title: Transcriptome profiles associated with selenium deficiency dependent oxidative stress identify potential diagnostic and therapeutic targets in liver cancer cells

status: Public on Dec 29 2020

city: Ankara

first: Damla

last: Gözen

state: Tr

address: Hayrabolu cad. Konutkent 2 sit. A5 blok No:6 Cayyolu

country: Turkey

institute: METU

summary:
Hepatocellular carcinoma (HCC) cells show increased resistance to various stress conditions such as oxidative stress. Conventional therapies have low efficacies due to resistance and off-target effects in HCC. Here we aimed to analyze oxidative stress-related gene expression profiles of two HCC cell lines.
To identify important genes that cause resistance to reactive oxygen species (ROS), a model of oxidative stress upon selenium (Se) deficiency was utilized in which an affymetrix microarray data was generated for two isogenic HCC cell lines - HepG2 and HepG2-2.2.15 (Head-to-Tail HBV genome integration) that are either sensitive or resistant to Se deficiency dependent oxidative stress, respectively.

overall_design: HepG2-2.2.15 and HepG2 cell lines were grown in either Se positive or Se negative mediums for 3 days in duplicates. They were harvested on the 1st, 2nd and 3rd days of the treatment, and RNA isolation was performed. For each day, gene expression analysis data by Human Genome U133 Plus 2.0 Array Affymetrix Array was acquired.

last: Gozen

first: Deniz

last: Cansen Kahraman

first: Kubra

last: Narci

first: Huma

last: Shehwana

first: Ozlen

last: Konu

first: Rengul

last: Cetin-Atalay


title: Response of CD14-derived macrophages to LPS

status: Public on Dec 15 2020

city: Moscow

first: Evgeny

last: Karpulevich

address: Alexandra Solzhenitsina

department: Information Systems

country: Russia

institute: ISP RAS

type: Expression profiling by high throughput sequencing

summary: We releaved the transcriptomic changes after LPS stimulation of CD14-derived macrophages from three donors.

overall_design: Blood samples from three femal donors were collected and CD14+ monocytes were isolated with magnetic beads. CD14-derived macrophages were obtained after 7 days of incubation of CD14+ monocytes with MCSF (5 ng/ml). After that LPS was added to experimental cells. Control cells were intact.

first: Polina

last: Vishnyakova

first: Anastasiya

last: Poltavets

first: Andrey

last: Elchaninov


title: Single-cell RNA sequencing of human nasal mucosa

status: Public on Jan 01 2021

city: Daejeon

first: Myung Jin

last: Yang

country: South Korea

institute: Institute for Basic Science

summary: Understanding on pathogenesis of COVID-19 is rapidly growing, but primary target cells of SARS-CoV-2 infection is still not known. Here, we performed single cell RNA sequencing on human nasal mucosa tissue to investigate the expression patterns of host cell entry factors of SARS-CoV-2.

overall_design: Single cells from human nasal mucosa were isolated and single-cell transcriptome libraries were generated

first: Gou

middle: Y

last: Koh

first: Ji

middle: H

last: Ahn

first: Myung

middle: J


title: Brain-infiltrating CD8+ T cell transcriptional activity after acute infection and during CNS autoimmunity [RNA-Seq]

status: Public on Dec 16 2020

city: Geneva

first: Sylvain

last: LEMEILLE

address: 1 rue Michel-Servet

department: Department of Pathology and Immunology

country: Switzerland

institute: University of Geneva Medical School

summary:
Gene expression of WT and Tox -/- P14 CD8+ T cells adoptively transfered in WT and MOG-GP recipient mice following intracranial infection with LCMV-GP.

CD8+ T cells play a pivotal role in eliminating pathogens or tumors, but they equally exert tissue damage in autoimmune or chronic inflammatory diseases. While the mechanisms regulating the differentiation of CD8+ T cells in chronic infections and cancer are just beginning to be unraveled, we still lack knowledge about how this unfolds in autoimmune diseases. Here, we investigated in a model of chronic central nervous system autoimmunity, the transcriptional and epigenetic landscape of autoreactive brain infiltrating CD8+ T cells and how these cells diverge from classical long-lived memory CD8+ T emerging after virus infection.

overall_design: Transcriptional activity of WT and Tox -/- P14 CD8+ T cells adoptively transfered in WT and MOG-GP recipient mice following intracranial infection with LCMV-GP.

first: Page

last: Nicolas

first: Merkler

last: Doron


title: Pioneer activity of an oncogenic fusion transcription factor at inaccessible chromatin

status: Public on Dec 12 2020

city: Colum bus

first: Stephanie

middle: D

last: LaHaye

state: OH

address: 575 Children's Crossroads

department: Institute for Genomic Medicine

country: USA

institute: Nationwide Children's Hospital

type: Genome binding/occupancy profiling by high throughput sequencing

summary: We developed a new method for ChIP-seq with per-cell normalization (pc-ChIP-seq) to  define PAX3-FOXO1 localization in rhabdomyosarcoma (RMS). We report novel pioneer function for the major driver oncogene in RMS, with nucleosome-motif targeting and kinetic displacement of nucleosomes in human cells.

overall_design: Genomic localization of the PAX3-FOXO1 fusion transcription factor in rhabdomyosarcoma cells and kinetic resolution of PAX3-FOXO1 chromatin occupancy and accessibility changes.

first: Benjamin

last: Stanton

first: Meng

last: Wang

last: Sunkel


title: Genomic and epigenomic 1α,25(OH)2D3 responses in human colonic organoids

status: Public on Dec 09 2020

city: Chicago

first: Sonia

last: Kupfer

state: IL

address: 900 East 57th Street, MB#9

department: Department of Medicine

institute: University of Chicago

summary:
Background & Aims: Active vitamin D, 1α,25(OH)2D3, is a nuclear hormone with roles in colonic homeostasis and carcinogenesis, yet mechanisms underlying these effects are incompletely understood. Human organoids are an ideal system to study genomic and epigenomic host-environment interactions. Here, we utilize human colonic organoids to measure 1α,25(OH)2D3 responses on genome-wide gene expression and chromatin accessibilityover time.
Methods: Human colonic organoids were cultured and treated in triplicate with either 100nM 1α,25(OH)2D3 or vehicle control for 4 and 18 hours (h) for chromatin accessibility, and 6 and 24hfor gene expression. DNA and RNA were extracted for ATAC- and RNA-sequencing, respectively. Differentially accessible peaks were analyzed using DiffBind and EdgeR; differentially expressed genes were analyzed using DESeq2. Motif enrichment was determined using HOMER.
Results: At 6h and 24h, 2870 and 2721 differentially expressed genes, respectively (FDR<5%) were identified with overall stronger responses with 1α,25(OH)2D3. Similarly, 1α,25(OH)2D3 treatment led to stronger chromatin accessibility especially at 4h. The vitamin D receptor (VDR) motif was strongly enriched among open chromatin peaks with 1α,25(OH)2D3 treatment accounting for 30.5% and 11% of target sequences at 4h and 18h, respectively (FDR<1%). A number of genes such as CYP24A1, FGF19, MYC, FOS and TGFBR2 showed significant transcriptional and chromatin accessibility responses to 1α,25(OH)2D3 treatment with open chromatin located distant from promoters for some gene regions.
Conclusions: Assessment of chromatin accessibility and transcriptional responses to 1α,25(OH)2D3 yielded new observations about vitamin D genome-wide effects in the colon facilitated by application of human colonic organoids. This framework can be applied to study host-environment interactions between individuals and populations in future.

overall_design: Human colonic organoids from single indvidual treated with 100nM active vitamin D for 4 and 18 hours for ATAC-seq and 6 and 24 hours for RNA-seq

first: Jinchao

last: Li

first: David

last: Witonsky

first: Emily

last: Sprague

first: Dereck

last: Alleyne

first: Maggie

last: Bielski

first: Kristi

last: Lawrence

middle: S


title: Aging stress and ceramide dependent RNA sequencing expression profiling depends on ceramide synthase 6

city: Charleston

first: Alhaji

middle: Harune

last: Janneh

state: SC

address: 86 Jonathan Lucas Street

department: Biochemistry and Molecular Biology

institute: Medical University of South Carolina

summary: The bioactive sphingolipid ceramide, generated by ceramide synthase, is an adaptive stress effector induced in response to various stress stimuli such as aging. In fact, ceramide synthase (CerS) was first discovered in yeast as a longevity assurance gene (LAG1), as the reduced ceramide generation consequent to LAG1 deletion increased longevity. There are six homologues of mammalian LAG1/ceramide synthases (CerS1-6) that generate ceramides with different fatty acid chain lengths with distinct functions and hydrophobicity. Ceramide can be metabolized by sphingosine kinase 1 or 2 for the generation of pro-survival sphingosine 1-phosphate (S1P), which enhances immune cell egress to the blood stream, activating immune function. Ceramide is known to induce mitophagy and cell death. Here we investigated the role of this sphingolipid pathway in immune function, specifically T cell functions, via high throughput expression profiling.

overall_design: Duplicate samples were analyzed from the WT and Ceramide Synthase deficient mouse. The splenic T cells were activated with anti-CD3, anti-CD28 and IL-2 for three days before the RNA was prepared from the activated T cells and RNA sequencing performed.

first: Silvia

middle: G

last: Vaena


title: Transcriptome-wide investigation of stop codon readthrough in Saccharomyces cerevisiae

status: Public on Dec 08 2020

city: Worcester

first: Kotchaphorn

last: Mangkalaphiban

state: MA

address: 368 Plantation St.

