title: Alteration of miRNome by recombinant ncRNA agents

summary:
Purpose: Identify whether recombinant ncRNAs are recognized by mammalian cells and processed into mature microRNAs, thus leading to the alteration of downstream target genes. In addition, we also explored whether cleavage is Dicer dependent.
Methods: HEK293T (Dicer-WT) and 424 (Dicer-KO) cell lines were treated with 20nM of control tRNA (MSA), nCAR/miR-34a-5p, or nCAR/miR-34a-3p in triplicate.
Results: Read counts of known miRNAs were analyzed using a miRDeep module, and read counts derived from nCAR/miR-34a-5p or nCAR/miR-124a-3p transfection were obtained by mapping sequence reads by an in-house developed PERL script. EdgeR was used to analyze significant (P-value < 0.05 and log2CPM > 6) differentially expressed miRNAs and sRNAs.
Conclusions: Upon transfection of recombinant RNA agents in mammalian cells, nCAR/miR-34a-5p became the most dominant miRNA and was processed in a Dicer dependent manner, while nCAR/miR-124a-3p become the 7th most abundant miRNA and displayed comparable levels with or without Dicer. In both cases, different isoforms of the mature miRNA sequence were generated.

overall_design: A total of 18 samples were analyzed. Three biological replicates in each group were subjected to treatment with control tRNA (MSA), nCAR/miR-34a-5p, or nCAR/miR-124a-3p. Therefore, each cell line had a total of 9 samples and the miRNome was explored in two different cell lines for a total of 18 samples.

middle: Y


title: ATAC-seq (Assay for Transposase-Accessible Chromatin using sequencing) assess genome-wide chromatin accessibility of Runx2fl/fl Mouse Molar Transcriptomes

status: Public on Aug 31 2020

city: Los Angeles

first: Quan

last: Wen

state: Ca

address: 2250 Alcazar St. CSA 123

country: USA

institute: University of Southern California

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Purpose: To study the function of Runx2 on mouse tooth root development. The goals of this study are to assess genome-wide chromatin accessibility of Runx2fl/fl Mouse Molar Transcriptomes to identify the potential downstream target of Runx2. Methods: the apical halves of lower 1st molars of PN7.5 control mice were lysed to get a single cell suspension. 50000 cells 50,000 cells were lysed, and followed by transposition reaction and purification, and PCR amplification. The library construction and sequencing were performed by NextSeq 500 platform.

overall_design: Briefly, the apical halves of 8 mandibular first molars of PN7.5 Runx2fl/fl control mice were collected in PBS on ice. Then the tissues were treated to obtain a single-cell suspension. 50,000 cells were lysed, and followed by transposition reaction and purification, and PCR amplification. The library construction and sequencing were performed by NextSeq 500 platform. The raw data was analyzed using Partek® Flow® software. Briefly, raw reads were aligned using Bowtie2 with the mm10 genome; MACS2 was used for detecting genomic enrichment regions; Runx2 binding motifs were analyzed using HOMER

first: Yang

last: Chai


title: Expression data from C. elegans wdr-23 mutants

status: Public on Sep 01 2020

city: Gainesville

first: Keith

last: Choe

state: Florida

address: 876 Newell Dr, 321 Bartram Hall

institute: University of Florida

summary:
The WDR-23 protein regulates the transcription factor SKN-1 directly.  C. elegans wdr-23 mutants have highly active SKN-1 and are stress resistant, long-lived, small, and reproduce poorly.
We used microarrays to measure global gene expression in wdr-23 mutants and to indentify genes regulated by SKN-1.

overall_design: RNA extraction and hybridization on Affymetrix microarrays was performed on three genetic backgrounds at the L4 larval stage.  The backgrounds were N2 (wildtype) with control RNAi, wdr-23(tm1817) mutants with skn-1 RNAi, and wdr-23(tm1817) mutants with control RNAi.

middle: P


title: Glioma escape signature and clonal development under immune pressure [microarray]

city: Hamburg

first: Krystian

middle: Daniel

last: Fita

address: Martinistrasse 52

department: Department of Neurosurgery

country: Germany

institute: University Medical Center Hamburg-Eppendorf

summary: Immunotherapeutic strategies are increasingly important in neuro-oncology and the elucidation of escape mechanisms which lead to treatment resistance is crucial. We investigated the impact of immune pressure on the clonal dynamics and immune escape signature by comparing glioma growth in immunocompetent versus immunodeficient mice. Glioma-bearing wildtype and PD-1-/- mice survived significantly longer than immunodeficient Pfp/Rag2-/- mice. While tumors in Pfp/Rag2-/- mice were highly polyclonal, immunoedited tumors in WT and PD-1-/- mice displayed reduced clonality with emergence of immune escape clones. Tumor cells in wildtype mice were distinguished by an interferon-gamma-mediated response signature with upregulation of genes involved in immunosuppression. Tumor-infiltrating stromal cells, which include macrophages/microglia, contributed even stronger to the immunosuppressive signature than the actual tumor cells. The identified murine immune escape signature was reflected in human patients and correlated with poor survival. In conclusion, cancer immune pressure profoundly shapes the clonal composition and gene regulation in malignant gliomas.

overall_design: Total RNA obtained from ex vivo GL261 tumors compared between WT and Pfp/Rag2 -/- mice, in vitro GL261 cell line and healthy mouse brain.

first: Cecile

middle: L

last: Maire

first: Malte

last: Mohme

first: Michael

last: Bockmayr

middle: D

first: Kristoffer

last: Riecken

first: Daniela

last: Börnigen

first: Malik

last: Alawi

first: Antonio

last: Failla

first: Katharina

last: Kolbe

first: Svenja

last: Zapf

first: Mareike

last: Holz

first: Katrin

last: Neumann

first: Lasse

last: Dührsen

first: Tobias

last: Lange

first: Boris

last: Fehse

first: Manfred

last: Westphal

last: Lamszus


title: Mycobacterium tuberculosis small RNA MTS1338 is required for bacterial virulence and regulates the expression of an immune-evasion molecules

city: Moscow

first: Artem

last: Grigorov

address: Ulitsa Miklukho-Maklaya, 16/10

department: Genomics and Postgenomic Technologies

country: Russia

institute: Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry

type: Expression profiling by high throughput sequencing

summary: Bacterial small non-coding RNAs play an essential role in adaptation to stresses. M. tuberculosis  small RNA MTS1338 was shown to contribute to successful persistence of mycobacteria within  host cells. In this study, we expressed MTS1338 in Mycobacterium smegmatis, and studied  transcriptomic and proteomic changes in the recombinant strain. We found that MTS1338  expression resulted to DosR regulon activation, metabolic changes characteristic for hypoxia,  mycolic acids synthesis, as well as rearrangement of iron metabolism. These metabolic changes  slow down bacterial growth in vitro, provide an increase of non-pathogenic M.smegmatis  survival in the infected THP-1macrophages, and also influence phagosome maturation.

overall_design: THP-1 Macrophages were infected with two strains of M. smegmatis. One strain contaned a vector that expresses M.tuberculosis small non-coding RNA MTS1338 (pMV_MTS1338) and other strain was a control and contained an empty vector (pMV_Empty). In each group, three independent biological samples were sequenced.

first: Elena

middle: G

last: Salina

first: Oksana

last: Bychenko

first: Yulia

last: Skvortsova

first: Tatyana

last: Azhikina


title: Checkpoint receptor TIGIT expressed on Tim-1+ B cells regulates tissue inflammation

city: Boston

first: Lloyd

last: Bod

state: MA

address: 60 Fenwood road

institute: Brigham and Women's Hospital and Harvard Medical School

summary: Tim-1, a phosphatidylserine receptor expressed on B cells induces IL-10 production by sensing apoptotic cells. Here we show that mice with B cell-specific Tim-1 deletion developed tissue inflammation in multiple organs including spontaneous paralysis with inflammation in the central nervous system (CNS). Transcriptomic analysis demonstrated that besides IL-10, Tim-1+ B cells also differentially expressed a number of co-inhibitory “checkpoint” receptors including TIGIT. Mice with B cell-specific TIGIT deletion developed spontaneous paralysis with CNS inflammation but with limited inflammation in other organs. Our findings suggest that Tim-1+ B cells are essential for maintaining self-tolerance and restraining tissue inflammation, and that Tim-1 dependent TIGIT expression on B cells is essential for maintaining CNS-specific tolerance. A possible critical role of AhR in regulating the B cell function is discussed, as we found that AhR is a top-ranked transcription factor expressed in Tim-1+ B cells and regulates their TIGIT and IL-10 expression.

overall_design: Samples from isolated Tim-1- and Tim-1+ B cells were processed with the SMART-Seq2 protocol (Picelli et al., 2013), and sequenced on Illumina Hi-Seq 2500. The paired-end 38bp reads sequenced (26.53M ± 1.62M pairs of reads per sample) for each of the 8 samples (4 Tim-1+ and 4 Tim-1- B cell samples derived from naïve WT mice) were aligned to the mouse mm10 UCSC reference genome using Tophat version 2.0.10 (Kim et al., 2013) with default settings (read alignment rate of 82.81% ± 0.44% properly mapped pairs per sample). Gene expression levels were quantified for 22,815 genes in the mouse mm10 UCSC reference gene annotations using Cuffquant in the Cufflinks suite version 2.2.1 (Trapnell et al., 2012). These levels were subsequently normalized across all 8 samples using Cuffnorm with default settings based on “geometric” normalization, which normalizes samples based on the median expression level in each sample, thus reporting normalized expression levels (FPKM: fragments per kilo bases of exons for per million mapped reads). Out of the 22,815 quantified reference genes, 15,519 genes were detected in at least one of the 8 samples at a normalized FPKM level > 0. Differential gene expression analysis comparing the 4 Tim-1+ and 4 Tim-1- B cell samples was performed by estimating the asymptotic t-test (Student's t-distribution) p-values, False Discovery Rate (FDR) (Benjamini and Hochberg) values and fold-changes using the ComparativeMarkerSelection module in GenePattern (Reich et al., 2006). Significantly differentially expressed genes were identified using the asymptotic t-test p-value ≤ 0.05 and absolute fold change ≥ 2 selection criteria. Heatmaps visualizing the normalized gene expression levels (zero mean centering and unit standard deviation scaling of the expression levels for each gene, followed by saturating these normalized gene expression levels at +1 and -1) were generated using GENE-E/Morpheus.

first: Sheng

last: Xiao

first: Nathalie

last: Pochet

first: Savithri

last: Balasubramanian Kota

first: Dan

last: Hu

first: Asaf

last: Madi

first: Jessica

last: Kilpatrick

first: Jingwen

last: Shi

first: Allen

last: Ho

first: Huiyuan

last: Zhang

first: Raymond

last: Sobel

first: Howard

last: Weiner

first: Terry

middle: B

last: Strom

first: Francisco

last: J Quintana

first: Nicole

last: Joller

first: Vijay

middle: K

last: Kuchroo


title: Glioma escape signature and clonal development under immune pressure [RNA-Seq]

first: Krystian

last: Fita

overall_design: Total RNA obtained from ex vivo GL261 tumors compared between bulk tumors versus sorted GFP+ tumor cells in WT and Pfp/Rag2 -/- mice, with in vitro GL261 GFP+ cell line and non GFP labelled tumors as controls.

