first: ruicong

last: wang


title: Integrative QTL analysis of gene expression and chromatin accessibility identifies multi-tissue patterns of genetic regulation

first: Terry

last: Furey

institute: University of North Carolina, Chapel Hill


title: Expression data from WT and KO of Myc in innate lymphoid cell 2 (ILC2) in mice

summary:
Group-2 innate lymphoid cells (ILC2) serve crucial function in allergy and asthma. Activated ILC2 rapidly proliferate and secret large amounts of type-2 cytokines, such as IL-5 and IL-13. Mechanisms underlying still remain ambiguous. Here we report that Myc is required for ILC2 proliferation and activation in allergic airway inflammation. Inhibition of Myc impair the ILC2 proliferation in vivo and prevented ILC2-mediated airway hyperresponsiveness in vivo.
We used microarrays to detail the global programme of different gene expression between WT and Myc KO in ILC2.

overall_design: Gene expression comparison between CRISPR-mediated non-target control and Myc knockout ILC2 line

first: Mingwei

last: Liang

first: Xiaofei

last: Shen


title: antigenic KPC model_bulk pancreas RNASeq

city: St Louis

first: Samarth

last: Hegde

state: MO

address: 660 South Euclid Avenue, Campus Box 8069

institute: Washington University in St. Louis

summary: We report the application of bulk RNAseq of live cells from pancreas of KPC or KPC-OG genetic mice at 6 weeks of age. These sponteneous tumors were unperturbed otherwise until timepoint.

overall_design: mRNA sequencing from pancreas of 6 week old KPC or KPC-OG mice, in quadruplets, run on Illumina HiSeq 3000 platform

first: David

middle: G

last: DeNardo


title: Setdb1 safeguards genome stability against stem cell necroptosis and bowel inflammation (RNA-seq)

overall_design: Crypts mRNA profiles of 8-weeks old wild type (setdb1fl/fl) and setdb1-iIEC-KO mice were generated by deep sequencing using BGI-500. ISC total mRNA from Lgr5–EGFP–ires–CreERT2;setdb1fl/+ and Lgr5–EGFP–ires–CreERT2;setdb1fl/fl  were generated for deep sequencing using Hiseq X10.


title: Setdb1 safeguards genome stability against stem cell necroptosis and bowel inflammation


title: AGO-bound virus-derived RNA in Influenza infected  WT and AGO4 knockout bone marrow derived macrophages performed by Immunoprecipitation following by High Throughput Sequencing

city: Boston

first: Kate

middle: L

last: Jeffrey

state: Massachusetts

address: 50 Blossom St, Thier 340

department: Medicine

institute: Massachusetts General Hospital, Harvard Medical School

summary: We identified virus-derived RNA bound to AGO in Influenza A  or Influenza delNS1-infected macrophages taken from wild-type or Ago4 KO littermate control mice

overall_design: Immunoprecipitation of AGO-bound RNA after UV crosslinking using pan AGO antibody in WT or Ago4 KO primary bone marrow-derived macrophages infected with Influenza A (PR/8/1934) or A/PR/8/1934 ΔNS1 (MOI 1 for 24h)

first: Fatemeh

last: Adiliaghdam


title: Systematic genome perturbation reveals coordination of lipid droplets and metabolic gene regulation

first: Niklas

last: Mejhert

address: 655 Huntington Avenue

institute: Harvard TH Chan School of Public Health

summary: Lipid droplets (LDs) store lipids for metabolic energy and are central to cellular lipid homeostasis. The mechanisms coordinating lipid storage in LDs with cellular metabolism are unclear but relevant to obesity-related diseases. Here we utilized genome-wide screening to identify genes that modulate lipid storage in human macrophages, a cell type relevant to metabolic diseases. Among ~550 genes regulating lipid storage, we identify MLX, a basic helix-loop-helix leucine-zipper transcription factor that regulates multiple metabolic processes. We show that MLX and family members MLXIP/MondoA and MLXIPL/ChREBP bind LDs via C-terminal amphipathic helices. When LDs increase in cells, LD binding of MLX, MLXIP/MondoA, and MLXIPL/ChREBP reduces their transcriptional activity, whereas the absence of LDs results in hyperactivation. Our findings uncover an unexpected component to a lipid storage response, in which binding of MLX transcription factors to the LD surface modulates their activity, adjusting the expression of metabolic genes to lipid storage levels.

overall_design: Transcriptional profiles of wild type and MLX depleted THP-1 macrophages and SUM159 cells following incubations with varying concentrations of acetylated LDL or glucose.

first: Leena

last: Kuruvilla

first: Robert

middle: V

last: Farese

first: Tobias

last: Walther


title: The epigenetic landscapes of histone modifications on HSV-1 genome in human THP-1 cells

city: Wuhan

first: gao

last: chuan

state: Hubei province

address: bayi road

department: College of life science

institute: Wuhan university

sample_organism:
Homo sapiens
Human alphaherpesvirus 1

platform_organism:
Homo sapiens
Human alphaherpesvirus 1


title: Gene regulation in wild and cultivated Vitis species during deacclimation and budbreak

city: Geneva

first: Alisson

last: Kovaleski

state: NY

address: 630 W. North Street

institute: Cornell University

summary: Dormant grapevine buds from 4 genotypes were collected over a variable time series after being exposed to growth room temperatures.  Buds were collected from the field during winter and assessed for the loss of cold hardiness (deacclimation) in tandem with analysis of gene expression changes.  DEGs are examined for various enriched pathways.  Wild grapevines deacclimate faster than cultivated grapevines and the entire gene regulation cascade was also faster in wild grapevines.  No keystone genes were identified that could explaint he faster phenotype, suggesting all the regulatory pathways are tuned faster in wild grapes.