institute: University of Massachusetts Medical School

summary: Translation of mRNA into a polypeptide is terminated when the release factor eRF1 recognizes a UAA, UAG, or UGA stop codon in the ribosomal A site and stimulates nascent peptide release. However, stop codon readthrough can occur when a near-cognate tRNA outcompetes eRF1 in decoding the stop codon, resulting in the continuation of the elongation phase of protein synthesis. At the end of a conventional mRNA coding region readthrough allows translation into the mRNA 3’-UTR. Previous studies with reporter systems have shown that the efficiency of termination or readthrough is modulated by cis-acting elements other than stop codon identity, including two nucleotides 5’ of the stop codon, six nucleotides 3’ of the stop codon in the ribosomal mRNA channel, and stem-loop structures in the mRNA 3’-UTR. It is unknown whether these elements are important at a genome-wide level and whether other mRNA features proximal to the stop codon significantly affect termination and readthrough efficiencies in vivo. Accordingly, we carried out ribosome profiling analyses of yeast cells expressing wild-type or temperature-sensitive eRF1 and developed bioinformatics strategies to calculate readthrough efficiency, and to identify mRNA and peptide features which influence that efficiency. We found that the stop codon (nt +1 to +3), the nucleotide after it (nt +4), the codon in the P site (nt -3 to -1), and 3’-UTR length are the most influential features in the control of readthrough efficiency, while nts +5 to +9 and mRNA secondary structure in the 3’-UTR had milder effects. Additionally, we found low readthrough genes to have shorter 3’-UTRs compared to high readthrough genes in cells with thermally inactivated eRF1, while this trend was reversed in wild-type cells. Together, our results demonstrated the general roles of known regulatory elements in genome-wide regulation and identified several new mRNA or peptide features important for translation termination and readthrough.

overall_design: Translation profiles of the WT and sup45-2 strains were generated by ribosome profiling in duplicates using Illumina NextSeq 500. Readthrough efficiency of each gene was defined as the rato of ribosome density of the extension (region between the canonical stop and the first in-frame downstream stop in the 3'-UTR) to ribosome density of the coding region. For each sample, random forest algorithm was used to identify significant predictors of readthrough efficiency..

first: Allan

last: Jacobson


title: bulk RNA-seq analysis comparing HEK293 and HBEC 5i cell lines

city: New York

first: Edmund

last: Au

address: 630 West 168th Street, room P&S14-401

institute: Columbia University Irving Medical Center

summary: We report expression data analysis comparing HEK 293 and HBEC 5i immortalized cell lines grown for 3 days in serum-free Neurobasal medium

overall_design: examination of differential expression between 2 immortalized human cell line grown in serum-free conditions identical to those used to obtain serum-free conditioned medium

first: Matthieu

last: Genestine


title: Immunologic characterization of COVID-19 patients with hematologic cancer: biologic and clinical significance

status: Public on Dec 31 2020

first: Juan-Jose

last: Garces

state: New York

address: 321 E 61st St

institute: Memorial Sloan Kettering Cancer Center

summary: Immunosuppression caused by cancer itself and cytotoxic treatment may pose a challenge to coronavirus disease 2019 (COVID-19) patients with hematological malignancies. Here, we use multidimensional flow cytometry (MFC) to analyze immune profiles in peripheral blood samples of 515 COVID-19 patients at presentation. In 14 cases, deep immunophenotyping of B- and T-cells was performed and six myeloid- and dendritic-cell subsets were FACSorted for transcriptome analysis using RNAseq. Of the 515 patients, 15 and 10 had solid and hematological tumors, respectively. Those with hematological cancer showed significantly higher rates of intensive care (50%) and death (30%) from COVID-19 vs cases with solid cancer and no tumor. Patients with hematological malignancies displayed altered immune profiles with significantly decreased absolute numbers of several subsets of myeloid and lymphoid cells. Myeloid- and dendritic-cell types from hematological cases showed differentially expression of genes coding transcription factors, toll-like receptors and proinflammatory interleukin receptors implicated in response to coronaviruses. The relative distribution of the B-cell compartment was notoriously altered in COVID-19 patients with hematological cancer, and progressively lower numbers of B- and T-cell subsets were observed in deceased cases. Altogether, our results suggest an association between impaired immune responses and poorer outcomes in COVID-19 patients with hematological malignancies.

overall_design: Six cell immune populations sorted from 13 patients with COVID

first: Catarina

last: Maia

first: Esperanza

last: Martin

first: Juan-José

last: Garcés

first: Ascension

last: Lopez-Diaz de Cerio

first: Susana

last: Inogés

first: Manuel

last: Landecho

first: Belén

last: Gil-Alzugaray

first: Cristina

last: Perez

first: Cirino

last: Botta

first: Aintzane

last: Zabaleta

first: Félix

last: Alegre

first: César

last: Rincón

first: Laura

last: Blanco

first: Sarai

last: Sarvide

first: Diego

last: Alignani

last: Moreno

first: Mónica

last: Olid

first: Andrés

first: Felipe

last: Prosper

first: Bruno

last: Paiva

first: José-Ramón

last: Yuste


title: Knockout of immunotherapy prognostic marker genes eliminates the effect of the anti-PD-1 treatment

status: Public on Dec 01 2020

city: Beijing

first: Liqing

last: Cheng

state: Beijing

address: Tsinghua University,Hai Dian, Beijing

department: school of medicine

country: China

institute: Tsinghua university

summary: We present a pan-cancer integrated analysis of transcriptome, exome and clinical records of patients. Herein, we identified candidate genes as potential biomarkers to predict clinical outcome of immunotherapy. We further explored the function of two IPM genes in vivo. Knockout of MALT1, which is critical for the T cell receptor signaling, can eliminate the anti-tumor effect of anti-PD-1 treatment completely by impairing the activation of CD8+ T cells. Notably, knockout of CLEC4D, a C-type lectin receptor that expressed on myeloid cells, also reduced the effect of anti-PD-1 treatment potentially through maintaining the immunosuppressive effects of myeloid cells.

overall_design: Macrophages and MDSCs cells sorted from wild type (WT) or Clec4d KO mice with anti-PD-1  or IgG treatment were used for RNA extraction and RNA-sequencing

first: Naixue

last: Yang

first: Fansen

last: Ji


title: Genome-wide occupancy of conditionally-expressed wild-type and Lys120-to-Arg (K120R) p53 in H1299 cells.

status: Public on Dec 02 2020

city: CHICAGO

first: Manoj

last: Kandpal

state: Illinois

address: 1256 W NORTHSHORE AVE, 3R

institute: Northwestern University

summary: The tumor suppressor p53 integrates stress response pathways, triggering the appropriate cell fate by selectively engaging one of a diverse set of transcriptomes (e.g., cell cycle arrest, apoptosis). Foundational to this process is the binding of tetrameric p53 to response elements (REs) in the genome, of the consensus sequence (RRRCWWGYYYN0-13RRRCWWGYYY). This unusually long and degenerate RE provides a potential mechanism for contextually activating distinct sets of downstream targets. However, the RE code underlying this selectivity remains undeciphered. p53 REs exhibit the highest sequence variability at the triplets RRR/YYY, which flank the core (CWWG) sequence, and p53 tetramers interact with REs at those bases via Arg248 from the DNA minor groove and Lys120 from the DNA major groove. This set of observations triggered our hypothesis that the interplay of Lys120 and Arg248 residues with variable sequences of REs might be a potential source of p53 selectivity. To gain insight into the role of Lys120 integrity in the recognition of specific RE sequences, we performed a ChIP-seq experiment and analyzed genome-wide occupancy of conditionally-expressed wild-type (WT) and K120R forms of p53 in isogenic human cell lines that are null for endogenous p53. Using an optimized data analysis workflow, we mapped about 35 million unique sequence reads per sample to the human genome, and the number of significantly enriched peaks in each replicate was detected by setting an enrichment cutoff of 0.3 counts per million reads. This study presents a foundation for analysis of differences in sequences between K120R-sensitive and K120R-insensitive p53 binding sites.

overall_design: ChiP-seq profiles of WT and K120R-p53 forms in untreated [Tet(-)] and Tetracycline treated [(Tet(+)] 1299 cells.

first: Marina

last: Farkas

first: Yingtao

last: Bi

first: Ramana

middle: V

last: Davuluri

first: Adam

last: Ertel


title: Next Generation Sequencing to understand the pathogenic role of synovial fibroblasts in experimental models of arthritis

status: Public on Nov 30 2020

city: Glasgow

first: Miguel

last: Pineda

address: University Avenue

country: United Kingdom

institute: University of Glasgow

summary:
The goal of this study is to define the pathophysiology of synovial fibroblasts in chroninc joint inflammation using NGS-derived synovial  transcriptome profiling (RNA-seq)
Methods: Synovial fibroblasts were isolated by FACs sorting [CD45-CD31-Popdoplanin+] from the synovium of healthy mice and mice undergoing experimental Collagen-Induced Arthritis (12 weeks old)(CIA).
Methods:  Whole Transcriptome Profiling  of healthy and CIA synovial fibroblasts  were generated by deep sequencing, in triplicate, using Illumina NextSeq™ 500 platform.  Libraries were prepared using polyA selection (TruSeq stranded mRNA kit).
Methods: The sequence reads that passed quality filters were  aligned to mouse reference genome (GRCM38) using Hisat2 version 2.1.0.  we mapped about 30 million sequence reads per sample (75 bp, paired-end)
Methods: Featurecounts version 1.4.6 was used to quantify reads counts. Data quality control, non-expressed gene filtering, median ratio normalization (MRN) implemented in DESeq2  package and identification of differentially expressed (DE) genes were done  using the R bioconductor project DEbrowser.
Results: Results: We detected several differentially expressed (DE) genes in CIA synovial fibroblasts compared to naïve cells. These included 298 up-regulated genes and 88 down-regulated (>2 fold, adjp <0.05). DE genes reflected the reflecting the hyperplasia and activation observed during chronic joint disease. Pathways associated to DE genes were cell cycle and cell division and inflammatory response.
Conclusions: our study shows pathophysiological changes associated to inflammatory synovial fibroblasts in a model of rheumatoid arthritis. Cells were directly isolated from fresh tissue, providing a valuable tool to study stromal inflammation.