first: Cecile

last: Maire

first: Malte

last: Mohme

first: Michael

last: Bockmayr

first: Kristoffer

last: Riecken

first: Daniela

last: Börnigen

first: Malik

last: Alawi

first: Antonio

last: Failla

first: Katharina

last: Kolbe

first: Svenja

last: Zapf

first: Mareike

last: Holz

first: Katrin

last: Neumann

first: Lasse

last: Dührsen

first: Tobias

last: Lange

first: Boris

last: Fehse

first: Manfred

last: Westphal

last: Lamszus


title: Transcriptional response of thyroid to iodide in wild-type and Nrf2 knockout mice

city: Lausanne

first: Dionysios

last: Chartoumpekis

address: Ave de la Sallaz 8

department: Service of Endocrinology Diabetes and Metabolism

country: Switzerland

institute: Lausanne University Hospital CHUV

summary: Analysis of differentially expressed mRNAs indicated that iodide overload upregulated inflammatory-, immune-, fibrosis- and oxidative stress-related pathways, including the Nrf2 pathway. Nrf2 KO mice showed a more pronounced inflammatory-autoimmune transcriptional response to iodide

overall_design: Thyroidal mRNA profiles after iodide exposure (0.05% NaI in their drinking water for 7 days) in C57Bl6j WT and Nrf2 knockout mice

first: Gerasimos

last: Sykiotis

first: Panos

last: Ziros

middle: V


title: Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and RBM7-/- HEK293 cell lines Transcriptomes and RBM7 targetted RNAs

city: Suita

first: Daisuke

last: Motooka

address: Yamadaoka

country: Japan

institute: Osaka University

summary: Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to compare NGS-derived  transcriptome profiling of Wild Type and RBM7-/- HEK293 cell lines and RNA immunoprecipitation (RIP) of Flag-tagged RBM7-expressing HEK293 cells

overall_design: Examination of RIP samples and total RNA from WT and Rbm7−/− HEK293 cells.

first: Kiyoharu

last: Fukushima

first: Shota

last: Nakamura

first: Fuminori

last: Sugihara

first: Takashi

last: Satoh


title: The protein modifier SUMO is critical for Arabidopsis shoot meristem maintenance at warmer ambient temperatures

city: Amsterdam

first: Martijs

last: Jonker

address: Science Park 904

department: Microarray Department

country: Netherlands

institute: University of Amsterdam

summary:
Short periods of heat (>37°C) are extremely damaging to non-acclimated plants and their capacity to acclimate to and recover from heat stress is a key parameter for their survival and longevity. To acclimate, the Heat Shock transcription Factor A1 (HSFA1) subfamily activates a transcriptional response that resolves the heat stress-induced protein damage. Importantly, HSFA1 activity is also critical for Arabidopsis to withstand sustained warmer periods of 28°C, a non-detrimental condition that triggers a thermomorphogenesis response. We find that SUMO, a protein modification whose adduct levels increase as a result of acute heat stress in eukaryotes, is also critical for plant longevity during warmer periods, in particular for shoot meristem development. The known E3 and E4 SUMO ligases (SIZ1, HPY1/MMS21, PIAL1/2) were not essential to endure these warmer periods, alone or in combination. Thermo-lethality was also not seen when plants lacked certain SUMO proteases (ESD4, OTS1/OTS2, SPF1/SPF2 combined) or when SUMO chain formation was blocked. Furthermore, SUMO thermo-resilience is not connected to the autoimmune phenotype found in the corresponding SUMO knockdown and a SIZ1 loss-of-function mutant. As acquired thermotolerance was normal in the SUMO knockdown mutant, we thus conclude that the role of SUMO in heat acclimation differs from that of HSFA1 and SIZ1. Combined, this study reveals that SUMO appears to be critical for shoot meristem integrity during warmer periods.
This experiment we have examined how gene expression is affected in two SUMO mutants (siz1-2; sumo1 amiR-SUMO1 [aka. sumo1/2KD] ), and a HsfA1a,b,d triple mutant, when the plants are placed at 28C constant ambient temperature, which is a condition normally used to induce thermomorphogenesis. We used as control the pad1-4 background, as the siz1-2 and sumo1/2KD mutants normally suffer from constitutive defence signalling due hyperaccumulation of SA, which is suppressed by introgression of pad4 in these backgrounds.

overall_design: Three biological replicates per samples; Arabidopsis plants were grown at 22C (t=0) and shifted to 28C (t=24hrs, and t=96 hrs post shift); In total four lines were compared: pad4-1 ('negative' control), siz1-2 pad4-1 double mutant, and sumo1-1 amiR-SUMO2 pad4-1  triple mutant, and eTK triple mutant.

first: Valentin

last: Hammoudi

first: Bas

last: Beerens

first: Georgios

last: Vlachakis

first: Sayantani

last: Chatterjee

first: Like

last: Fokkens

first: Manuel

last: Arroyo-Mateos

first: Paul

middle: F

last: Wackers

middle: J


title: Immune Gene Expression Profile of Tr1 Skewed Tregs

first: Todd

middle: M

last: Brusko

state: FL

address: 1275 Center Dr BMSB J515

department: Pathology

summary: IL-12 and IL-18 synergize to promote TH1 responses and have been implicated as accelerators of autoimmune pathogenesis in type 1 diabetes (T1D). We therefore investigated the influence of these cytokines on phenotype and function of immune cells that are involved in disease progression. To understand how IL-12 and IL-18 may synergize to impair Treg function and phenotype, we conducted transcriptional profiling of Treg expanded under normal conditions or in the presence of IL-12 and IL-18. This analysis revealed increased expression of IFNG, GZMB, GZMA, and IL18RAP and decreased FOXP3 in Tregs expanded with IL-12 and IL-18.

overall_design: Transcriptional profiling of Treg expanded under normal conditions or in the presence of IL-12 and IL-18.  3 samples, 2 conditions each

first: Joseph

middle: W

last: Dean

first: Leeana

last: Peters

first: Christopher

middle: A

last: Fuhrman

first: Howard

middle: R

last: Seay

first: Amanda

last: Posgai

first: Scott

middle: E

last: Stimpson

first: Maigan

first: Daniel

last: Perry

first: Wen-I

last: Yeh

first: Brittney

middle: N

last: Newby

first: Michael

last: Haller

first: Andrew

last: Muir

first: Mark

last: Atkinson

first: Clayton

last: Mathews


title: Genome-wide association between DNA methylation and fiber type composition in skeletal muscle

first: Rinat

middle: I

last: Sultanov

address: Malaya Pirogovskaya, 1a

institute: FRCC of Physical Chemical Medicine of the FMBA of the Russian Federation

summary: Genome wide DNA methylation profiling of skeletal muscle

overall_design: Bisulphite converted DNA from the 47 samples were hybridised to the Illumina Infinium Human Methylation EPIC

first: R

first: E

last: Generozov


city: Qingdao

first: Yong

last: Zhao

state: Shandong

address: 700 Changcheng Road

department: College of Life Sciences

country: China

institute: Qingdao Agricultural University

type: Methylation profiling by high throughput sequencing

summary: The aim of this study was to explore  DNA methylation status in the ovarian samples of two different population of goat (high fertility and low fertility groups). We profiled DNA methylation sequencing of the goat ovarian samples  from high fertility group (>three babys per birth) and low fertility group (< two babys per birth).

first: Beining

last: Kang

first: Lingjiang

last: Min


title: Microarray-based miRNA expression in fluid shear stress-stimulated collateral blood vessels

city: Bad Nauheim

first: Kerstin

last: Troidl

address: Ludwigstr. 43

institute: Max-Planck-Institute for Heart and Lung Research

summary:
collateral tissue from rats was isolated to assess miRNA expression changes between quiescent and growing blood vessels
A total of ten total RNA samples were processed. 800 ng of total RNA from each sample was subjected to two different labeling reactions (AF665 and AF546). Two differentially labeld reactions were combined into one tube and loaded onto the arrays.

platform_organism:
Caenorhabditis elegans
Drosophila melanogaster
Danio rerio
Homo sapiens
Mus musculus
Rattus norvegicus

overall_design: Two condition experiment: growing vs quiescent; 5 biological replicates in each group.

first: Wilma

last: Schierling


title: Pleiotrophic roles of VEGF during neonatal thymic development

first: Gay

middle: Miriam

last: Crooks

state: California

address: 610 Charles E. Young Drive, East

department: Pathology & Laboratory Med

institute: University of California Los Angeles

summary: In this report, we show that VEGF-blockade in neonates induces rapid changes in all three stromal compartments of the thymus, and that these changes recapitulate the maturation of these cell types seen during the transition from the neonatal to the young adult thymus. These changes in the microenvironment occur prior to the loss of thymocytes, and include not only a loss of cTECs relative to mTECs but also depletion of CD146+CD140aneg cells, a novel population of pericytes we have identified in the neonatal thymus. Importantly, we show that based on receptor expression and receptor blockade, VEGF drives the fetal/neonatal phenotype of the thymus through distinct VEGFR2- and NRP1-dependent pathways in thymic endothelial and mesenchymal cells respectively. In addition, receptor signaling induces changes in critical pathways affecting the function of all three compartments, with consequences on thymopoieisis.

overall_design: To investigate the role of VEGF in the neonatal thymic microenvironment, we treated P1 mice with either Aflibercept (aka VEGF-Trap), a VEGFR1/R2 fusion protein which acts as a decoy molecule for all VEGF-A isoforms, or an hFC control antibody. Thymi were harvested 2 days post injection (P3), digested and depleted from thymocytes (CD45 magnetic depletion). Thymic stromal cells were isolated by fluorescence activated cell sorting as follows: within the stromal cell gate (CD3-CD4-CD8-CD45-Ter119- cells), thymic epithelial cells (TECs) were identified by their expression of the epithelial cell adhesion molecule (EpCAM) and were further subdivided into cortical thymic epithelial cells (cTECs) and medullary thymic epithelial cells (mTECs) based on high to low/negative Ly51 expression, respectively. Platelet endothelial cell adhesion molecule (PECAM-1 aka CD31) expression allowed the isolation of endothelial cells. By excluding endothelial, epithelial and hematopoietic cells, we next examined the mesenchymal population of the thymus. CD146+ and CD140a+ mesenchymal cells were isolated within the EpCAMnegCD31neg stromal cell gate.

first: Stephanie

middle: C

last: de Barros

first: Batul

middle: T

last: Suterwala

first: Chongbin

last: He

first: Shundi

last: Ge

first: Garrett

last: Blumberg

first: Kenneth

last: Kim

first: Sam

last: Klein

first: Yuhua

last: Zhu

first: Brent

last: Chick

first: Xiaoyan

last: Wang

first: David

last: Casero


title: Identification of a Four-lncRNA Signature Predicting Prognosis of Patients with NSCLC

city: GZ

first: Rui-qi

last: WANG

state: Guangdong

address: 651 DONGFENG EAST ROAD, GUANGZHOU 510060, P. R. CH

department: state key lab

institute: SUN YAN-SEN UNIVERSITY CANCER CENTER

summary: LncRNA expression profiles were sucessfully contructed in 370 patients with NSCLC and 198 corresponding adjacent noncancerous tissues using a custom microarray containing probes for 2412 lncRNAs and an outcome prediction of  lncRNA signature was successfully identified for predicting prognosis in NSCLC.

overall_design: In the study presented here, a consecutively operated, well-defined cohort of 370 NSCLC cases, followed up more than three ears, was used to acquire lncRNA expression profiles of 2412 lncRNAs and identify a lncRNA signature, leading to the successfully for predicting prognosis in NSCLC.

first: Xiao-ran

last: Long

first: Huiyun


title: Effect of QTRT2 KO  on mitochondrial translation

status: Public on Aug 28 2020

city: Hirosawa Wako city, Saitama prefecture, Japan

first: Shintaro

last: Iwasaki

department: Cluster for Pioneering Research

institute: RIKEN

type: Non-coding RNA profiling by high throughput sequencing

summary: Mitochondria are organelles that generate most of the energy in eukaryotic cells in the form of ATP via oxidative phosphorylation in eukaryote. Twenty-two species of mitochondrial (mt-)tRNAs encoded in mtDNA are required to translate essential subunits of the respiratory chain complexes involved in oxidative phosphorylation. mt-tRNAs contain post-transcriptional modifications introduced by nuclear-encoded tRNA-modifying enzymes. These modifications are required for deciphering genetic code accurately, as well as stabilizing tRNA. Loss of tRNA modifications frequently results in severe pathological consequences. We performed a comprehensive analysis of post-transcriptional modifications of all human mt-tRNAs, including 14 previously-uncharacterized species, and revised the modification status of some of the previously studied species. In total, we found 17 kinds of RNA modifications at 137 positions (8.7% in 1,575 nucleobases) in 22 species of human mt-tRNAs. An up-to-date list of 34 genes responsible for human mt-tRNA modifications are provided. We here demonstrated that both QTRT1 and QTRT2 are required for biogenesis of queuosine (Q) at position 34 of four mt-tRNAs. Our results provide insight into the molecular mechanisms underlying the mitochondrial decoding system, and could help to elucidate the molecular pathogenesis of human mitochondrial diseases caused by aberrant tRNA modifications.