sample_organism:
Vitis vinifera
Vitis riparia
Vitis amurensis

overall_design: Examination of dormant buds from 4 genotypes of grapevine; 2 wild and 2 cultivated.

first: Jason

last: Londo


title: Gene expression data of human chord blood hematopoietic progenior cells subjected to myeloid differentiation and transduced with lncRNA HOXA10-AS

city: Frankfurt

first: Jan-Henning

last: Klusmann

address: Theodor-Stern-Kai 7

department: Pediatric Hematology and Oncology

institute: Goehte University Frankfurt

summary: Long non-coding RNAs (lncRNAs) have emerged as crucial regulators of gene expression and cell fate decisions. We predicted HOXA10-AS as a novel non-coding regulator of hematopoiesis and leukemogenesis We overexpressed HOXA10-AS using a bidirectional vector construct, which resulted in impaired monocytic in vitro differentiation and myeloid colony-formation. In addition, shRNA, sgRNA and LNA-GapmeR mediated downregulation of HOXA10-AS resulted in reduced leukemic growth of KMT2A-r cell lines and patient blasts. (p≤0.05). Here we assess early effects of ectopic HOXA10-AS expression in human HSPC subjected to myeloid differentiation conditions.

overall_design: Human CD34+ HSPC from cord blood were transduced with HOXA10-AS or a empty control vector. Transduced cells were subjected to myeloid differentiation conditions. CD11b+CD117-CD14- myeloid precursors were FACS sorted on day 5 of culture. We analyzed early gene expression changes upon ectopic HOXA10-AS expression using the Affymetrix Microarray Clariom™ S Pico Assay

first: Adrian

last: Schwarzer


title: The epigenetic landscapes of histone modifications on HSV-1 genome in human THP-1 cells [ChIP-seq]

summary: To study the dynamic changes of histone modifications across HSV-1 genome during lytic infection in THP-1 cells. Totally 20 maps of HSV-1 epigenome were generated at 5 different time points after infection, together with their corresponding gene expression profiles.We found that histone modifications were detected on HSV-1 genome soon after infection; all four studied modifications, H3K9me3, H3K27me3, H3K4me3 and H3K27ac, change rapidly along with virus life cycle progression.The transcription repression marks, H3K9me3 and H3K27me3, tended to decrease along with infection process, and the transcription activation mark H3K27ac increased on viral genome, which were aligned with increased expression of viral genes. Moreover, treatment with C646, an inhibitor for H3K27ac transferase p300, inhibited expression of viral genes; and EPZ6438, an inhibitor for H3K27 methyltransferase EZH2, enhanced viral gene expression.

overall_design: To study the distribution of histone modifications on HSV-1 genome during lytic stage, as well as their dynamic changes, we used HSV-1 (1×108 / 10 cm dish) to infect THP-1 cells, and collected samples at 1 hr, 2 hr, 4 hr, 8 hr and 24 hr after infection. The cells were then subjected to RNA-Seq and ChIP-Seq with antibodies recognizing H3K27ac, H3K4me3, H3K9me3 and H3K27me3. Two biological replications were performed for each sequencing and data quality control was done before analysis.

first: Chuan

last: Gao

first: Shanbo

last: Tang


title: Expression data from in-vitro cultured mouse stress erythroid progenitors

city: University Pk

last: Paulson

address: 115 Henning Building, Penn State University

department: Veterinary and Biomedical Sciences

institute: Penn State University

summary:
Rapid expansion of stress erythroid progenitors is a key response to acute anemia during stress erythropoiesis. Besides the rapidly amplifying progenitors, a small population of stem-cell like erythroid progenitors undergoes limited number of cell divisions and maintains their stemness. In this study, we addressed the differences in expression profiles of regulatory genes between two stress erythroid progenitor populations that were identified by proliferation capacity and physiological status.
We used microarrays to detail the gene expression profiles of stem-cell like stress erythroid progenitors and rapidly amplifying stress erythroid progenitors during stress erythropoiesis.

overall_design: Stem-cell like stress erythroid progenitors and rapidly amplifying stress erythroid progenitors were obtained from unfractionated mouse bone marrow culture by staining with cell division marker, PKH26, and erythroid progenitor cell surface markers and selecting by cell sorting. Total RNA was extracted from purified cell population and hybridized on Affymetrix microarrays for analysis.

first: Siyang

last: Hao

middle: F


title: Epithelial Vegfa specifies a distinct endothelial population in the mouse lung [bulk RNA-seq]

city: Chicago

first: Lisandra

last: Vila Ellis

state: IL

address: 303 E Superior

department: Cell and Developmental Biology

institute: Northwestern University

summary: The lung microvasculature is essential for gas exchange and commonly considered homogeneous.  We show that Vascular endothelial growth factor A (Vegfa) from the epithelium specifies a distinct endothelial cell (EC) population in the postnatal mouse lung.  Vegfa is predominantly expressed by alveolar type 1 (AT1) cells and locally required to specify a subset of ECs.  Single cell RNA-seq identified 15~20% lung ECs as transcriptionally distinct and marked by Carbonic anhydrase 4 (Car4), which are specifically lost upon epithelial Vegfa deletion.  Car4 ECs, unlike bulk ECs, have extensive cellular projections and are separated from AT1 cells by a limited basement membrane without any intervening pericytes.  Without Car4 ECs, the alveolar space is aberrantly enlarged despite the normal appearance of myofibroblasts.  Lung Car4 ECs and retina tip ECs have common and distinct transcriptional profiles.  These findings support a signaling role of AT1 cells and shed light on alveologenesis.

overall_design: FACS-purified lung endothelial cells were obtained from control and mutant lungs, and analyzed to look for differences in gene expression.

first: Jichao

last: Chen