overall_design: Synovial fibroblast mRNA profiles of naïve healthy DBA1J and CIA mice.


title: The regulatory genome of the malaria vector Anopheles gambiae: integrating chromatin accessibility and gene expression

status: Public on Dec 22 2020

city: Granada

first: Jose

middle: Luis

last: Ruiz Rodriguez

state: Granada

address: Avda. del Conocimiento 17. P. T. Ciencias de la Salud

country: Spain

institute: IPBLN-CSIC

summary: Anopheles gambiae mosquitoes are primary human malaria vectors, but we know very little about their mechanisms of transcriptional regulation. We profiled chromatin accessibility by ATAC-seq in laboratory-reared An. gambiae mosquitoes experimentally infected with the human malaria parasite Plasmodium falciparum. By integrating ATAC-seq, RNA-seq and ChIP-seq data we showed a positive correlation between accessibility at promoters and introns, gene expression and active histone marks. By comparing expression and chromatin structure patterns in different tissues, we were able to infer cis-regulatory elements controlling tissue specific gene expression and to predict the in vivo binding sites of relevant transcription factors. The ATAC-seq assay also allowed the precise mapping of active regulatory regions, including novel transcription start sites and enhancers that annotate to mosquito immune-response genes. This study is important not only for advancing our understanding of mechanisms of transcriptional regulation in the mosquito vector of human malaria, but also the information we produced is of great potential for developing new mosquito-control and anti-malaria strategies.

overall_design: Genome-wide profiling of chromatin accessibility (ATAC-seq) in mosquito tissues infected by P. falciparum coupled with transcriptome profiling (RNA-seq).

first: José

middle: L

last: Ruiz

first: Lisa

middle: C

last: Ranford-Cartwright

first: Elena

last: Gómez-Díaz


title: Transcriptional analysis of THs-responding genes during human hepatocyte development

city: beijing

first: liang

middle: sheng

last: xian

address: shuangqing road

institute: chinese academy of science

summary: Liver is one of the major targets of thyroid hormones (THs), as well as many industrial chemicals reported as THs-analogues. A number of THs-analogues could permeate the placenta barrier, risking the embryonic development, which may be more susceptible and cause sustained health risk. Therefore, to study the toxicology of industrial chemicals during the hepatocytes developmental stage is necessary, and may be more efficiency. In this study, we differentiated human embryonic stem cells (hESCs) into functional hepatocyte-like cells in presence of two thyroid hormones (THs, T3 and T4), with the purposes trying to dissect genes regulated by THs during human hepatocyte development and identifying biomarkers for THs-analogues. THs exposure finally induced dysfunctional hepatocytes with 19 days differentiation, with impaired glycogen storage ability and lipid droplets accumulation. Global gene expression analysis by RNA-seq identified a number of genes responding to THs were responsible for hepatic differentiation and function, enriched in many metabolic related pathways.

overall_design: Total of 12 samples with 6 groups, including control on day 7 and day 19, T3 treatment on day 7 and day 19, T4 treatment on day7 and day 19; each group contains two replicates.

first: Shengxian

last: Liang

first: Francesco

last: Faiola


title: A role for the Mre11–Rad50–Xrs2 complex in gene expression and chromosome organization

city: Montpellier

first: romain

last: forey

address: 142 avenue de la Cardonille

country: France

institute: IGH

summary: Mre11–Rad50–Xrs2 (MRX) is a highly conserved complex with key roles in various aspects of DNA repair. Here, we report a new function for MRX in limiting transcription in budding yeast. We show that MRX interacts physically and colocalizes on chromatin with the transcriptional co-regulator Mediator. It restricts transcription of coding and noncoding DNA independently by a mechanism that does not require the nuclease activity of Mre11. MRX is required to tether the active GAL locus to the nuclear pore complex (NPC) and also promotes large-scale gene–NPC interactions. Moreover, MRX-mediated chromatin anchoring to the NPC contributes to chromosome folding and helps to control gene expression. Together, these findings indicate that MRX has a role in transcription and chromosome organization that is distinct from its known function in DNA repair.

overall_design: NGS-based analysis of Rad50, Mlp1 (ChIP-seq). Chromosome folding with MicroC-XL. Analysis of gene expression by strand specific, total RNA-seq of G1 synchronised S. cerevisiae cells, wild type and mutants for the MRX complex and degradation pathways.

first: Romain

last: Forey

first: Antoine

last: Barthe

first: Mireille

last: Tittel-Elmer

first: Maxime

last: Wery

first: Marie-Bénédicte

last: Barrault

first: Cécile

last: Ducrot

first: Andrew

last: Seeber

first: Nils

last: Krietenstein

first: Ugo

last: Szachnowski

first: Magdalena

last: Skryzpczak

first: Krzysztof

last: Ginalski

first: Maga

last: Rowicka

first: Jennifer

last: Cobb

first: Oliver

last: Rando

first: Julie

last: Soutourina

first: Karine

last: Dubrana

first: Susan

middle: M

last: Gasser

first: Antonin

last: Morillon

first: Philippe

last: Pasero

first: Armelle

last: Lengronne

first: Jérôme

last: Poli


title: The role of topical containing camellia sinensis in the cosmetic management of skin scars in healthy human volunteers: part 2

city: Manchester

first: Ardeshir

last: Bayat

address: Oxford Road

institute: University of Manchester

summary: Epigallocatechin-3-gallate (EGCG), a polyphenol, influences cutaneous wound healing due to its anti-angiogenic, anti-inflammatory and anti-oxidant properties. We previously showed EGCG is effective in improving skin scarring when applied immediately post-injury. Here, this double-blinded randomized placebo-controlled trial compared the effects of pre-emptive priming versus immediate and delayed topical EGCG compared with placebo.  This was evaluated in 40 healthy human volunteers, over 8 weeks, in scars created in their upper inner arms using skin punch biopsies. Participants were split into 4 groups; each undergoing different modes of application versus placebo: Group 1=priming (7D) pre-injury, Group 2=priming (3D) pre-injury, Group 3=immediate (0D) day-of-injury, Group 4=delayed application (14D) post-injury. RNA sequencing was used here based on 72 samples: Group 1: day 0 (n=3 placebo, n=3 EGCG), week 4 (n=3 placebo, n=3 EGCG), week 8 (n=3 placebo, n=3 EGCG); Group 2: day 0 (n=3 placebo, n=3 EGCG), week 4 (n=3 placebo, n=3 EGCG), week 8 (n=3 placebo, n=3 EGCG), Group 3:  day 0 (n=6), week 4 (n=3 placebo, n=3 EGCG), week 8 (n=3 placebo, n=3 EGCG), Group 4: day 0 (n=6), week 4 (n=3 placebo, n=3 EGCG), week 8 (n=3 placebo, n=3 EGCG). The common differentially expressed genes identified between treated (EGCG) and placebo samples were grouped according to time point and group. Further exploration of these genes using Gene Ontology and Ingenuity Pathway Analysis revealed gene signatures of relevant biological processes and pathways. Here, RNA sequencing revealed a number of angiogenic, inflammatory genes that were significantly downregulated with EGCG particularly in pre-emptive priming Group 1 at week 4 including; hemoglobin subunit beta (HBB), hemoglobin subunit alpha-1 (HBA1) and hemoglobin subunit alpha-2 (HBA2) at week-4.

overall_design: mRNA profiles of human skin scars over 8 weeks (treatment EGCG versus placebo) were generated by sequencing, in triplicate, using Illumina PE150 Platform.

first: Sara

last: Ud-Din


title: Spurious regulatory connections dictate the expression-fitness landscape of translation factors

status: Public on Nov 28 2020

city: Montreal

first: Jean-Benoit

last: Lalanne

state: QC

address: 5155 Chem. de la Rampe

department: Biochemistry and Molecular Medicine

country: Canada

institute: University of Montreal

summary: During steady-state cell growth, individual enzymatic fluxes can be directly inferred from growth rate by mass conservation, but the inverse problem remains unsolved. Perturbing the flux and expression of a single enzyme could have pleiotropic effects that may or may not dominate the impact on cell fitness. Here, we quantitatively dissect the molecular and global responses to varied expression of translation termination factors (peptide release factors, RFs) in the bacterium Bacillus subtilis. While endogenous RF expression maximizes proliferation, deviations in expression lead to unexpected distal regulatory responses that dictate fitness reduction. Molecularly, RF depletion causes expression imbalance at specific operons, which activates master regulators and detrimentally overrides the transcriptome. Through these spurious connections, RF abundances are thus entrenched by focal points within the regulatory network, in one case located at a single stop codon. Such regulatory entrenchment suggests that predictive bottom-up models of expression-fitness landscapes will require near-exhaustive characterization

overall_design: 26 competition experiment amplicon libraries from strain pools, 12 matched Rend-seq and ribosome profiling datasets, one individual Rend-seq dataset, 56 RNA-seq datasets. All data from Bacillus subtilis samples.