overall_design: Ribosome profiling and RNA-seq

first: Ding

last: Ma

first: Hironori

last: Saito

first: Mari

last: Mito

first: Takeo

last: Suzuki

first: Tsutomu


title: RNA-sequencing analysis of naive and primed hPSCs in static and stirred suspension conditions

city: Calgary

first: Leili

last: Rohani

state: Alberta

address: 405J, 1919 University Drive, NW

department: Biochemistry and Molecular Biology

country: Canada

institute: University of Calgary

summary: We have pioneered human pluripotent stem cell (hPSC) manufacturing in stirred suspension bioreactors. Cell therapies require large numbers of quality-controlled hPSCs yet technologies are limited in their ability to efficiently grow and scale clinically-viable hPSCs. We report here that naive hPSCs exhibit superior growth in suspension bioreactors compared to their primed counterpart. Naive hPSCs exhibited a shorter lag phase, and grew into more uniform, homogenous aggregates. Compared to static culture, gene expression analyses revealed that the bioreactor environment promoted the upregulation of naïve- and downregulation of primed-associated transcripts in both primed and naive hPSCs. Bioreactor-cultured naive hPSCs similarly showed more hypomethylated DNA and less primed hPSC-associated surface protein marker compared to statically-cultured naive hPSCs. Gene expression, epigenetic, and cell surface protein marker analyses all suggest that the bioreactor environment promotes the transition from primed-to-naive pluripotent state. Our research shows that reprogramming conventional hPSCs to the naive pluripotent state enhances hPSC manufacturing.

overall_design: RNA-seq experiment was performed on naïve and primed human pluripotent stem cells from static culture and stirred suspension bioreactors. The aliquots of static-cultured cells for each naïve (P4) and primed hPSC sample were collected for RNA-seq analysis before inoculating them into stirred suspension bioreactors. Those cells were counted as “static-cultured donor cells”. The aggregates of day four post-inoculation for each bioreactor-cultured naïve and primed hPSC were collected for RNAseq analysis. We performed a pairwise comparison for transcriptomic analysis: 1) bioreactor-cultured naïve hPSCs and their statically-cultured naïve counterpart, 2) bioreactor-cultured primed hPSCs and their statically-cultured primed cognate. This study was exploratory (n =1), no p-values are given and therefore the simple normalized expression (normalization) value comparison was done as Reads per Kilobase per Million (RPKM). Expression levels for each gene were determined by RPKM in each sample. The RPKM values for each gene within each sample were compared to generate fold change and log2 fold changes values between the samples in a pair-wise comparison (a python script was used to compare RPKM values and to generate the foldchange tables). For the pairwise comparison between static- and bioreactor-cultured naïve hPSCs, the fold change > 1.95 was measured relative to static-cultured naïve hPSCs to derive differentially expressed genes among those samples. For the pairwise comparison between static- and bioreactor-cultured primed hPSCs, the fold change > 1.95 was measured relative to static-cultured primed hPSCs to derive differentially expressed genes among those samples. The differentially expressed genes (fold change > 1.95) among the pair-wise comparisons were used for enriched gene ontology (GO) terms (Molecular Function (MF), Biological Process (BP), and Biological Pathway (rea)) through g:Profiler.

first: Derrick

last: Rancourt


title: High throughput RNA-sequencing identifies Differentially Expressed Genes (DGS) of control and shATXN7L3 HCCLM3 cells

status: Public on Aug 30 2020

city: Shenyang

first: Ning

last: Sun

state: Liao Ning Province

address: No.77, Puhe Road

department: Institute of life sciences

institute: China Medical University

summary:
Purpose: High throughput RNA-seq has provided an efficient and convenient access to screening substantial differentially expressed genes (DEGs). The goals of this study are to screen out ATXN7L3-regulated DEGs of HCC cells by utilizing RNA-sequencing and to furtherly identify specific biological function-related gene subset from ATXN7L3-regulated DEGs
Method: Firstly, HCCLM3 cells were seperated into two groups and respectively transfected with lentivirus-delivered shATXN7L3 and shcontrol. We cultured these cells with DMEM containing 10% FBS serum. 48 hours later, cells of two groups were harvested by Trizol RNA isolater and analyzed by High througput RNA-sequencing.
Conclusion: Using next generation high throughput RNA-sequencing technology, we identified a total of 3831 genes were significantly changed after ATXN7L3 knockdown. There were 1914 ATXN7L3 positive-regulated genes and 1917 ATXN7L3 negative-regulated genes. Then we focused on top DEGs using ±0.8-log2(fold change) as a cutoff for statistic analysis. Furtherly, 246 positive-regulated DEGs and 187 negative-regulated DEGs were identified. Finally, pathway and process enrichment analysis identified a series of cell growth-related DEGs.

overall_design: Cellular mRNA sequencing of shcontrol and shATXN7L3 HCCLM3 cells

first: Yue

last: Zhao


title: Comparison of the expression profiles of 630E strain and a sig54 mutant after 6h of growth

city: Paris

first: Monot

last: Marc

address: 25, rue du docteur roux

department: Genomes and Genetics

country: France

institute: Institut Pasteur

summary: Transcriptional profiling of C. difficile 630E strain vs. a sig54 mutant after 6h of growth in TY.

overall_design: Two-conditions experiments, 630E strain 6h vs. sig54 mutant 6h, 4 biological replicates for each condition.

first: Thomas

last: Dubois

first: Marc

last: Monot


title: RNA sequencing of nos-gal4>phf7ey Drosophila ovaries

first: Mitzi

last: Kuroda

address: 77 Avenue Louis Pasteur

department: Genetics

institute: Brigham and Women's Hospital/Harvard Medical School

summary: Next generation sequencing of mRNA for nos-gal4>phf7ey Drosophila ovaries

overall_design: Total mRNA profiles for whole ovaries in 10 day old Drosophila females

first: Anne

last: Smolko

first: Helen

last: Salz


title: Expression data of TS cells with inactive MAP3K4 (TSKI4 cells) and TSKI4 cells re-expressing REL (TSKI4R cells)

city: Memphis

first: Amy

last: Abell

state: TN

address: 3774 Walker Ave

department: Biological Sciences

institute: The University of Memphis

summary:
Trophoblast stem cells lack MAP3K4 activity (TSKI4 cells) switch from epithelial phenotype to intermediate phenotype. Loss of epithelial phenotype is due to the loss of CBP histone acetyltransferase activity and the gain of histone deacetylase HDAC6 expression and activity. In our work, we identify a small network of 183 genes whose expression is co-regulated by MAP3K4, CBP, and HDAC6. Further, we define the key role of one of these co-regulated genes, Rel, in inducing epithelial phenotype in intermediate TSKI4 cells.
We used microarrays to compare gene expression betweeen mesenchymal TS cells with inactive MAP3K4 (TSKI4 cells) and epithelial TSKI4 cells re-expressing REL (TSKI4R cells)

overall_design: TS cells with inactive MAP3K4 (TSKI4 cells) and TSKI4 cells re-expressing REL (TSKI4R cells) were cultured for 4 days under stemness conditions. Total RNA was extracted and hybridized on Affymetrix microarrays.

first: Noha

last: Shendy

first: Deepthi

last: Raghu

first: Sujoy

last: Roy

first: Charles

middle: H

last: Perry

first: Adiba

last: Safi

first: Miguel

last: Branco

first: Ramin

last: Homayouni


title: Coordination of Rel Expression by MAP3K4, CBP, and HDAC6   Controls TS cell Phenotype (RNA-seq)

summary: Trophoblast stem cells lack MAP3K4 activity (TSKI4 cells) switch from epithelial phenotype to intermediate phenotype. Loss of epithelial phenotype is due to the loss of CBP histone acetyltransferase activity and the gain of histone deacetylase HDAC6 expression and activity. In our work, we identify a small network of 183 genes whose expression is co-regulated by MAP3K4, CBP, and HDAC6. Further, we define the key role of one of these co-regulated genes, Rel, in inducing epithelial phenotype in intermediate TSKI4 cells.

overall_design: RNA-seq samples from TSWT, TSWTCBsh,TSKI4, TSKI4H6sh

first: Deepth


title: Heterogeneity in antiviral responses in the upper respiratory tract promotes differential non-lytic clearance of influenza viruses

city: Durham

first: Nicholas

last: Heaton

state: NC

address: 213 Research Drive, 426 CARL Building

department: Molecular Genetics and Microbiology

institute: Duke University School of Medicine

summary: Analysis of RNA from different cells in the olfactory epithelium of mice after influenza virus infection.

overall_design: Olfactory epithelium of mice after infuenza virus infection

middle: S

first: Rebekah

last: Dumm


title: Maintenance DNA methylation is essential for regulatory T cell development and stability of suppressive function [gene expression/RNA-seq]

status: Public on Aug 27 2020

city: Chicago

first: Benjamin

middle: D.

last: Singer

state: Illinois

address: 303 E. Superior St., Simpson Querrey 5th floor

institute: Northwestern University Feinberg School of Medicine

summary: Regulatory T (Treg) cells require Foxp3 expression and induction of a specific DNA hypomethylation signature during development, after which Treg cells persist as a self-renewing population that regulates immune system activation. Whether maintenance DNA methylation is required for Treg cell lineage development and stability and how methylation patterns are maintained during lineage self-renewal remain unclear. Here, we demonstrate that the epigenetic regulator Uhrf1 is essential for maintenance of methyl-DNA marks that stabilize Treg cellular identity by repressing effector T cell transcriptional programs. Constitutive and induced deficiency of Uhrf1 within Foxp3+ cells resulted in global yet non-uniform loss of DNA methylation, derepression of inflammatory transcriptional programs, destabilization of the Treg cell lineage, and spontaneous inflammation. These findings support a paradigm in which maintenance DNA methylation is required in distinct regions of the Treg cell genome for both lineage establishment and stability of identity and suppressive function.

overall_design: Treg cells (CD25hiFoxp3-YFP+) from 8-week-old female Uhrf1fl/flFoxp3+/YFP-Cre (Uhrf1 chimeric knockout) or Uhrf1+/+Foxp3YFP-Cre/YFP-Cre (control) mice with n = 4  biological replicates per group; labeled Treg cells (Foxp3-GFP+tdTomato+), ex-Foxp3 cells (Foxp3-GFP–tdTomato+), and post-tamoxifen Treg cells (Foxp3-GFP+tdTomato–) from Uhrf1fl/flFoxp3GFP-CreERT2Rosa26SorCAG-tdTomato (a.k.a. iUhrf1fl/fl) or Uhrf1+/+Foxp3GFP-CreERT2Rosa26SorCAG-tdTomato (a.k.a iUhrf1+/+ control) mice with n = 6 (iUhrf1fl/fl) and 5 (iUhrf1+/+) biological replicates per cell type in 8-week-old mice that received tamoxifen chow for 2 weeks followed by standard chow for 4 weeks

middle: D


title: Maintenance DNA methylation is essential for regulatory T cell development and stability of suppressive function [DNA methylation/RRBS]

overall_design: Treg cells (CD25hiFoxp3-YFP+) from 8-week-old female Uhrf1fl/flFoxp3+/YFP-Cre (Uhrf1 chimeric knockout) or Uhrf1+/+Foxp3YFP-Cre/YFP-Cre (control) mice with n = 4  biological replicates per group; labeled Treg cells (Foxp3-GFP+tdTomato+) and ex-Foxp3 cells (Foxp3-GFP–tdTomato+) from Uhrf1fl/flFoxp3GFP-CreERT2Rosa26SorCAG-tdTomato (a.k.a. iUhrf1fl/fl) or Uhrf1+/+Foxp3GFP-CreERT2Rosa26SorCAG-tdTomato (a.k.a iUhrf1+/+ control) mice with n = 4  biological replicates per cell type in 8-week-old mice that received tamoxifen chow for 2 weeks followed by standard chow for 4 weeks


title: Maintenance DNA methylation is essential for regulatory T cell development and stability of suppressive function