first: Darren

last: Parker

first: Gene-Wei

last: Li


title: Genome-scale identification of SARS-CoV-2 and pan-coronavirus host factor networks

status: Public on Nov 24 2020

first: John

last: Poirier

address: 550 1st Ave

institute: NYU Langone Health

summary: The COVID-19 pandemic has claimed the lives of more than one million people worldwide. The causative agent, SARS-CoV-2, is a member of the Coronaviridae family, which are viruses that cause respiratory infections of varying severity. The cellular host factors and pathways co-opted by SARS- CoV-2 and other coronaviruses in the execution of their life cycles remain ill-defined. To develop an extensive compendium of host factors required for infection by SARS-CoV-2 and three seasonal coronaviruses (HCoV-OC43, HCoV-NL63, and HCoV-229E), we performed parallel genome-scale CRISPR knockout screens. These screens uncovered multiple host factors and pathways with pan- coronavirus and virus-specific functional roles, including major dependency on glycosaminoglycan biosynthesis, SREBP signaling, BMP signaling, and glycosylphosphatidylinositol biosynthesis, as well as a requirement for several poorly characterized proteins. We identified an absolute requirement for the VTT-domain containing protein TMEM41B for infection by SARS-CoV-2 and three seasonal coronaviruses. This human Coronaviridae host factor compendium represents a rich resource to develop new therapeutic strategies for acute COVID-19 and potential future coronavirus spillover events.

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct

overall_design: Genome-wide CRISPR screens for SARS-CoV-2, HCoV-OC43, HCoV-NL63, and HCoV-229E coronavirus host factors.

first: William

last: Schneider

first: Joseph

last: Luna

first: Hans-Heinrich

last: Hoffmann

first: Francisco

last: Sánchez-Rivera

first: Andrew

middle: A

last: Leal

first: Alison

middle: W

last: Ashbrook

first: Jérémie

last: Le Pen

first: Inna

last: Ricardo-Lax

first: Eleftherios

last: Michailidis

first: Avery

last: Peace

first: Ansgar

middle: F

last: Stenzel

first: Scott

last: Lowe

first: Margaret

middle: R

last: MacDonald

first: Charles

last: Rice

middle: T


title: Gene and miRNA expression in radioiodine refractory and avid papillary thyroid carcinomas

city: Milan

first: Matteo

last: Dugo

address: via Olgettina 60

department: Department of Medical Oncology

country: Italy

institute: IRCCS Ospedale San Raffaele

type:
Expression profiling by array
Non-coding RNA profiling by array

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

overall_design: Refer to individual Series


title: Genetic determinants of EGFR-Driven Lung Cancer Growth and Therapeutic Response In Vivo

status: Public on Feb 01 2021

city: Stanford

first: Chuan

last: Li

state: CA

address: 327 Campus Drive

department: Biology

institute: Stanford University

summary: Cancer genome sequencing has uncovered substantial complexity in the mutational landscape of tumors. Given this complexity, experimental approaches are necessary to establish the impact of combinations of genetic alterations on tumor biology and to uncover genotype-dependent effects on drug sensitivity. In lung adenocarcinoma, EGFR mutations co-occur with many putative tumor suppressor gene alterations, however the extent to which these alterations contribute to tumor growth and their response to therapy in vivo has not been explored experimentally. By integrating a novel mouse model of oncogenic EGFR-driven Trp53-deficient lung adenocarcinoma with multiplexed CRISPR–Cas9-mediated genome editing and tumor barcode sequencing, we quantified the effects of inactivation of ten putative tumor suppressor genes. Inactivation of Apc, Rb1, or Rbm10 most strongly promoted tumor growth. Unexpectedly, inactivation of Lkb1 or Setd2 – which were the strongest drivers of tumor growth in an oncogenic Kras-driven model – reduced EGFR-driven tumors growth. These results were consistent with the relative frequency of these tumor suppressor gene alterations in human EGFR and KRAS-driven lung adenocarcinomas. Furthermore, Keap1 inactivation reduced the sensitivity of tumors to osimertinib in the EGFRL858R;p53flox/flox model. Importantly, in human EGFR/TP53 mutant lung adenocarcinomas, mutations in the KEAP1 pathway correlated with decreased time on tyrosine kinase inhibitor treatment. Our study highlights how genetic alterations can have dramatically different biological consequences depending on the oncogenic context and that the fitness landscape can shift upon drug treatment.

overall_design: We initiated tumors with a lentiviral-Cre vector in EGFR;p53 mice, EGFR;p53;Cas9 mice and KRAS;p53 mice at different time points with or without osimertinib treatments.

first: Giorgia

last: Foggetti

first: Hongchen

last: Cai

first: Jessica

last: Hellyer

first: Wen-Yang

last: Lin

first: Deborah

last: Ayeni

first: Katherine

last: Hastings

first: Jungmin

last: Choi

first: Anna

last: Wurtz

first: Laura

last: Andrejka

first: Dylan

last: Maghini

first: Nicholas

last: Rashleigh

first: Stellar

last: Levy

first: Robert

last: Homer

last: Gettinger

first: Maximilian

last: Diehn

first: Heather

last: Wakelee

first: Dmitri

last: Petrov

first: Monte

last: Winslow

first: Katerina

last: Politi


title: Age dependent effects of hypoxia-ischemia in neonatal mice brain

status: Public on Nov 13 2020

city: Rouen cedex

first: Celine

last: Derambure

address: 22 Bvd Gambetta

institute: INSERM U905

summary:
Goal was to determine differences in response to hypoxa-ischemia in the mouse brain at development stages representative of human preterm or term neonates, and to detect putative interference with spontaneous development from 2 to 15 days post-natal.
Hypoxia-ischemia was realized in 5-day old and 10 day-old mice by unilateral permanent carotid ligature followed by 40 min oxygen restriction (8%). Ipsilateral hemisphere were collected at 3h, 6h 12h or 24h after hypoxia and pools of sex-equilibrated 4-6 brains were used for total RNA extraction at each time point. Co-hybridization of ishemic mRNA (labelled Cy5) and control tissues (labelled Cy3) were performed on Agilent (design 026655) micro-arrays in three independent experiments. In order to evaluate hypoxia ischemia interference during development, we evaluated spontaneous expression at P5 and P10 (Cy3) and extented the observations in a separate set of tissues extracted at P2 and P15.

overall_design: Two color experiments : ischemic hemisphere (Cy5)/control tissue from untreated animals (Cy3). Moreover, P2 and P15 extracts each hybridized in two colors dye-swap.

first: Nicolas

last: Dupré

first: Céline

first: Bérénice

last: Le Dieu-Lugon

first: Philippe

last: Leroux


title: Reproductive history determines ErbB2 locus amplification, WNT signalling and tumor phenotype in a murine breast cancer model

city: Cardiff

first: Matthew

middle: John

last: Smalley

state: Cardiff

address: Hadyn Ellis Building, Maindy Road

department: School of Biosciences

institute: Cardiff University

type: Genome variation profiling by high throughput sequencing


title: Drug-drug interaction between metformin and sorafenib alters antitumor effect in hepatocellular carcinoma cells

status: Public on Dec 03 2020

city: Paris

first: Christèle

last: Desbois-Mouthon

address: 15 rue de l'école de médecine

institute: INSERM

summary:
Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and is one of the leading causes of cancer-related deaths worldwide. The multi‐target inhibitor sorafenib is a first-line treatment for patients with advanced unresectable HCC. Recent clinical studies have evidenced that patients treated with sorafenib together with the anti-diabetic drug metformin have a survival disadvantage compared to patients receiving sorafenib only. Here, we examined whether a clinically relevant dose of metformin (50 mg/kg/d) could influence the antitumoral effects of sorafenib (15 mg/kg/d) in a subcutaneous xenograft model of human HCC growth using two different sequences of administration, i.e concomitant versus sequential dosing regimens. We observed that the administration of metformin six hours prior to sorafenib was significantly less effective in inhibiting tumor growth than concomitant administration of the two drugs. In vitro experiments confirmed that pretreatment of different human HCC cell lines with metformin reduced the effects of sorafenib on cell viability, proliferation and signaling. Transcriptomic analysis confirmed significant differences between xenografted tumors obtained under the concomitant and the sequential dosing regimens. Taken together, these observations call into question the benefit of parallel use of metformin and sorafenib in patients with advanced HCC and diabetes, as the interaction between the two drugs could ultimately compromise patient survival.
To better characterize the molecular signatures driving the differential responses to concomitant and sequential biotherapies, we conducted a transcriptomic analysis on RNA extracted from tumor xenografts mice xenografed and treated with vehicle (control, n=3), metformin (50 mg/kg/day) combined to sorafenib (15 mg/kg/day) (concomitant schedule, n=3) or metformin (50 mg/kg/day) followed 6 hours later by sorafenib (15 mg/kg/day) (sequential schedule, n=3).

overall_design: Total RNA was extracted from tumors collected from mice xenografed subcutaneously in the right flank with PLC/PRF5 cells and treated with vehicle (control, n=3), metformin (50 mg/kg/day) combined to sorafenib (15 mg/kg/day) (concomitant schedule, n=3) or metformin (50 mg/kg/day) followed 6 hours later by sorafenib (15 mg/kg/day) (sequential schedule, n=3). Total RNA was amplified and labelled using the GeneChipTM WT PLUS Reagent Kit (ThermoFischer Scientific). Each RNA sample was hybridized to Human ClariomTM S GeneChip (ThermoFischer Scientific). Arrays were scanned, and images were analyzed and controlled for hybridization artefacts. The microarray data were normalized using Signal Space Transformation-RMA (SST-RMA) which is optimized for under-estimation of true fold changes. Following normalization, differential expression was carried out using eBayes function and One-Way Anova statistical analysis. The analysis was carried out using Transcriptome Analysis Console software (ThermoFischer Scientific, version 4.0.2) with p < 0.05 considered as statistically significant. The differentially expressed genes were then subjected to absolute GSEA searching through more than 10,000 different cellular pathways.

first: Rania

last: Harati

first: Marc

last: Vandamme

first: Benoit

last: Blanchet

first: Christophe

last: Bardin

first: Françoise

last: Praz

first: Rifat

last: Hamoudi


title: EWSR1 regulates PRDM9-dependent histone 3 methylation and links recombinant hotspots to the chromosomal axis

status: Public on Dec 17 2020

city: Bar Harbor

first: Hui

last: Tian

state: ME

address: 600 Main Street

department: Research

institute: The Jackson Laboratory

summary: Meiotic recombination in mammals requires recombination hotspot activation through the action of the histone 3 lysine-4 and lysine-36 methyltransferase PRDM9 to ensure successful double-strand break initiation and repair. Here using a Ewsr1 male germ cell conditional knockout mouse, we show that EWSR1 increases the efficiency of PRDM9-dependent H3K4/K36 trimethylation at the adjacent nucleosomes in vivo, and directs the hotspot choices for double-strand breaks formation and their subsequent repair into sufficient number of crossovers properly positioned along the centromere-telomere axis.

overall_design: We performed ChIP-seq with antibodies against H3K4me3 and H3K36me3 using chromatin extracted from control, Prdm9 PR/SET domain mutant and Ewsr1 conditional knockout mouse spermatocytes, and compared it to previously generated ChIP-seq data for H3K4me3 and H3K36me3 in the same cell type. We also performed ChIP-seq with antibody against single strand DNA binding protein DMC1 in spermatocytes isolated from control, Prdm9 PR/SET domain mutant, Prdm9 heterozygous and Ewsr1 conditional knockout mouse to analyze meiotic DNA double strand formation level

first: Petko

last: Petkov


title: Reproductive history determines ErbB2 locus amplification, WNT signalling and tumor phenotype in a murine breast cancer model [RNA-seq]

status: Public on Nov 12 2020

summary: Understanding the mechanisms underlying tumor heterogeneity is key to development of treatments that can target specific tumor subtypes. We have previously targeted CRE recombinase-dependent conditional deletion of the tumor suppressor genes Brca1, Brca2, p53 and/or Pten to basal or luminal ER- cells of the mouse mammary epithelium. We demonstrated that both the cell-of-origin and the tumor-initiating genetic lesions co-operate to influence mammary tumor phenotype. Here, we use a CRE-activated HER2 orthologue to specifically target HER2/ERBB2 oncogenic activity to basal or luminal ER- mammary epithelial cells and carry out a detailed analysis of the tumors which develop. We find that in contrast to our previous studies, basal epithelial cells are refractory to transformation by the activated NeuKI allele, with mammary epithelial tumor formation largely confined to luminal ER- cells. Histologically, the majority of tumors that developed were classified as either adenocarcinomas of no special type or metaplastic adenosquamous tumors. Remarkably, the former were more strongly associated with virgin animals and were typically characterised by amplification of the NeuNT/ErbB2 locus and activation of non-canonical WNT signalling. In contrast, tumors characterised by squamous metaplasia were associated with animals that had been through at least one pregnancy and typically had lower levels of NeuNT/ErbB2 locus amplification but had activated canonical WNT signalling. Squamous changes in these tumors were associated with activation of the Epidermal Differentiation Cluster. Thus, in this model of HER2 breast cancer, cell-of-origin, reproductive history, NeuNT/ErbB2 locus amplification, and the activation of specific branches of the WNT signalling pathway all interact to drive inter-tumor heterogeneity.

overall_design: Mice carrying an engineered active orthologue of the HER2/ERBB2 gene knocked into the endogenous locus but with expression blocked by a FLOX-STOP-FLOX cassette were crossed with mice carrying the Beta-lactoglobulin promoter-driven CRE recombinase transgene, which targets CRE expression to the luminal cells of the mammary gland. Some mice were put through one or more rounds of pregnancy. Ten tumors which developed in the mammary glands of ten mice were snap frozen and processed for gene expression analysis by RNA-seq. One sample failed QC. Non-negative matrix factorization was used to cluster RNAseq data to determine which characterstic (reproductive history, tumor diagnosis or amplification status of the Erbb2/Neu locus) accounted for the greatest difference between samples. Amplification status of the Erbb2/Neu locus was selected as giving the greatest difference between samples, with one sample, B67_1006, proving to be an outlier. Therefore, differential expression comparing high amplification against non/low amplification samples was carried out, excluding B67_1006.

middle: J


title: Different expression of lipid metabolism-related genes in Black cattle and Luxi cattle based on transcriptome analysis

city: Chengyang District, Qingdao City, Shandong Province

first: xianxun

last: liu

address: 700 Changcheng Road

institute: Qingdao Agricultural University

summary: Black cattle is a new breed of beef cattle developed by combining modern biotechnologies such as somatic cell cloning and conventional breeding methods. To provide new ideas for improving meat quality and generating new breeds of cattle, the important candidate genes affecting fat deposition in two kinds of cattle were identified. Eighteen months Black cattles and Luxi cattles were randomly assigned into two environmental. The longissimus dorsi muscle were collected on Black cattle and Luxi cattle,for analyses including fatty acid determinationrs, high-throughput sequencing metagenomics, qRT-PCR expression profile and western blot.The ratio of unsaturated fatty acids to saturated fatty acids was 1.37:1 and 1.24:1 in the muscle tissues of Black cattle and Luxi cattle, respectively. The results of RNA-Seq analysis revealed 1,415 DEGs(fold change ≥ ± 2, P<0.05) between the longissimus dorsi of Black cattle and yellow cattle. A total of 939 genes were upregulated, and the other 476 genes were downregulated. With GO enrichment analysis, it was found that the identified DEGs were significantly enriched in biological regulation, regulation of the Wnt signaling pathway, negative regulation of the Wnt signaling pathway, cAMP metabolic process, fat cell differentiation, and brown fat cell differentiation, among other functions. Regulation of lipolysis in adipocytes, AMPK signaling pathway, adipocytokine signaling pathway and PPAR signaling pathway in the KEGG pathway database were significantly enriched. PPI network analysis showed that the downregulated genes FABP4, ADIPOQ, PLIN1, PLIN2 and LIPE were closely linked to other DEGs and were the key sites of multiple metabolic pathways. Combined with qRT-PCR and protein expression profile analysis, the expression level of fat acid metabolism related genes (FABP4, ADIPOQ) in black cattle was high and the difference was significant. Changes in the expression of fatty acid metabolism-related genes in Black cattle and Luxi cattle were analyzed and important candidate marker genes (such as ADIPOQ and FABP4) that affect fat deposition were identified in order to provide a genetic basis for the efficient breeding of production performance, establish a molecular marker database for local cattle breeds and support the cultivation of new breeds.

overall_design: Different expression of lipid metabolism-related genes in muscle from  Black cattle and Luxi cattle

first: Ruili

last: Liu

first: Xianxun

first: Xuejin

last: Bai

first: Yajuan

last: Dong


title: Estrogen-related receptor γ agonist DY131 ameliorates lipopolysaccharide-induced acute liver injury

status: Public on Nov 06 2020

city: Nanjing

first: Ma

last: Yang

state: Jiangsu

address: No. 140 Hanzhong Rd

institute: Department of Nephrology

summary: DY131 is a pharmacological agonist of the orphan receptor estrogen-related receptor (ERR) γ which plays a crucial role in regulating energy generation, oxidative metabolism, cell apoptosis, inflammatory responses, etc. However, its role in acute liver injury is unknown. In the study, we evaluated the effect of DY131 on lipopolysaccharide (LPS)-induced liver injury. Transcriptomics analysis revealed that the dysregulated pathways associated with inflammation and metabolism were significantly reversed by DY131 in LPS-treated mice, providing more evidence in favor of the protective effect of DY131 against LPS-induced liver injury.

overall_design: Mice were pretreated with DY131 through intraperitoneal injection at a dose of 5 mg/kg/day for three days prior to LPS challenge (10mg/kg). 24 hours later, they were anesthetized and sacrificed. Blood and liver tissues were collected for further studies.

first: H

middle: Y

last: Ma

first: J

first: W

last: Gong


title: Glycosyltransferase ST6Gal-I promotes the epithelial to mesenchymal transition in pancreatic cancer cells

status: Public on Nov 02 2020

city: Birmingham

first: Susan

last: Bellis

state: ALABAMA

address: MCLM 350, 1918 University Boulevard

institute: University of Alabama at Birmingham

summary: RNA-Seq results suggested that epithelial to mesenchymal transition (EMT) -related pathways were upregulated in the metastatic subclones.  Furthermore, forced expression of ST6Gal-I in the parental Suit2 line was sufficient to upregulate EMT, indicating a functional role for ST6Gal-I in this process.