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

overall_design: Refer to individual Series


title: Early-life environmental enrichment generates persistent individualized behavior in mice

status: Public on Aug 26 2020

city: Dresden

first: Gerd

last: Kempermann

state: Saxony

address: Tatzberg 41

institute: German Center for Neurodegenerative Diseases (DZNE)

summary: Analysis of hippocampal DNA methylation changes that were induced by environmental enrichment and maintained for long-term after withdrawal from the enriched environment.

overall_design: DNA methylation patterns were compared between three groups of mice. One group was housed in standard housing conditions for 6 months, one group in environmental enrichment for six months, and a third group was housed for three months in environmental enrichment followed by three months of standard housing. DNA methylation profiling was performed by Reduced Representation Bisulfite Sequencing of micro-dissected dentate gyrus tissue.

first: Sara

last: Zocher


title: A dichotomy of gene regulatory associations during the activated B-cell to plasmablast transition [Microarray]

status: Public on Aug 24 2020

city: York

first: Matthew

middle: Anthony

last: Care

state: North Yorkshire

address: Wentworth Way

department: Department of Biology

country: United Kingdom

institute: The University of York

summary: The activated B-cell (ABC) to plasmablast transition is the cusp of antibody secreting cell (ASC) differentiation but is incompletely defined. We apply expression time-courses, parsimonious gene correlation network analysis, and ChIP-seq to explore this in human cells. The transition initiates with input signal loss leading within hours from cell growth dominant programs to enhanced proliferation, accompanied from 24h by ER-stress response, secretory optimization and upregulation of ASC features. Clustering of genomic occupancy for ASC transcription factors (TFs) IRF4, BLIMP1 and XBP1 with CTCF and histone marks defines distinct patterns for each factor in plasmablasts. Integrating TF-associated clusters and modular gene expression identifies a dichotomy: XBP1 and IRF4 significantly link to gene modules induced in plasmablasts, but not to modules of repressed genes, while BLIMP1 links to modules of ABC genes repressed in plasmablasts but is not significantly associated with modules induced in plasmablasts. Pharmacological inhibition of the G9A (EHMT2) histone-methytransferase, a BLIMP1 co-factor that catalyzes repressive H3K9me2 marks, leaves functional ASC differentiation intact but de-represses ABC-state genes. Thus, in human plasmablasts IRF4 and XBP1 emerge as the dominant association with ASC gene expression, while BLIMP1 links to repressed modules with particular focus in repression of the B-cell activation state.

overall_design: B-cells were isolated from the peripheral blood of 3 adult donors and differentiated in vitro until day 6 (see individual samples for culture conditions)

first: Reuben

last: Tooze

first: Mario

last: Cocco


title: A dichotomy of gene regulatory associations during the activated B-cell to plasmablast transition [ChIP-seq]

overall_design: ChIP-Seq; 28 samples


title: TWIST1 direct targets during embryonic stem cell differentiation [ChIP-seq]

status: Public on Aug 23 2020

city: WESTMEAD

first: PIERRE

last: OSTEIL

state: NSW

address: 214 Hawkesbury Road

department: Embryology

country: Australia

institute: CMRI

summary: To understand TWIST1 function in early neural development, we analyzed genome-wide binding sites of TWIST1 in mouse embryonic stem cells (mESC) induced for neurogenic differentiation by chromatin immunoprecipitation, followed by high-throughput sequencing

overall_design: A2loxcre mESC cell line with doxycycline-inducible Flag-Twist1 under Twist1-/- background was generated (Twist1-/-; Twist1 O/E). mESC with genotype Twist1-/-; Flag-Twist1 O/E and Twist1-/- were differentiated into neuroepithelium cells for 3 days following established protocol. Neurogenic differentiation was initiated by plating mESC in AggreWells (1x 10^6 per well) using feeder independent mESC. Colonies were then lifted from AggreWells and grown in suspension in Neurogenic Differentiation Media supplemented with 15% FBS with gentle shaking. Doxycycline was added 16 hours to induce FLAG-TWIST1 induction before cells were collected for ChIP-seq analysis. Control cell line without inducible cassette (Twist1-/-) were treated under the same condition. Four biological replicate experiments were performed.

first: Xiaochen

last: Fan

first: Pierre

last: Osteil

first: Ashley

last: Waardenberg


title: Brd4-bound enhancers drive cell intrinsic sex differences in glioblastoma

status: Public on Aug 25 2020

city: St. Louis

first: Zongtai

last: Qi

state: Missouri

address: 4515 Mckinley Ave

institute: Washington University in St Louis


title: Senataxin, the helicase mutated in AOA2, plays a role in autophagy regulation

city: Davis

first: Frederic

last: Chedin

state: CA

address: 1 Shields Avenue

department: MCB

institute: UC Davis

summary: SETX (senataxin) is an RNA/DNA helicase that has been implicated in transcriptional regulation and the DNA damage response through resolution of R-loop structures. Mutations in SETX result in either of two distinct neurodegenerative disorders. SETX dominant mutations result in a juvenile form of amyotrophic lateral sclerosis (ALS) called ALS4, whereas recessive mutations are responsible for ataxia called ataxia with oculomotor apraxia type 2 (AOA2). How mutations in the same protein can lead to different phenotypes is still unclear. To elucidate AOA2 disease mechanisms, we first examined gene expression changes following SETX depletion. We observed the effects on both transcription and RNA processing, but surprisingly observed decreased R-loop accumulation in SETX-depleted cells. Importantly, we discovered a strong connection between SETX and the macroautophagy/autophagy pathway, reflecting a direct effect on transcription of autophagy genes. We show that SETX depletion inhibits the progression of autophagy, leading to an accumulation of ubiquitinated proteins, decreased ability to clear protein aggregates, as well as mitochondrial defects. Analysis of AOA2 patient fibroblasts also revealed a perturbation of the autophagy pathway. Our work has thus identified a novel function for SETX in the regulation of autophagy, whose modulation may have a therapeutic impact for AOA2.

overall_design: DRIP-seq was performed on asynchronous U87 cell lines to map R-loop formation genome-wide, 50bp single-reads on HiSeq 2500: 2 replicates for WT and 2 replicates for SETX knockdowns.

first: Patricia

last: Richard

first: Shuang

last: Feng

first: Wencheng

last: Li

first: Paola

last: Rinchetti

first: Ubayed

last: Muhith

first: Juan

last: Irizarry-Cole

first: Katharine

last: Stolz

first: Lionel

last: Sanz

first: Stella

last: Hartono

first: Mainul

last: Hoque

first: Saba

last: Tadesse

first: Hervé

last: Seitz

first: Francesco

last: Lotti

first: Michio

last: Hirano

first: Bin

last: Tian

first: James

middle: L

last: Manley


title: MiR-302b as a Combinatorial Therapeutic Approach to Improve Cisplatin Chemotherapy Efficacy in Human Triple Negative Breast Cancer

status: Public on Aug 21 2020

city: Mexico

first: Sandra

middle: Lorena

last: Romero-Cordoba

state: Mexico

address: Periférico Sur No. 4809, Col. Arenal Tepepan, Delegación Tlalpan. México, D.F.

department: Cancer Genomics

country: Mexico

institute: National Instute of Genomic Medicine

summary:
Chemotherapy is still the standard–of-care for triple-negative breast cancers (TNBCs). Here, we investigated miR-302b as therapeutic tool to enhance cisplatin sensitivity in vivo and unraveled the molecular mechanism.
Material and method: TNBC xenografted mice were treated with miR-302b or control, alone or with cisplatin. Genome-wide transcriptome analysis and independent-validation of ITGA6 expression was assessed on mice tumor samples. SiRNA-knockdown of ITGA6 was performed to evaluate cisplatin response in vitro. Further, potential transcription factors of ITGA6 (E2F1, E2F2 and YY1) were explored to define the miRNA molecular mechanism. MiR-302b expression was also assessed in TNBC patients treated with chemotherapy.
Results: MiR-302b-cisplatin combination significantly impaired tumor growth versus control, through indirect ITGA6 downregulation. Indeed, ITGA6 was downmodulated in mice treated with miR-302b-cisplatin and ITGA6 silencing increased drug sensitivity in TNBC cells. In silico analyses and preclinical assays pointed out the regulatory role of E2F family and YY1 on ITGA6 expression under miR-302b-cisplatin treatment. Finally, miR-302b enrichment correlates with better overall survival in 118 TNBC patients.
Conclusion: MiR-302b can be exploited as a new therapeutic tool to improve the response to chemotherapy, modulating E2F family, YY1 and ITGA6 expression. Moreover, miR-302b could be defined as new prognostic factor in TNBC patients.

overall_design: Through in vivo and in vitro preclinical models and genome-wide mRNA profiling, we investigated the mechanism behind miR-302b activity in combination with cisplatin and identified a biological axis involving Integrin Subunit Alpha 6 (ITGA6), Yin Yang 1 (YY1) and the E2F family, which resulted downmodulated by the combinational treatment of miR-302b and cisplatin.

first: Alessandra

last: Cataldo

first: Ilaria

last: Plantamura

first: Giulia

last: Cosentino

first: Alfredo

last: Hidalgo-Miranda

first: Elda

last: Tagliabue

first: Marilena

middle: V

last: Iorio


title: Single-Cell Transcriptome Profiling Reveals β Cell Maturation in Stem Cell-Derived Islets after Transplantation

first: Jeffrey

middle: Robert

last: Millman

state: MO

address: 660 South Euclid

institute: Washington University in St. Louis

summary: SC-β cells and native β cells from cadaveric islets have different transcriptome profiles. In this study, we demonstrate that SC-β cells can acquire mature gene signatures via transplantation in mice for 6 months. MAFA, a marker for mature β cells, is expressed and resulted in significant improvements in insulin release and glucose regulation in diabetic mice.

overall_design: Stem cell derived islets were generated and transplanted into the kidney capsule of each diabetic mouse. We harvested the kidneys with SC islets from mice at 6 months post-transplantation and excised graft sections for tissue dissociation. We used the mouse cell depletion kit to enrich human cells from the extracted graph cell suspension. Isolated and enriched human cell samples were then submitted for single cell RNA sequencing. We performed this with 5 mice, including SC islet cells generated from the Hues8 and IPS lines and pre-transplated in Vitro Stage 6 control.

first: Punn

last: Augsornworawat

first: Kristina

middle: G

last: Maxwell


title: SOX1 is required for the specification of rostral hindbrain neural progenitor cells from human embryonic stem cells

city: Stanford

first: Zhuoqing

last: Fang

address: 300 Pasteur Drive

institute: Stanford

summary: Region-specific neural progenitor cells (NPCs) can be generated from human embryonic stem cells (hESCs) through modulating signaling pathways. However, how intrinsic transcriptional factors contribute to the neural regionalization are not well characterized. Here, we generate region-specific NPCs from hESCs and find that SOX1 is highly expressed in NPCs with the rostral hindbrain identity. Moreover, we find that OTX2 inhibits SOX1 expression, displaying exclusive expression between the two factors. Furthermore, SOX1 knockout (KO) leads to up-regulation of midbrain genes and down-regulation of rostral hindbrain genes, indicating that SOX1 is necessary for specification of rostral hindbrain NPCs. Our SOX1 ChIP-sequencing analysis reveals that SOX1 binds to the distal region of GBX2 to activate its expression. Overexpression of GBX2 largely abrogates SOX1-KO induced aberrant gene expression. Taken together, this study uncovers previously unappreciated role of SOX1 in early neural regionalization and provides new information for the precise control of the OTX2/GBX2 interface.