overall_design: RNA-Seq was conducted on poorly-metastatic Suit2 pancreatic cancer cells and Suit2-derived metastatic subclones, as well as Suit2 cells with forced expression of ST6Gal-I.

first: Yvonne

last: Edwards


title: Landscape of genomic alterations in Indian Urinary Bladder cancer patients

status: Public on Nov 01 2020

city: KOLKATA

first: CHINMAY

middle: KUMAR

last: PANDA

state: WEST BENGAL

address: 37, SP MUKHERJEE ROAD, KOLKATA

department: ONCOGENE REGULATION UNIT

country: India

institute: CHITTARANJAN NATIONAL CANCER INSTITUTE, INDIA

type: Genome binding/occupancy profiling by genome tiling array

type: Genome binding/occupancy profiling by SNP array

summary: Urinary bladder is an open system and is under constant carcinogenic insults leading to chromosomal aberrations. Chronic retention of urine in the bladder is   considered one of the reasons for initiation of bladder cancer. Here, we have explored the genomic alterations in Indian BlCa patients by CGH-SNP microarray.   The microarray data (n=10) revealed several alterations throughout the genome. Genomic regions like 2p25.3 [chr2: 3557810-3575411],   4p16.3 [chr4: 656211-657032], 7p22.3 [chr7: 990730-1000575], 10q26.13 [chr10: 123137874-123148372], 16q24.3 [chr16: 88835825-88837868],   17p11.2 [chr17: 17477220-17506264], 19q13.33 [chr19: 49861971-49862491], 20q13.33 [chr20: 63406442-63415195; 63415195-63495407] etc.   were found to be frequently (≥40%) amplified among the 10 BlCa samples. Regions like 2q37.2 [chr2: 235384329- 235479250],   8p23.3-11.21 [chr 8: 236477- 39291988], 10q21.1-26.3 [chr 10: 55309072- 133391562], 11p12 [chr 11: 37327799- 38803561],   12q24.31-24.33 [chr 12: 128515048- 133047983], and 13q13.3-34 [chr 13: 35583974- 113996673] etc. are identified as common   deletion (30%). Chromosomal regions like 2p25.3, 16q24.3 and 20q13.33 showed the highest frequency of amplification (70%),   whereas 2q37.2, 10q21.1, 11p12, 11q22.3, etc. showed the highest frequency of deletion (30%). Chromosome 7, 15 and 21 showed   the least alterations in these samples. Almost a complete loss in 8p chromosomal arm (8p23.1-11.22), chromosome 11q (11q14.1-25)   and chromosome 13 (13q13.3-34) were found in 30% of samples, signifying their plausible role in development of BlCa.

overall_design: This study was done with human urinary bladder tumor tissue and respective Peripheral Blood Lymphocyte high-quality DNA using Agilent two-color CGH+SNP  2 X 400K Array (AMADID:028081). The results implicated great number of genomic aberrations in the tissues, which may be involved in the progression of the disease.

first: Mukta

last: Basu

first: Amlan

last: Ghosh

first: Chinmay

middle: K

last: Panda


title: Reproductive history determines ErbB2 locus amplification, WNT signalling and tumor phenotype in a murine breast cancer model [exome sequencing]

first: Matthew

last: Smalley

overall_design: Mice carrying an engineered active orthologue of the HER2/ERBB2 gene knocked into the endogenous locus but with expression blocked by a FLOX-STOP-FLOX cassette were crossed with mice carrying the Beta-lactoglobulin promoter-driven CRE recombinase transgene, which targets CRE expression to the luminal cells of the mammary gland. Some mice were put through one or more rounds of pregnancy. Ten tumors which developed in the mammary glands of ten mice were snap frozen and processed for gene expression analysis by exome sequencing together with matched spleens. One sample failed QC. Exome sequencing was used to assess point mutations in tumors and also genomic copy number variations.


title: PfAP2-HC, an unusual, heterochromatin-associated ApiAP2 factor of Plasmodium falciparum

status: Public on Nov 04 2020

city: Nijmegen

first: Roel

last: Cooijmans

address: Geert Grooteplein Zuid 30

department: Molecular biology

country: Netherlands

institute: RIMLS

summary: In this study we have investigated PfAP2-HC (PF3D7_1456000), a protein that was identified by co-immunoprecipitation with PfHP1 coupled with liquid chromatography-tandem mass spectrometry. PfAP2-HC belongs to the ApiAP2 family, the main transcription factor family in Apicomplexan parasites. We have confirmed that AP2-HC colocalises with HP1 with the use of immunofluorescence assays and chromatin immunoprecipitation-sequencing. We show that PfAP2-HC is not required for heterochromatin maintenance and inheritance with the use of PfAP2-HC Kock out and knockdown. We show with transcriptome-wide microarray time course analysis that PfAP2-HC does not act as a transcription factor in blood stage parasites. We demonstrate that the AP2 domain is dispensable for heterochromatin targeting by introducing a premature stop codon before the AP2 domain. We show that PfAP2-HC binding to heterochromatin dependents on PfHP1. and PfAP2-HC is likely not involved in de novo heterochromatin formation

overall_design: 17 samples were analysed, 11 ChIP samples (7 PfHP1 and 4 GFP) and 6 input control samples. Biological replicates for the HP1mSglmSON samples were performed, to eliminate experimental error, due to the abstains of signal in the GFP ChIP (AP2-HC/AP2-14)

first: Eilidh

last: Carrington

first: Dominique

last: Keller

first: Christa

middle: G

last: Toenhake

first: Alexia

last: Loynton-Ferrand

first: Richárd

last: Bártfai

first: Till

middle: S

last: Voss


title: Identification of cellular context sensitive regulatory variation in mouse genomes

status: Public on Nov 25 2020

city: New York

last: Maurano

state: NY

address: 435 East 30th Street

department: Institute for Systems Genetics

institute: NYU School of Medicine

summary: RNA-seq and DHS mapping in hybrid mouse kidney, liver, lung, and B cells

overall_design: 138 DNase-seq and 17 RNA-seq samples collected from 4 tissues in 5 hybrid mouse strains


title: Schengen-pathway controls spatially separated and chemically distinct lignin deposition in the endodermis

city: Chapel Hill

first: Isai

last: Salas Gonzalez

address: 250 Bell Tower Drive, Room 4244

institute: The University of North Carolina at Chapel Hill

summary: Schengen-pathway modulates the phenylpropanoid pathway and induces defense-related mechanisms

overall_design: To investigate the biosynthesis of this H-rich stress lignin we performed both RNA-seq on root tips (5 mm) of WT plants, mutants and treatment showing a strong activation of the Schengen-pathway (WT  treated with exogenous CIF2, myb36.2 and esb1.1) and mutants with no Schengen signalling (sgn3.3, esb1.1 sgn3.3, sgn3.3 myb36.2 and sgn3.3 treated with exogenous CIF2).

first: Guilhem

last: Reyt

last: Salas-Gonzalez


title: Targeted suppression of metastasis controlling transcription factor SOX2 in various cancers using a sequence-specific designer Pyrrole-Imidazole Polyamide

status: Public on Oct 31 2020

city: Kyoto

first: Madhu

last: Malinee

state: Kyoto

address: Kitashirakawa-Oiwakecho,

department: Graduate School of Science

country: Japan

institute: Kyoto University

summary: SOX2 is a well-known transcription factor that plays a central role in maintaiing self renewal, pluripotency of embryonic stem cells, and cell fate determination during embryonic development. N-Methylpyrrole-N-methylimidazole polyamides (PIPs) are chemical compounds that can selectively recognize DNA sequence and achieve operative control over gene expression. In this study, the antitumor activity of a PIP targeting SOX2 binding sequence (SOX2i), which was recently shown to induce the targeted differentiation of stem cells into cardiomyocytes, was evaluated.

overall_design: Genome-wide gene expression change in SW620 by treatment of SOX2i were examined.

first: Hiroshi

last: Sugiyama


title: RBMX suppresses tumorigenicity and progression of bladder cancer via interaction with hnRNP A1 protein to regulate PKM alternative splicing

status: Public on Nov 23 2020

city: guangzhou

first: jizhong

last: wang

address: 63 Duobao Road

institute: Guangzhou Medical University

summary: Bladder cancer (BCa) is one of the most common malignant tumors of urinary system and has high incidence rate and mortality but there is a lack of effective treatment. Therefore, an in-depth study of the molecular mechanisms involved in BCa is of great significance for improving the survival of patients with advanced BCa. We found that the expression of RBMX was significantly declined in BCa, especially in muscle-invasive BCa. BCa patients with low levels of RBMX had poor prognoses. By in vitro and in vivo experiments, RBMX was shown to inhibit BCa cells growth and metastasis. Co-IP coupled with MS and GO analysis identified hnRNP A1 as a RBMX-binding protein. Mechanistically, RBMX competitively bond to the RGG box of hnRNP A1 and antagonized the hnRNP A1-mediated regulation of PKM splicing by blocking the binding of the RGG motif of hnRNP A1 to the sequences flanking PKM exon 9, resulting in downregulation of PKM2 and upregulation of PKM1. By decreasing the PKM2/PKM1 ratio, RBMX suppressed BCa cells aerobic glycolysis, which further inhibited tumorigenicity and progression of BCa.

overall_design: Overexpression of RBMX by plasmid transfection followed by examining alternative splicing changes by RNA-seq or examining cancer phenotype by established assays.

first: Qiuxia

last: Yan


title: Sex-Specific Differences of Humoral Immunity and Transcriptome Diversification in Older Adults Vaccinated with Inactivated Quadrivalent Influenza Vaccines

status: Public on Nov 09 2020

city: Wuhan

first: Jing

last: Yang

address: No. 1, Gold industrial park, Zhengdian street, Jiangxia district

institute: Wuhan Institute of Biological Products Co., Ltd.

summary: Clinical data showed sex variability in the immune response to influenza vaccination, this study aimed to investigate differentially expressed genes that contribute to sex-bias immune responses to quadrivalent inactivated influenza vaccines (QIVs) in older subjects. A cohort of 60 healthy older adults aged 60 to 80 years old were vaccinated with quadrivalent inactivated influenza vaccines (QIVs), and gene expression was analyzed at Day 0 pre-vaccination, Day 3, Day 28 and Day 180 post-vaccination. Sexual dimorphism of humoral immunity was analyzed by HAI assay, and the correlation of gene expression patterns of two sex groups with humoral immune response to vaccination was analyzed. The differentially expressed genes involved in type I interferon signaling pathway and complement activation of classical pathway were upregulated within 3 days post-vaccination in females. At Day 28 after immunization, the immune response showed a man-bias pattern associated with the regulation of protein processing and complement activation of classical pathway. A list of differentially expressed genes associated with variant responses to influenza vaccination between women and men were identified by biology-driven clustering. Old women have a greater immune response to QIVs but rapid decline of antibody levels, while old men have the advantages to sustain a durable response to influenza vaccination. In addition, we identified genes that may contribute to the sex variations toward influenza vaccination in the aged. Our findings highlight the importance of developing personalized seasonal influenza vaccines.

overall_design: Peripheral whole Blood mRNA profiles of 16 older adults aged 60 year or older after inactivated quadrivalent influenza vaccines inoculation(pre-vaccination Day 0, and Day 3, Day 28, Day 180 post-vacciantion)

first: Xiaoyuan

last: Huang

first: Jiayou

last: Zhang

first: Renfeng

last: Fan

first: Wei

last: Zhao

first: Jikai

first: Xiaoming


title: Primate cell fusion disentangles gene regulatory divergence in neural development

status: Public on Nov 17 2020

first: Rachel

last: Agoglia

address: 371 Jane Stanford Way

summary: Among primates, humans display a unique trajectory of development responsible for the many traits specific to our species. However, the inaccessibility of human and chimpanzee primary tissues has limited our ability to study human evolution. Comparative in vitro approaches using primate-derived induced pluripotent stem cells have begun to reveal species differences on the cellular and molecular levels. In particular, brain organoids have emerged as a promising platform to study primate neural development in vitro, although cross-species comparisons of organoids are complicated by differences in developmental timing and variability of differentiation. Here, we developed a new platform to address these limitations. We first generated a panel of tetraploid hybrid stem cells by fusing human and chimpanzee induced pluripotent stem cells. We next applied this approach to study species divergence in cerebral cortical development by differentiating them into neural organoids. We found that hybrid organoids provide a controlled system for disentangling cis- and trans-acting gene expression divergence across cell types and developmental stages, revealing a signature of selection on astrocyte-related genes. In addition, we identified an up-regulation of human somatostatin receptor 2 (SSTR2), which regulates neuronal calcium signaling and is associated with neuropsychiatric disorders.  We discovered a human-specific response to modulation of SSTR2 function in cortical neurons, underscoring the potential of this unique platform to reveal the molecular basis of human evolution.

sample_organism:
Pan troglodytes
Homo sapiens

platform_organism:
Pan troglodytes
Homo sapiens

overall_design: RNA sequencing of iPS cells and iPS cell-derived cortical spheroids from three human, three chimpanzee and five hybrid cell lines

first: Hunter

last: Fraser

first: Sergiu

last: Pasca


title: RNA-sequencing data for 923 enhancer deletion newborn mouse whole skin

status: Public on Oct 30 2020

city: St Louis

first: Cristina

last: de Guzman Strong

state: MO

address: 660 South Euclid

department: Dermatology

institute: Washington University in St Louis

summary: We examined the RNA expression profiles of whole skin from newborn CRISPR-edited mice that have the 923 enhancer located within the Epidermial Differentiation Complex deleted. We examined two independent deletion lines, 923del and 923large, both homozygous mutants and heterozygotes, as well as wild type mice and involucrin Ivl knockout mice. We find that Ivl, the gene most proximal to 923, is downregulated upon the loss of the enhancer in both independent deletions, and 4 additional genes are also downregulated in both deletion lines, a secondary effect of the loss of Ivl, as concluded by comparing to the Ivl KO mice. We thus conclude that Ivl is the primary target gene of the 923 enhancer and establish a 923-Ivl regulatory module.

overall_design: Examination of RNA expression profiles in two  923 enhancer deletion lines, the Ivl knockout line and WT  newborn mouse whole skins.

first: Mary

middle: E

last: Mathyer


title: Comprehensive analysis of coding and non-Coding RNA transcriptomes related to hypoxic adaptation in the Tibetan chickens

status: Public on Oct 28 2020

city: Beijing

first: Ying

address: Haidian District, Yuanmingyuan West Road 2

institute: China Agricultural University

type: Non-coding RNA profiling by high throughput sequencing

summary: Tibetan chicken has a suite of adaptive features to tolerate the high-altitude hypoxic environment as a unique native breed in Qinghai-Tibet Plateau of China. Increasing evidence suggests that long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) have roles in hypoxic adaptation of high-altitude animals, though their exact contributions remain unclear. This study aims to uncover the global landscape of mRNAs, lncRNAs and miRNAs using transcriptome sequencing so as to construct a regulatory network of competing endogenous RNAs (ceRNAs) provide a new sight for the hypoxic adaptation of Tibetan chicken embryos. In the study, 354 differentially expressed mRNAs (DEGs), 389 differentially expressed lncRNAs (DELs) and 73 differentially expressed miRNAs (DEMs) were identified between Tibetan (TC) and Chahua chicken (CH). The functional analysis showed that several important DEMs and their targets of DELs and DEMs are involved in angiogenesis (include blood vessel development and blood circulation) and energy metabolism (include glucose, carbohydrate and lipid metabolism). The ceRNA network was then constructed with the predicted pairs of DEGs-DEMs-DELs which further revealed regulatory roles of these differentially expressed RNAs in hypoxic adaptation of Tibetan chicken.

overall_design: Chorioallantoic membrane (CAM) of embryos coding and non-coding RNA profiles of 11-day old Tibetan chickens (TCs) and Chahua chickens (CHs) were generated by deep sequencing, in triplicate.

first: Woyu

last: Su

first: Bo

first: Hao


title: miRNA profiling in RAW264.7 cells treated without or with LPS and IFNγ

city: Jinan

first: Xiaoxiao

last: Zhu

address: 18877 Jingshi Road

institute: Shandong First Medical University & Shandong Academy of Medical Sciences

summary:
We have identified a number of miRNAs that are differentially expressed in LPS and IFNγ treated RAW264.7 cells, as compared to untreated cells.
Microarray and quantitative real-time PCR (qRT-PCR) results showed that miR-103 decreased while the STAT1 expression increased substantially in RAW264.7 derived M1 macrophage, and there was a significant negative correlation between miR-103 and STAT1.Overexpression of miR-103 suppressed M1 polarization by inhibiting STAT1/IRF1 signaling pathway and vice versa in vitro. STAT1 is a direct target of of miR-103.

overall_design: RAW264.7 cells treated with  LPS and IFNγ for 24h to generate M1-polarized macrophages. RAW264.7 (n= 3) and RAW264.7-derived M1 macrophages (n= 3) were used to evaluate miRNAs expression. RNA was isolated and miRNA array assays were performed by OE Biotech (mouse miRNA, release 21.0, 8*60K, Design ID: 070155))

first: Qiang

last: Guo

first: Ran

last: Wei

first: Lin

first: Zhen

first: Xia

last: Li


title: Single-cell RNA sequencing reveals the mesangial identity and species diversity of glomerular cell transcriptomes

status: Public on Nov 21 2020

city: Huddinge

first: Ping

last: Chen

address: Alfred Nobels allé 8

department: Department of Laboratory Medicine

country: Sweden

institute: Karolinska Institutet

summary: Molecular characterization of the individual cell types in human kidney as well as model organisms are critical in defining organ function and understanding translational aspects of biomedical research. Previous studies have uncovered gene expression profiles of several kidney glomerular cell types, however, important cells, including mesangial (MCs) and glomerular parietal epithelial cells (PECs), were missing or incompletely described, and a systematic comparison between mouse and human kidney is lacking. To this end, we used Smart-seq2 to profile 4332 individual glomerulus-associated cells isolated from human living donor renal biopsies and mouse kidney. The analysis revealed genetic programs for all four glomerular cell types (podocytes, glomerular endothelial cells, MCs and PECs) as well as rare glomerulus-associated macula densa cells (MDCs). Importantly, we detected heterogeneity in glomerulus-associated Pdgfrb-expressing cells, including bona fide intraglomerular MCs with the functionally active phagocytic molecular machinery, as well as a unique mural cell type located in the central stalk region of the glomerulus tuft. Furthermore, we observed remarkable species differences in the individual gene expression profiles of defined glomerular cell types that highlight translational challenges in the field and provide a guide to design translational studies.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design:
Smart-seq2 single-cell RNA-seq