title: Brd4-bound enhancers drive cell intrinsic sex differences in glioblastoma.

first: Zongtai

last: Qi

summary: We report the application of Calling Card sequencing technology for high-throughput profiling of Brd4-bound enhancers in male and female mouse glioblastoma.

overall_design: Examination of Brd4-bound enhancer in male and female astrocytes.

first: Najla

last: Kfoury


title: Sexually dimorphic characteristics of the small intestinal transcriptome of Fabp-gene ablated mice

status: Public on Aug 19 2020

city: Ste-Anne-de-Bellevue

first: Luis

middle: B

last: Agellon

state: QC

address: 21111 Lakeshore Road

department: School of Human Nutrition

institute: McGill University

summary: Analysis of whole small intestine tissue of 10-week old C57BL/6J mice, C57BL/6J mice with the disruption of either Fabp2 gene or Fabp6 gene, and C57BL/6J mice with the simultaneous disruption of Fabp2 and Fabp6 genes. Fabp2 and Fabp6 are two types of fatty acid binding proteins expressed primarily in the small intestine.

overall_design: Small intestine were isolated from ten-week old WT and Fabp-disrupted male and female mice fed a standard lab diet and subjected to gene expression profiling using microarray

first: Yiheng

last: Chen


title: Comparative transcriptome analysis of human skeletal muscle in response to cold acclimation and exercise training in human volunteers. [A391]

status: Public on Aug 18 2020

city: Wageningen

first: Guido

last: Hooiveld

address: HELIX, Stippeneng 4

department: Div. Human Nutrition & Health

institute: Wageningen University

summary: Background: Cold acclimation and exercise training were previously shown to increase peripheral insulin sensitivity in human volunteers with type 2 diabetes. Although cold is a potent activator of brown adipose tissue, the increase in peripheral insulin sensitivity by cold is largely mediated by events occurring in skeletal muscle and at least partly involves GLUT4 translocation, as is also observed for exercise training. Results: To investigate if cold acclimation and exercise training overlap in the molecular adaptive response in skeletal muscle, we performed transcriptomics analysis on vastus lateralis muscle collected from human subjects before and after 10 days of cold acclimation, as well as before and after a 12-week exercise training intervention. Methods: Cold acclimation altered the expression of 756 genes (422 up, 334 down, P<0.01), while exercise training altered the expression of 665 genes (444 up, 221 down, P<0.01). Principal Component Analysis, Venn diagram, similarity analysis and Rank–rank Hypergeometric Overlap all indicated significant overlap between cold acclimation and exercise training in upregulated genes, but not in downregulated genes. Overlapping gene regulation was especially evident for genes and pathways associated with extracellular matrix remodeling. Interestingly, the genes most highly induced by cold acclimation were involved in contraction and in signal transduction between nerve and muscle cells, while no significant changes were observed in genes and pathways related to insulin signaling or glucose metabolism.   Conclusions: Overall, our results indicate that cold acclimation and exercise training have overlapping effects on gene expression in human skeletal muscle, but strikingly these overlapping genes are designated to pathways related to cell remodeling rather than metabolic pathways.

overall_design: Microarray analysis was performed on a muscle biopsies taken from overweight male individuals with type 2 diabetes before and after a 10 day cold acclimation period.

first: Emmani

last: Nascimento

first: Roland

middle: W

last: Hangelbroek

middle: J

first: Joris

last: Hoeks

first: Wouter

last: Van Marken Lichtenbelt

first: Matthijs

last: Hesselink

first: Patrick

last: Schrauwen

first: Sander

last: Kersten


title: Comparative transcriptome analysis of human skeletal muscle in response to cold acclimation and exercise training in human volunteers.


title: Identification and characterization of a novel double-strand RNA mycovirus from Malassezia restricta KCTC 27540

city: Seoul

first: Yong-Joon

last: Cho

address: 1 Gwanak-ro, Gwanak-gu

country: South Korea

institute: Seoul National University

summary: To understand the overall effect of the virus on fungal host, we compared the transcriptomes between virus-infected and virus-cured M. restricta strain by using RNA-Seq.

overall_design: Comparison of the transcriptomes between virus-infected and virus-cured M. restricta KCTC 27540

first: Minji

last: Park

first: Won

last: Jung


title: Single-Cell RNA Sequencing Uncovers Heterogenous Transcriptional Signatures in Macrophages during Efferocytosis

status: Public on Aug 22 2020

city: Evanston

first: Behram

middle: Sarosh

last: Radmanesh

state: IL

address: 633 Clark St

institute: Northwestern University

summary: Efferocytosis triggers cellular reprogramming, including the induction of mRNA transcripts which encode anti-inflammatory cytokines that promote inflammation resolution. Our current understanding of this transcriptional response is largely informed from analysis of bulk phagocyte populations; however, this precludes the resolution of heterogeneity between individual macrophages and macrophage subsets. Moreover, phagocytes may contain so called “passenger” transcripts that originate from engulfed apoptotic bodies, thus obscuring the true transcriptional reprogramming of the phagocyte. To define the transcriptional diversity during efferocytosis, we utilized single-cell mRNA sequencing after co-cultivating macrophages with apoptotic cells. Importantly, transcriptomic analyses were performed after validating the disappearance of apoptotic cell-derived RNA sequences. Our findings reveal new heterogeneity of the efferocytic response at a single-cell resolution, particularly evident between F4/80+ MHCIILO and F4/80- MHCIIHI macrophage sub-populations. After exposure to apoptotic cells, the F4/80+ MHCIILO subset significantly induced pathways associated with tissue and cellular homeostasis, while the F4/80- MHCIIHI subset downregulated these putative signaling axes. Ablation of a canonical efferocytosis receptor, MerTK, blunted efferocytic signatures and led to the escalation of cell death-associated transcriptional signatures in F4/80+ MHCIILO macrophages. Taken together, our results newly elucidate the heterogenous transcriptional response of single-cell peritoneal macrophages after exposure to apoptotic cells.

overall_design: Examination of efferocytosis at three timepoints between two genotypes of primary macrophages.

first: Connor

last: Lantz

first: Esther

last: Liu

first: Edward

last: Thorp

first: Jennie

last: Lin


title: Comparative transcriptome analysis of human skeletal muscle in response to cold acclimation and exercise training in human volunteers. [A294]

overall_design: Microarray analysis was performed on a muscle biopsies taken from overweight middle-aged male subjects before and after a 12 week combined exercise training intervention.


title: Gene expression changes in human cord blood-derived hematopoietic stem and progenitor cells upon Centrifugation enhanced Nanostraw Transfection or Conventional Electroporation

status: Public on Aug 17 2020

city: Lund

first: Ludwig

last: Schmiderer

address: BMC A12

department: Laboratory Medicine / Molecular Medicine and Gene Therapy

country: Sweden

institute: Lund University

summary: CD34+ human cord blood-derived cells were subjected to GFP mRNA delivery or mock treatment using Centrifugation enhanced Nanostraw Transfection (CeNT) or conventional electroporation.

overall_design: Gene expression levels of three independent replicates of the following samples were measured by Affymetrix Human Gene 2.0 arrays: untreated, CeNT-mock, CeNT-GFP, Electroporation-mock, Electroporation-GFP. All conditions were compared to the untreated condition

first: Agatheeswaran

last: Subramaniam

first: Kristijonas

last: Žemaitis

first: Alexandra

last: Bäckström

first: David

last: Yudovich

first: Svetlana

last: Soboleva

first: Roman

last: Galeev

first: Christelle

middle: N

last: Prinz

first: Jonas

last: Larsson

first: Martin

last: Hjort


title: A Uromodulin mutation resulting in kidney fibrosis due to activation of the innate immune system through mononuclear phagocyte ATF4/integrated stress response signaling

city: Little Rock

first: Annjanette

last: Stone

email: annjanettemc@yahoo.com, astone@uams.edu

state: AR

address: 4300 West 7th Street

department: Research

institute: CAVHS

summary:
Background: Uromodulin (Tamm-Horsfall) protein is secreted by the thick ascending limb into urine and crosses the basolateral membrane into the kidney interstitium and blood. Uromodulin may engage the kidney’s innate immune system with pro and anti-inflammatory effects described. Mutations in the UMOD gene are a cause of autosomal dominant tubulointerstitial kidney disease (ADTKD) due to protein misfolding and endoplasmic reticulum stress. We identified a family with a UMOD mutation (C106F) that leads to glomerular and interstitial inflammation atypical for ADTKD.
Methods: To determine if the observed phenotype is due to mutant uromodulin induced changes in the innate or adaptive immune system, we developed a transgenic (tg) mouse with a homologous cysteine to phenylalanine mutation (C105F) in the UMOD gene using CRISPR-Cas9 gene editing.
Results: LLC-PK1 cells expressing wt and mutant protein had increased basolateral secretion of protein compared with cells expressing wt or mutant protein alone. EM examination of mutant medium showed protein aggregates in contrast to filaments in wt medium. Immunoprecipitation of plasma uromodulin from mutant mice demonstrated aggregate formation not seen in wt mice. Tg/+ mice displayed increased F4/80+ mononuclear phagocyte (MP) and podocyte inflammasome activity at baseline. Tg/+ mice developed glomerular and interstitial matrix deposition, myofibroblast proliferation and increased Creatinine with aging. Following ischemia-reperfusion injury, tg/+ mice had no increase in inflammasome activation over baseline in contrast to wt littermates and displayed improved tubular repair and renal function at 7 days. Differential gene expression analysis by RNA sequencing of kidney MP revealed activation of the ATF4 mediated stress response pathway and genes associated with the development of tissue fibrosis.
Conclusions: The C106F mutation results in glomerular and interstitial inflammatory kidney disease due to interaction of aggregated misfolded protein with mononuclear phagocytic cells resulting in ER stress response. The resulting alteration in the immune system leads to improved kidney repair following acute injury but fibrosis and chronic renal failure with aging.

overall_design: To determine if changes in macrophage gene expression is a mechanism linking phagocytosis of aggregated uromodulin to the pathogenesis of disease in transgenic mice, we performed RNA-sequencing of mRNA isolated from F4/80+ cells purified from 4 mutant mice and 6 wt littermate controls at 12-16 weeks using anti-F4/80 conjugated magnetic microbeads. Total N=10.

first: Matthew

last: Plotkin

first: Charles

last: Obrien

first: Joseph

last: Goellner

first: Joshua

last: Williams

first: Weleetka

last: Carter


title: A dichotomy of gene regulatory associations during the activated B-cell to plasmablast transition

last: Care


title: Aneuploidy-induced proteotoxic stress can be effectively tolerated without dosage compensation, genetic mutations or stress responses

city: Singapore

first: Katherine

last: Larrimore

address: 1 Research Link

department: Cell Biology

country: Singapore

institute: Temasek Life Sciences Laboratory

summary: The protein homeostasis (proteostasis) network maintains balanced protein synthesis, folding, transport and degradation within a cell. Because failure to maintain proteostasis is associated with aging and disease, a concerted effort has been placed on studying how the proteostasis network responds to various stresses. Typically, this is accomplished using ectopic overexpression of well-characterized, model misfolded protein substrates; however, how cells tolerate large-scale, diverse burden to the proteostasis network is not understood. Aneuploidy, the state of imbalanced chromosome content, adversely affects the proteostasis network by dysregulating the expression of hundreds of proteins simultaneously. Using aneuploid yeast cells as a model, we address what compensatory adjustments enable cells to tolerate large-scale, diverse challenges to the proteostasis network. Here we show adapted aneuploid Saccharomyces cerevisiae strains that exhibit robust growth and enhanced stress tolerance associated with enhancement of translation, folding, and quality control systems without genetic changes or activation of canonical stress response pathways.

overall_design: Total mRNA and ribosome-protected footprints were analyzed by RNA-Seq.total RNA and RPFs were extracted following manufacturer's instructions from the TruSeq Ribo Profile kit (formerly ARTseq) from Illumina  Libraries were prepared with the TruSeq Ribo Profile kit (formerly ARTseq) from Illumina, according to manufacturer's instructions. In one run conducted in 2015, WT (RLY2626), D2 (NVY1) and D1/2 (NVY2) were run together (no biological replicates). Another run was conducted in 2017 in which one biological replicate each of D1/2/8 (NVY3) and D13 (KLY196), and two biological replicates each of D1/2/8/11(KLY193), D1/8 (KLY194) and D13 (KLY195) were run.

middle: E

first: Natalia

last: Barattin-Voynova

last: Reid


title: STARR-seq enhancer activity in mouse embryonic stem cells cultured in 2i+LIF or serum+LIF [STARR-seq]

status: Public on Aug 16 2020

city: Utrecht

first: Wout

last: Megchelenbrink

address: Heidelberglaan 25

institute: Princess Maxima Centre for Pediatric Oncology

summary: Enhancers are distal regulators of gene expression that shape cell identity and regulate cell fate transitions. Mouse embryonic stem cells (mESCs) are a typical example of cells whose pluripotent identity is maintained by a complex enhancer landscape, that is drastically altered upon differentiation. Genome-wide chromatin accessibility and histone modification assays are commonly used as a proxy for enhancer location, strength and dynamics. Here, we applied STARR-seq, a genome-wide plasmid-based assay, to measure the enhancer potential of genomic loci in a plasmid context in “ground-state” (2i+LIF; 2iL-ESCs) and “metastable” (serum+LIF; SL-ESCs) embryonic stem cells.

overall_design: 8 samples were analyzed. 2 biological replicates were performed in either 2i+LIF or serum+LIF condition cultured mESCs. Each STARR-seq libarary has it's own input library as control.

first: Tianran

last: Peng

first: Yanan

last: Zhai

first: Menno

last: ter Huurne

first: Yaser

last: Atlasi

first: Hendrik

last: Marks

last: Stunnenberg


title: Genome-wide analysis of the FOXA1 transcriptional network identifies novel protein coding and lncRNA targets in colorectal cancer cells

city: Bethesda

first: Lorinc

last: Pongor

state: MD

address: 37 Convent Dr,

department: DTB

institute: NIH

summary: Differentiation status of tumors is correlated with metastatic potential and malignancy. FOXA1 (forkhead box A1) is a transcription factor known to regulate differentiation in certain tissues. Here, we investigate FOXA1 function in human colorectal cancer (CRC). We found that FOXA1 is robustly expressed in the normal human colon but significantly downregulated in colon adenocarcinoma (COAD). Applying FOXA1 ChIP-seq and RNA-seq upon FOXA1 knockdown in well-differentiated colon-like cells, and FOXA1 overexpression in poorly differentiated CRC cells, we identified novel protein-coding and lncRNA genes regulated by FOXA1. Among the numerous novel FOXA1 targets we identified, we focused on CEACAM5, a tumor marker and facilitator of cell adhesion. We show that FOXA1 binds to a distal enhancer downstream of CEACAM5 and strongly activates its expression. Consistent with these data, we show that FOXA1 inhibits anoikis in CRC cells. Collectively, our results uncover novel protein-coding and non-coding targets of FOXA1 and suggest a vital role of FOXA1 in enhancing CEACAM5 expression and anoikis resistance in CRC cells.

overall_design: RNA-seq data of colon adenocarcinoma cell lines. SW1222 and LS180 cell lines were compared before and after siFOXA1. In HCT116, cells were compared before and after FOXA1 oveerexpression

first: Sarah

last: Lazar

first: Wei

last: Tang

first: Ashish

last: Lal


title: YTHDF2/3 are required for somatic reprogramming through different RNA deadenylation pathways

city: Guangzhou

first: Jiadong

last: Liu

state: Guangdong province

address: 190 Kai Yuan Avenue, Science Park

department: South China Institute for Stem Cell Biology and Regenerative Medicine

institute: GIBH

summary: N6-methyladenosine (m6A), the most abundant reversible modification on eukaryote messenger RNA, is recognized by a series of readers, including the YT521-B homology domain family (YTHDF) proteins, which are coupled to perform physiological functions. Here, we report that YTHDF2 and YTHDF3, but not YTHDF1, are required for reprogramming of somatic cells into induced pluripotent stem cells (iPSCs). Mechanistically, we found that YTHDF3 recruits the PAN2-PAN3 deadenylase complex and conduces to reprogramming by promoting mRNA clearance of somatic genes, including Tead2 and Tgfb1, which parallels the activity of the YTHDF2-CCR4-NOT deadenylase complex. Ythdf2/3 deficiency further represses mesenchymal-to-epithelial transition (MET) and chromatin silencing at loci containing the TEAD motif, contributing to decreased reprogramming efficiency. Moreover, RNA interference of Tgfb1 or the Hippo signaling effectors Yap1, Taz, and Tead2 rescues Ythdf2/3-defective reprogramming. Overall, YTHDF2/3 couple RNA deadenylation and regulation with the clearance of somatic genes provides insights into iPSC reprogramming at the posttranscriptional level.

overall_design: ATAC-seq, ChIP-seq, RNA-seq and mRNA lifetime profiling of reprogramming cells. SRA Study accession: SRP233516, BioProject accession: PRJNA592060


title: STARR-seq identifies chromatin masked enhancers in pluripotent mouse embryonic stem cells [single_cell_RNA-Seq]

overall_design: Single cell RNA-seq (Celseq2) of one 2i+LIF WT, one  serum+LIF WT and two independent Zic3 knockout cell line were cultured  in 2i+LIF or serum+LIF (biological replicates). 6 plates were sequenced in total.


title: The genetic basis of diurnal preference in Drosophila melanogaster

city: Liverpool

first: Mirko

last: Pegoraro

state: Liverpool

address: Byrom Street

department: School of Natural Science and Psychology

institute: Liverpool John Moores University

summary: Most animals restrict their activity to specific part of the day, being either diurnal, nocturnal, or crepuscular. The genetic basis underlying this diurnal preference is largely unknown.   Under laboratory conditions, Drosophila melanogaster is crepuscular, showing a bi-modal activity profile. However, recent experiments in our lab indicated that high variability among individuals exist, particularly in strains that derive from different wild populations. By assembling together flies from various geographical strains, we have generated a highly diverse population whose progeny exhibited extreme diurnal preference, including diurnal and nocturnal flies. We have used this population as a starting point for an artificial selection experiment in which we selected males that show the most extreme diurnal preference and mated them to their sisters. The response to selection was strong, and already after 10 selection cycles we obtained highly diurnal (D) and nocturnal (N) strains. Another strain that was not selected and showed intermediate behaviour (crepuscular) served as a control (C). These strains provide us with a unique opportunity to understand the genetics of diurnal preference.

overall_design: 3 lines: Nocturnal (NDN), Diurnal (RLD) and control intermidiate (PJN). Sample collected at ZT0 and 12. Each sample has 2 replicates (A,B). Total single end libraries 12.

first: Laura

middle: M

last: Flavell

first: Pamela

last: Menegazzi

first: Perrine

last: Colombi

first: Pauline

last: Dao

first: Charlotte

last: Helfrich-Förster

first: Eran

last: Tauber


title: STARR-seq identifies chromatin masked enhancers in pluripotent mouse embryonic stem cells

first: Wout

last: Megchelenbrink

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

overall_design: Refer to individual Series


title: Gene expression changes associated with malignant transformation of oral potentially malignant disorders

status: Public on Aug 14 2020

city: Newcastle

first: John

last: Casement

address: Framlington Place

department: Bioinformatics Support Unit

institute: Newcastle University

summary: A large number of oral squamous cell carcinomas (OSCCs) are believed to be preceded by oral potentially malignant disorders (OPMD) that have an increased likelihood of malignant transformation compared to clinically normal mucosa. This study was performed to identify differentially expressed genes between OPMDs that underwent malignant transformation (MT) and those that did not, termed ‘non-transforming’ (NT) cases. Total RNA was extracted from formalin-fixed paraffin-embedded tissue biopsies of 20 OPMD cases with known clinical outcomes (10 MT vs. 10 NT). Samples were assessed for quantity, quality and integrity of RNA prior to sequencing. Analysis for differential gene expression between MT and NT was performed using statistical packages in R. Genes were considered to be significantly differentially expressed if the False Discovery Rate corrected p-value was < 0.05. RNA yield was variable but RNA purity was good (A260/A280 >1.90). Analysis of RNA-Sequencing outputs revealed 41 genes (34 protein-coding; 7 non-coding) that were significantly differentially expressed between MT and NT cases. The log2 fold change for the statistically significant differentially expressed genes ranged from -2.63 to 2.48, with 23 protein-coding genes being downregulated and 11 protein-coding genes being upregulated in MT cases compared to NT cases. Several candidate genes that may play a role in malignant transformation of OPMD have been identified. Experiments to validate these candidates are underway. It is anticipated that this work will contribute to better understanding of the aetiopathogenesis of OPMD and development of novel biomarkers.

overall_design: 20 OPMD cases with known clinical outcomes were investigated by RNA sequencing. The gene expression profile of 10 cases that underwent malignant transformation were compared with 10 cases that did not.

first: Hans

middle: P

last: Sathasivam

first: Ralf

last: Kist


title: STARR-seq identifies chromatin masked enhancers in pluripotent mouse embryonic stem cells [RNA-Seq]

overall_design: RNA-seq of Zic3 KO, P53 KO and WT mouse embryonic stem cells cultured in 2i+LIF or serum+LIF. 2 biological replicates per condition, 12 samples in total.

first: Tianran

last: Peng

first: Yanan

last: Zhai

first: Menno

last: ter Huurne

first: Yaser

last: Atlasi

first: Hendrik

last: Marks

last: Stunnenberg


title: Targeted RNA Knockdown by a Type III CRISPR-Cas Complex in Zebrafish

status: Public on Aug 13 2020

city: Warsaw

first: Cecilia

middle: Lanny

last: Winata

state: Mazovia

address: 4 Ks. Trojdena

country: Poland

institute: International Institute of Molecular and Cell Biology in Warsaw

summary: RNA interference (RNAi) is a powerful experimental tool for RNA knockdown, but not all organisms are amenable. Here, we provide a “proof of principle” demonstration that CRISPR endoribonuclease can be used for programmable mRNA transcript degradation in eukaryotes. Using zebrafish as the animal model and Csm complex as the CRISPR endoribonuclease, we targeted EGFP transcript expressed from a variety of promoters. A drastic decrease of fluorescence was achieved in germ cells of the vasa:EGFP line. Weaker effects were also seen in fish lines that express EGFP zygotically. Knockdown was statistically significant in cmcl2:EGFP and fli1:EGFP zebrafish lines at 1 day post fertilization (dpf), but reduced to background levels at 2 dpf. The nkx2.5:EGFP fish line was least susceptible to Csm mediated EGFP knockdown. We also tested Csm mediated knockdown on the endogenous tdgf1 (oep) transcript. At optimal Csm dose, we observed a penetrance of the characteristic one-eyed phenotype at greater than 50% penetrance, and hence with similar efficiency to morpholino-mediated knockdown. We conclude that Csm mediated knockdown is very efficient for maternal transcripts and can also be used for mixed maternal/early zygotic and early zygotic transcripts, in some cases reaching comparable efficiency to morpholino-based knockdown with no significant off-target effects in our model.

overall_design: Zebrafish embryos from the transgenic line Tg(ddx4:ddx4-EGFP) on the background of ABTL were injected with the Csm complex targeted against the EGFP transcript or mock solution (StCsm buffer without the proteins). The RNA was isolated from the pooled embryos at three time points: 128 cell stage, 5 hpf and 24 hpf. Three replicates were obtained from each experimental condition and time point. Pair-end sequencing was performed on the Illumina NextSeq platform.

first: Maciej

middle: T

last: Łapiński

first: Matthias

last: Bochtler


title: miRNA profile of single hormone receptor positive breast cancer patients

status: Public on Aug 15 2020

city: Gdańsk

first: Wojciech

last: Biernat

address: Smoluchowskiego 17

department: Department of Pathomorphology

institute: Medical Univeristy of Gdańsk

summary: In the current pilot study we aimed to identify differentially expressed miRNAs in two types of single hormone receptor-positive breast cancer [ER(+)/PgR(-) and ER(-)/PgR(+)] with further distinction into HER2-overexpressing/amplified and HER2-negative tumors in a well-established cohort collected at Medical University of Gdańsk.

overall_design: Owing to the complexity of breast cancer and the essential role of both HER2 and the hormone receptors (ER and PgR) in its biology, we were interested in evaluating the differences in miRNA profiles between the 4 ER/PgR/HER2 phenotypes of single hormone receptor positive primary breast tumors.

first: Michał

last: Kunc

first: Marta

last: Popęda

first: Anna

last: Szałkowska

first: Magdalena

last: Niemira

last: Bieńkowski

first: Rafał

last: Pęksa

first: Aleksandra

last: Łacko

first: Barbara

middle: S

last: Radecka

first: Marcin

last: Braun

first: Joanna

last: Pikiel

first: Maria

last: Litwiniuk

first: Katarzyna

last: Pogoda

first: Ewa

last: Iżycka-Świeszewska

first: Adam

last: Krętowski

last: Żaczek

first: Elżbieta

last: Senkus-Konefka


title: STARR-seq identifies chromatin masked enhancers in pluripotent mouse embryonic stem cells [ChIP-seq]

first: Wout

last: Megchelenbrink

overall_design: ZIC3 ChIP-seq of 2 biological replicates in mouse embryonic stem cells cultured in 2i+LIF or serum+LIF (4 samples total)

first: Tianran

last: Peng

first: Yanan

last: Zhai

first: Menno

last: ter Huurne

first: Yaser

last: Atlasi

first: Hendrik

last: Marks

last: Stunnenberg


title: The role of the NMD factor UPF3B in olfactory sensory neurons

city: La Jolla

first: Kun

last: Tan

address: 2880 Torrey Pines Scenic Dr. Room 4812

department: Department of Obstetrics, Gynecology, and Reproductive Sciences

institute: UCSD

summary: The UPF3B-dependent branch of the nonsense-mediated RNA decay (NMD) pathway is critical for human cognition. Here, we examined the role of UPF3B in the olfactory system. Single-cell RNA-sequencing (scRNA-seq) analysis demonstrated considerable heterogeneity of olfactory sensory neuron (OSN) cell populations in wild type (WT) mice, and revealed that UPF3B loss influences specific subsets of these cell populations. UPF3B also regulates the expression of a large cadre of anti-microbial genes in OSNs, and promotes the selection of specific olfactory receptor (Olfr) genes for expression in mature OSNs (mOSNs). RNA-seq and Ribotag analyses identified classes of mRNAs expressed and translated at different levels in WT and Upf3b-null mOSNs. Integrating multiple computational approaches, UPF3B-dependent NMD target transcripts that are candidates to mediate the functions of NMD in mOSNs were identified in vivo. Together, our data provides a valuable resource for the olfactory field and insights into the roles of NMD in vivo.

overall_design: Single cell sequencing (10X genomics) from olfactory epithelium from adult mice (with four biological replicates) were performed for this study. For bulk RNAseq, purified mOSNs from four different donor mice were used for library prep and sequencing (pair-end 100). For translome, purified riboTag-labeled RNAs from mOSNs from three different donor mice were used for library prep and sequencing (single-end 50).

first: Samantha

middle: H

last: Jones

first: Miles

middle: F

last: Wilkinson


title: Transcriptional responses of fungi (Rhizopus microsporus host [ATCC5 2813] and non-host [ATCC 11559] isolates) and bacteria (Burkholderia sp B13 ) to interaction with each other at a distance (pre-contact) and during physical contact.

city: Ithaca

first: Olga

last: Lastovetsky

state: NY

address: 334 Plant Science Building, 236 Tower Rd, Cornell University

institute: Cornell University

summary: The goal of this study was to identify reciprocal commnication between fungi and bacteria during a mutualistic and antagonistin interaction at two time points.

sample_organism:
Rhizopus microsporus ATCC 52813
Burkholderia sp. b13

overall_design: Methods: Host (ATCC 52813) and non-host (ATCC 11559) isolates of R. microsporus were grown alone or in the presence of Burkholderia sp. B13 endobacteria (wb) isolated from the host isolate. Fungal mycelia and bacterial cells were harvested before fungi have contacted bacteria (pre-contact) and  when the fungal colony has just begun to come into physical contact with the bacteria (physical contact). Total RNA was extracted, rRNA depleted with RiboZero, and sequenced on the Illumina Hi-Seq 100bp-paired end reads platform

middle: A

first: Teresa

last: Pawlowska


title: Ubiquitin ligase COP1 suppresses microglia-mediated inflammation

city: South San Francisco

first: Rohit

last: Reja

address: 1 DNA Way

institute: Genentech

summary: Dysregulated microglia are intimately involved in neurodegeneration including Alzheimer’s disease (AD) pathogenesis, but the mechanisms controlling pathogenic microglial gene expression remain poorly understood. The transcription factor CCAAT/enhancer binding protein beta (c/EBPß) regulates pro-inflammatory genes in microglia and is upregulated in AD. We show expression of c/EBPß in microglia is regulated post-translationally by the ubiquitin ligase COP1 (also called RFWD2). Ubiquitination of c/EBPß by COP1 targets it for proteasomal degradation. In the absence of COP1, c/EBPß accumulates rapidly and drives a potent pro-inflammatory and ApoE gene-expression program, evidenced by increased neurotoxicity in microglia-neuronal co-cultures. Antibody blocking studies reveal that neurotoxicity is almost entirely attributable to complement. Remarkably, loss of a single allele of Cebpb prevented the pro-inflammatory phenotype. COP1-deficient microglia markedly accelerated tau-mediated neurodegeneration in a mouse model where elevated ApoE plays a deleterious role. Collectively, these results identify c/EBPß as a potential therapeutic target for inflammation-driven neurodegeneration.


title: Loss Of Neural Stem Cell O-Glcnacylation Regulates An Age-Related Fate Switch In The Mature Hippocampus

city: Stanford

first: Julien

last: Couthouis

address: 279 Campus Dr

department: Genetics

institute: Stanford University

summary: To gain mechanistic insight into the role of O-GlcNAc in adult hippocampal neural stem cells (NSCs), we treated primary NSCs with a pharmacological inhibitor of Ogt or a lentivirus encoding an shRNA construct targeting Ogt. We determined that loss of O-GlcNAc in NSCs impairs proliferation and activates programs associated with glial differentiation.

overall_design: mRNA of NSCs was isolated by lysis with TRIzol Reagent (Thermo Fisher Scientific), separation with chloroform (0.2mL per mL TRIzol), and precipitated with isopropyl alcohol according to manufacturer’s instructions. RNA-Seq libraries were constructed using the Smart-Seq2 protocol from Trombetta et al. 2014. Final libraries were purified with Ampure beads and quantified using a qPCR Library Quantification Kit (Kapa Biosystems). Libraries were pooled for sequencing on an Illumina HiSeq 2500 (paired reads 2x100bp). Alignment of RNA-sequencing reads to the transcriptome was performed using STAR with ENCODE standard options, read counts were generated using rsem, and differential expression analysis was performed in R using DESeq2 package (Love et al., 2014). All bioinformatics analyses were performed on Sherlock, a Stanford HPC cluster.

first: Charles

last: White III

first: Saul

last: Villeda


title: Tumor-Infiltrating Regulatory T Cell Accumulation in the Tumor Microenvironment is Mediated by IL33/ST2 Signaling

status: Public on Aug 12 2020

city: Boston

first: JAE WON

last: CHO

state: MA

address: 60 Fenwood Road

department: Neurology

institute: Brigham and Women's Hospital

summary: Regulatory T cells (Tregs) are enriched in the tumor microenvironment (TME) and suppress anti-tumor immunity. However, the origin of tumor-infiltrating (TI) Tregs and the molecular mechanism underlying their TME accumulation are poorly understood. Although IL-33 was reported to regulate the expansion and function of Tregs, its direct role on TI Tregs and its contribution to tumor progression remain uncertain. Firstly, we demonstrated TI Tregs were primarily thymus-derived. More importantly, we found using comparative transcriptome analysis that proliferation-related genes were prominently upregulated in TI Tregs compared with TI CD4+Foxp3− conventional T cells or splenic Tregs of same tumor-bearing mice. One of these genes, ST2, an interleukin-33 (IL-33) receptor, was identified as a potential factor driving Treg accumulation in the TME. Indeed, IL-33-directed ST2 signaling induced the preferential proliferation of TI Tregs and enhanced tumor progression, while genetic deletion of ST2 in Tregs limited their TME accumulation and delayed tumor growth. These data demonstrate the IL-33/ST2 axis in Tregs as one of the critical pathways for the preferential accumulation of thymus-derived Tregs in the TME and suggest the axis as a potential therapeutic target for cancer immunotherapy.

overall_design: CD4 T cell from mouse lung cancer and CD4 T cell from mouse normal lung. From each sample, scRNA-seq and scVDJ-seq were performed

first: Jimin

last: Son

first: Jae-Won

last: Cho

first: Hyo-Jin

last: Park

first: Jihyun

last: Moon

first: Seyeon

first: Hoyoung

last: Lee

first: Jeewon

first: Gamin

last: Kim

first: Su

first: Sergio

last: Lira

first: Andrew

last: Mckenzie

first: Hye

middle: Y

first: Cheol

last: Choi

first: Yong

last: Lim

first: Seong

first: Su-Hyung

first: Eui-Cheol

last: Shin

first: Insuk

first: Sang-Jun

last: Ha


title: miR-24 induces human T cell exhaustion though mitochondria energy metabolic reprograming via suppressing MYC and FGF11

status: Public on Aug 11 2020

first: Jiang

last: Li

state: Guangdong

address: Dongfengdong Road NO.651, Yuexiu District

institute: State Key Laboratory of Oncology in South China, Sun Yat-sen University Cancer Center

summary: Exhausted T cells (TExh) is a barrier for cancer immunotherapy, evidences demonstrate that transcriptional regulation is essential for TEX induction; however the factors that manipulate exhausted phenotype are largely unexplored. Here we showed that the microRNA (miR)-24, previously established to be enriched in eoxosmes from nasopharyngeal carcinoma (NPC) cells and promote T cell dysfunction, induced T cells to express high level of TIM-3, PD-1 and CD39 but to display low IFNg and GrB secretion and diminished proliferation ability ( a TExh phenotype) through inhibiting energy metabolism in vitro. Forced miR-24 expression inhibited ATP production through mitochondrial oxidative phosphorylation, while blockade of endogenous miR-24 the mitochondrial formation became massive and tubular and the levels of mitochondrial constituent proteins including Mfn1, Mfn2, P-Drp1 and TOMM20 was increased in OKT-3 stimulated T cells. MiR-24-mediated TEX and mitochondria metabolism reprogramming were regulated by suppressing MYC and FGF11 expression. Moreover, MYC enhanced the FGF11 transcription to promote mitochondria ATP production. Importantly, clinical data showed an increased TEX phenotype in circuiting and tumor-infiltrating T cells from NPC patients. Thus, our findings support that inhibition of miR-24-mediated mitochondria metabolism reprogramming can be titrated to break TExh immune barrier in NPC immunotherapy.

overall_design: Examination of 2 different treatments in human T cells.

first: Yina

last: Liu


title: RRBS, RNA-seq of placenta in women with PCOS and RNA-seq of hippocampus and hypothalamus in femal rats born to PCOS rats.

city: Hangzhou

first: Ningning

last: Xie

address: 1 Xueshi Road

institute: Women’s Hospital, School of Medicine, Zhejiang University

summary: Polycystic ovarian syndrome (PCOS) is characterized by hyperandrogenism, irregular ovulation and polycystic ovaries. In the present study, we utilized integrated analysis of whole transcriptome and DNA methylome by reduced representation bisulfite sequencing (RRBS) and RNA sequencing (RNA-seq) of the fetal side of placenta in women with PCOS. A cluster of neural function related genes, such as LIN7A and MAP1B, were identified with integrative analysis of methylome and transcriptome. Inflammatory disease and response process were spotlighted with GO enrichments analysis of differentially transcriptional gene cluster in PCOS, and the cellular growth and the development and function of nervous and endocrine system were also closely related to PCOS. With respect to neural function, the enriched GO terms in differential expressed gene cluster of PCOS group involved brain development, the proliferation and differentiation of neural precursor cell, neuron and neuroblast. Consistently, the enriched GO terms in differential methylated gene cluster of PCOS group were associated with neuropeptide and synaptic transmission, as well as cognition and learning or memory. As the gene expression changes in hypothalamus and hippocampus probably result in long standing neurobehavioural consequences in rodents, we conducted a global analysis of transcriptomic data of F1 pubertal female rats born to PCOS rats.

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design: RRBS and RNA-seq were performed on fetal side of placentae in women with PCOS and corresponding controls (n=3 in each group). RNA-seq was performed on hippocampus and hypothalamus in femal rats born to PCOS rats (n=3 in each group).

first: Fangfang

last: Wang

first: Jue

last: Zhou

first: Minchen

last: Dai

first: Qing

last: Zhang

first: Paul

last: Hardiman

first: Fan

last: Qu


title: CD28 Co-stimulation Drives Tumor-Infiltrating T Cell Glycolysis to Promote Inflammation

status: Public on Aug 10 2020

city: Nashville

first: Jeff

last: Rathmell

state: TN

address: 1211 Medical Center Drive

department: PMI

institute: Vanderbilt University Medical Center

summary: Metabolic reprogramming dictates the fate and function of stimulated T cells, yet these pathways can be suppressed in T cells subjected to unique microenvironments, such as in a tumor. We previously showed that glycolytic and mitochondrial adaptations directly contribute to reduced effector functions of Renal Cell Carcinoma (RCC) CD8 tumor infiltrating lymphocytes (TIL). Here we define the role of these metabolic pathways in the activation and effector functions of RCC CD8 TIL. CD28 co-stimulation plays a key role to augment T cell activation and metabolism and is antagonized by inhibitory and checkpoint immunotherapy receptors CTLA4 and PD-1. While RCC CD8 TIL activated poorly when stimulated through the T cell receptor alone, addition of CD28 co-stimulation greatly enhanced activation, function, and proliferation. CD28 co-stimulation reprogrammed RCC CD8 TIL metabolism with increased glycolysis and mitochondrial oxidative metabolism, in part through upregulation of GLUT3. Mitochondria also became more fused, with higher membrane potential and overall mass. These phenotypes were dependent on glucose metabolism, as the glycolytic inhibitor 2-deoxyglucose both prevented changes to mitochondria and suppressed RCC CD8 TIL activation and function. These data show that CD28 co-stimulation can restore RCC CD8 TIL metabolism and function through rescue of T cell glycolysis that supports mitochondrial mass and activity.

overall_design: CD8+ T cells were isolated from patient peripheral blood and ccRCC tumor. Then the CD8+ T cells were treated with IL7, anti-CD3, anti-CD3/CD28 and anti-CD3/CD28/IL2 for 5 days and subject to single cell RNA sequencing. Bulk RNA sequencing done on sorted peripheral CD8 T cell and ccRCC TIL CD8 T cells without treatment.

first: Kathryn

last: Beckermann

first: Kirsten

last: Young

first: Xiang

last: Ye

first: Rachel

last: Hongo

first: Kimryn

first: Jeffrey

middle: C


title: RACK7 ChIP-seq in pediatric glioblastomas cell lines.

city: Shanghai

first: jiao

last: fangfang

address: DongAn Road 131

institute: fudan university

summary: We identified that RACK7 recognizes the histone H3.3G34R mutaion in vitro. In order to determine the interaction between RACK7 and H3.3G34R mutaion in cells, we used three pediatric glioblastoma (pGBM) cell lines. SJ-HGGx6c(R6) and SJ-HGGx42c(R42) have heterozygous G34R mutation, while SJ-HGGx39c(WT39) has wildtype H3F3A, which encodes H3.3, and we also corrected H3.3G34R in R6 and R42 cells to wildtype H3.3 by CRISPR/Cas9 mediated knock-in. Finally, we performed RACK7 ChIP-seq in these cell lines to explore the function of RACK7 and H3.3G34R mutation.

overall_design: RACK7 ChIP-seq was carried out in three pGBM cell lines and two H3.3 CRISPR/Cas9 knock-in cell lines.

last: Jiao

first: Chen

last: He

first: Wenqi

last: Xu

first: Gensheng

last: Yang

first: Suzanne

last: Baker

first: Yang

last: Shi

first: Rui

last: Guo


title: RNA-seq analysis of feeder cells for NK cells expansion

status: Public on Aug 07 2020

city: Newark

first: Dongfang

last: Liu

state: New Jersey

address: 205 S Orange Ave

department: Department of Pathology, Immunology and Laboratory Medicine

institute: Rutgers-The State University of New Jersey

summary: K562 and 221 cells

overall_design: mRNA profiles of K562, 221, K562-mIL21, and 221-mIL21

first: Yan


title: ALKBH5 regulates anti-PD-1 therapy response by modulating lactate and suppressive immune cell accumulation in tumor microenvironment

last: Li

state: California

address: 9500 Gilman Dr

department: Pediatrics

institute: University of California, San Diego

summary: We performed RNA-Seq and m6A RNA immunoprecipitation followed by high-throughput sequencing (MeRIP-Seq) to determine whether the altered gene expression and the regulatory functions in cancer immunotherapy could be a consequence of Alkbh5 or Fto-mediated m6A/m6Am demethylation. We used two m6A peak calling algorithms to find the common peaks between the two methods, and only kept the conserved peaks detected in all the biological replicates of each group to obtain the most stringent results. Our data suggest that the enzymatic activity and the altered m6A/m6Am peaks of Fto and Alkbh5 knockout had similarity but also had obvious distinct features, which may lead to the different mechanisms of the two protein to exert their functions in regulating immunotherapy.  In addition, we performed single cell RNA-seq on a tumor obtained from a patient who had stage IV melanoma progressive disease and responded well to PD-1 blockage therapy.  We observed substantial immune cell infiltration and very few residual melanoma cells, indicators of good immunotherapy responses. We then examined ALKBH5 expression and found that there were more melanoma cells expressing ALKBH5 than the surrounding normal control cells.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: NTC control, Alkbh5 KO, Fto KO mouse tumors after cancer immunotherapy were analyzed by RNA-Seq and MeRIP-Seq. Tumors from a melanoma patient who had stage IV melanoma progressive disease and responded well to PD-1 blockage therapy were analyzed by single cell RNA-seq.

first: Yuqi

last: Kang

first: Tariq

last: Rana


title: RNA sequencing of cell free saliva

status: Public on Aug 05 2020

city: Jerusalem

first: Maayan

last: Salton

address: Ein-Kerem Canpus

country: Israel

institute: Hebrew University

summary: We sequenced RNA from a pool of ten RNA samples originally extracted from cell free saliva of healthy women. The aim was to find the most abundant genes. We sequenced 1254 genes (with more than 49 reads). Out of them, 28 were splicing factors (2.23%).

overall_design: mRNA profiling of one pooled sample of cell free saliva RNA.

first: Mercedes

last: Bentata


title: Effects of successful aerobic and strength training on gene expression of lymphomonocytes of stable CAD patients

city: Jyväskylä

first: Maarit

last: Lehti

address: Rautpohjankatu 8

country: Finland

institute: LIKES Research Cetre for Physical Activity and Health

summary: Analysis of impact of successful combined aerobic and strength training intervention of CAD patients on lymphomonocyte gene expression. The purpose of this study was to identify the most up/down-regulated gene sets related to the training to increase our knowledge on the mechanisms mediating the exercise-induced effects.

overall_design: Total RNA obtained from lymphomonocytes isolated before and after 21 weeks successfull aerobic and strength taining intervention of stable coronary artery disease patients

first: Heikki

last: Kainulainen

first: Urho

last: Kujala


title: The S enantiomer of 2-hydroxyglutarate increases central memory CD8 populations and improves CAR-T therapy outcome

city: Cambridge

first: Iosifina

middle: Petrina

last: Foskolou

address: Downing Site

department: Physiology, Development and Neuroscience

institute: Cambridge University

summary:
Study aim: Investigate the effect of S-2HG (2-hydroxyglutarate) metabolite on the mRNA levels of human CD8 T cells
Methods: Naïve CD8 T cells were isolated from human healthy donors. The CD8 T cells were aCD3/CD28 beads (1:1 bead-to-cell ratio) and IL-2 (30 U/ml) for 4 days. The cells were treated with S-2HG (0.4 mM) or vehicle (H2O) from the day of isolation and for 12 days before collection. Library preparation and paired-end RNA sequencing using Illumina NextSeq 500, as well as downstream data analysis, were performed by Active Motif.

overall_design: human CD8 mRNA profiles of 3 independent healthy donors treated with S-2HG (0.4 mM) or vehicle (H2O) for 12 days

first: David

last: Bargiela

first: Randall

last: Johnson


title: Gene expression changes in cardiomyocytes from normal mice and diabetic, high fat diet-fed mice exposed to the DPP-4 inhibitor linagliptin

city: Toronto

first: Andrew

last: Advani

state: Ontario

address: 6-151 61 Queen Street East

department: Medicine

institute: St. Michael's Hospital

summary: Cardiomyocytes were isolated from the hearts of C57BL/6 mice or C57BL/6 mice that had been fed a high fat diet for 26 weeks, receiving an intraperitoneal injection of streptozotocin 12 weeks into the study. Cells were exposed to either linagliptin (9nmol/L) or vehicle (0.1% DMSO) for 24 hours before comparison of RNA levels by RNA-sequencing.

overall_design: RNA-sequencing using Ilumina Novaseq 6000 in primary cultured cardiomyocytes from C57BL/6 (Control) and diabetic, high fat diet-fed (DMHFD) mice treated with linagliptin 9nmol/L (Treated) or 0.1% DMSO (Vehicle) for 24 hours.

first: Sri

last: Batchu

first: Veera

middle: G

last: Yerra


title: Genomic and Long-Term Transcriptomic Imprints Related to the Daptomycin Mechanism of Action Occurring in Daptomycin- and Methicillin-Resistant Staphylococcus aureus Under Daptomycin Exposure

city: Catania

first: Viviana

last: Cafiso

state: Italy

address: Via Santa Sofia,97

department: Biometec

country: Italy

institute: University of Catania

summary: Daptomycin (DAP) is the last-resort treatment for heterogeneous Vancomycin-Intermediate-Staphylococcus aureus (hVISA) and Vancomycin-Intermediate-S.aureus (VISA), and DAP-resistance onset which is also linked to reduced vancomycin susceptibility, is an increasing public health problem. To have more insight into the mechanisms of daptomycin resistance, the comparative transcriptomes of two DAP-R (1C-3B) clinical isogenic isolates vs their DAP-S (1A-3A) counterparts were investigated by Illumina RNA-seq, the Rockhopper tool, computational filtering analyses and bioinformatic tools.

overall_design: Two Italian Daptomycin-susceptible/resistant (DAP-S/R) Staphylococcus aureus clinical strain pairs were investigated.

first: Stefano

last: Stracquadanio

first: Flavia

last: Lo Verde

first: Irene

last: De Guidi

first: Alessandra

last: Zega

first: Giuseppe

last: Pigola

first: Stefania

last: Stefani