Raw data for GSM4855911/Human_kidney_GLOM will be submitted to EGA due to patient privacy concerns.

first: Bing

last: He

first: Jaakko

last: Patrakka


title: Identification of required host factors for SARS-CoV-2 infection in human cells

status: Public on Oct 24 2020

city: New York City

first: Zharko

last: Daniloski

address: 1 Washington square village, 2P

institute: New York University

summary: We performed a genome-wide CRISPR knockout screen to identify human genes required for SARS-CoV-2 infection.

overall_design: Human A549-ACE2 CRISPR-Cas9 perturbed cells were infected with SARS-CoV-2 at two MOIs (0.01 and 0.3), and cells were collected for genomic DNA isolation 6 days post-infection.

first: Tristan

middle: X

last: Jordan

first: Ben

last: tenOever

first: Neville

last: Sanjana


title: Transcriptomic data of MDA-MB-231 cells adapted to culture in media containing 0 mM or 2 mM metformin and cultured as mammospheres

status: Public on Oct 23 2020

city: Southampton

first: Jeremy

middle: Paul

last: Blaydes

state: Hampshire

address: Southampton General Hospital, Tremona Road

department: Faculty of Medicine

institute: University of Southampton

summary: We report the single-cell RNA sequencing data obtained from MDA-MB-231 breast cancer cells cultured in standard DMEM (high glucose) media, or adapted to culture in standard DMEM (high glucose) media containing 2 mM metformin, and then cultured as mammospheres

overall_design: Examination of transcriptomic changes in MDA-MB-231 breast cancer cells mammospheres in response to adaptation to culture in 2 mM metformin

middle: P

first: Matthew

last: Rose-Zerilli


title: Genome-wide DNA methylation and RNA-seq analyses identify genes and pathways associated with doxorubicin resistance in canine diffuse large B-cell lymphoma cell line

status: Public on Oct 22 2020

city: Taipei

first: Chen-Si

last: Lin

address: No. 1, Sec. 4, Roosevelt Road

department: Department of Veterinary Medicine

country: Taiwan

institute: Nationwide Taiwan University

type: Methylation profiling by high throughput sequencing

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

overall_design: Refer to individual Series


title: Kidney Intercalated Cells Phagocytose and Acidify Escherichia coli

city: Indianapolis

first: Andrew

last: Schwaderer

state: IN

address: 1044 West Walnut Street

department: Pediatric Nephrology

institute: Indiana University School of Medicine

summary: Intercalated cells are known to be involved in acid-base homeostasis via vacuolar ATPase (H+-ATPase or V-ATPase) expression. Increasing evidence supports an innate immune role for ICs along with their traditional function of pH regulation. In this study, human kidney tissue was enriched for viable intercalated cells then exposed to uropathogenic E. coli versus saline control. Single cell transcriptomics was performed. Six intercalated cell subtypes were identified including hybrid principal-intercalated cells. Cell specific cluster marker gene list generated from this sequencing data was put through ingenuity pathway analysis pipeline which predicted “phagosome maturation” as a key biological pathway that increased in rank following exposure to uropathogenic E. coli in two of the intercalated cell subtypes. Uptake of  E. coli and pHrodo coated E. coli BioParticlesTM during live animal intravital microscopy demonstrated that intercalated cell phagocytosis of bacteria was an active process that involved acidification. Taken together, our finding indicate that intercalated cells represent an epithelial cell with characteristics of professional phagocytes like macrophages or neutrophils, which includes the ability to phagocytose E. coli and acidify phagolysosomes.

overall_design: Normal margin of the human kidney biopsy sample was processed for single cell suspension with enzymatic digestion (Liberase TL and DNAse I) and rapid dissociation using GentleMacs (Miltenyi Biotec). Dead cells were then removed from single cell suspension using dead cell removal microbead (Miltenyi Biotec). CD45+ traditional immune cells were then removed using anti-human CD45 microeads (Miltenyi Biotec). Intercalated cells were then enriched using anti-human C-KIT (CD117) microbeads (Miltenyi Biotec). Cell viability was tested on hemocytometer. Viable cells were equally divided into 2 wells of the 96-well U bottom plate and exposed to uropathogenic E.coli (UPEC) for 1 hr and sterile saline at 370C and 5% CO2 environment. After washing with sterile PBS and re-suspension in PBS (without Ca2+ and Mg2+) both cells samples (Saline control and UPEC exposed) were immediately processed for single cell sequencing using 10x genomics platform. Single cells of both samples were sequenced on Illumina Novaseq 6000 instrument.

first: Vijay

last: Saxena

first: Hongyu

last: Gao

first: Samuel

last: Arregui

first: Amy

last: Zollman

first: Malgorzata

middle: M

last: Kamocka

first: Xiaoling

last: Xuei

first: Patrick

last: McGuire

first: Michael

last: Hutchens

first: Takashi

last: Hato

first: David

last: Hains

middle: L


title: Genome-wide RNA-seq analysis in canine diffuse large B-cell (cDLBCL) cell lines

summary: To analyze the biological relevance of methylation changes, gene expression from both CLBL-1 and CLBL-1 8.0, a doxorubicin-resistanct cDLBCL cell line, were examined. In total, we obtained 24.8 M and 23.7 M sequence reads for CLBL-1 and CLBL-1 8.0, respectively. After filtering out low-quality bases and reads from the datasets prior to read mapping, 19.8 M reads for CLBL-1 and 19.1 M reads for CLBL-1 8.0 were mapped to canine genome. The criteria that transcripts per million (TPM) >5 in one of sample and the log2 fold-change ≥ 0.67 or ≤ -0.67 were applied to identified differentially expressed genes. Upregulated and downregulated genes were 1127 and 602, respectively.

overall_design: DNA expression profiles of CLBL-1 8.0 and CLBL-1 as control were generated by deep sequencing using Illumina Novaseq 6000.

first: Chia-Hsin

last: Hsu

first: Wei-Chi

last: Wang


title: Genome-wide DNA methylation analysis using MethylCap-seq in canine diffuse large B-cell (cDLBCL) cell lines

summary: MethylCap-seq was performed to characterize the genome-wide DNA methylation in CLBL-1 8.0, a doxorubicin-resistant cDLBCL cell line, and CLBL-1 as control, and aimed to investigate underlying mechanisms of doxorubicin resistance in cDLBCL. Strong difference in methylation pattern of both promoter and gene-body regions was detected between CLBL-1 8.0 and CLBL-1. Methylated peaks with fold-change greater than 4 and a P value of less than 0.01 were defined as differentially methylated regions (DMRs); there were a total of 20289 hypermethylated and 38362 hypomethylated DMRs detected. Among these, 1339 hypermethylated and 4861 hypomethylated DMRs were in promoter regions, while 12855 hypermethylated and 18904 hypomethylated DMRs were in gene-body regions. There were 6.6% hypermethylated DMRs located on the promoter regions. A high percentage (63.4%) of hypermethylated DMRs were distributed within the gene body, and 7.8%, 5.1% and 23.8% hypermethylated DMRs were found to be clustered in upstream, downstream, and intergenic regions, respectively.

overall_design: DNA methylation profiles of CLBL-1 8.0 and CLBL-1 as control were generated by deep sequencing using Illumina Novaseq.


title: A potential role for PLAGL1 in the regulation of blood vessel development

status: Public on Nov 18 2020

city: Ames

first: Geetu

last: Tuteja

state: IA

address: 2106 Molecular Biology Building

department: Genetics, Development and Cell Biology

institute: Iowa State University

summary: Throughout pregnancy, the placenta is important for the well-being of the fetus, transporting nutrients and waste between the maternal and fetal blood supply, secreting hormones, and serving as a protective barrier. To obtain a more complete understanding of placental development, we must understand how placental gene expression is regulated. We used RNA-seq data and ChIP-seq data for the enhancer associated mark, H3k27Ac, to study gene regulation in the mouse placenta at embryonic day (e) 9.5, when the placenta is developing a complex network of blood vessels which facilitate maternal-fetal exchange of material. We identified several transcription factors upregulated at e9.5 with enriched binding sites in e9.5-specific enhancers. We performed follow up studies to investigate the role of the most enriched transcription factor, PLAGL1, in placental development. The PLAGL1 binding motif was found in 233 regions which were significantly associated with genes involved in vasculature development. To further understand the role of PLAGL1, we performed an siRNA knockdown in a human placental cell line and analyzed the genes that significantly changed in expression compared to cells treated with a negative control siRNA. Genes that significantly decreased in expression along with PLAGL1 were associated with placental vasculature development terms. Using a tube assay, we found that decreased PLAGL1 expression lead to reduced tube formation. By identifying PLAGL1 as an important regulator in the placenta, our analysis improves our understanding of gene regulation in the placenta, an important step towards developing methods to diagnose and treat placental diseases.

overall_design:
Examine changes in gene expression upon PLAGL1 knockdown.
We performed RNA-seq in the human placental cell line, HTR-8/Svneo, with PLAGL1 siRNA and a negative control siRNA. We identified 4,003 differentially expressed genes, of which 1,964 were upregulated in the knockdown and 2,039 were downregulated in the knockdown. Genes downregulated along with PLAGL1 were associated with placenta vasculture development.

first: Rebekah

last: Starks

first: Rabab

last: Abu-Alhasan

first: Haninder

last: Kaur


title: Keratinocyte-Macrophage Crosstalk by the Nrf2/Ccl2/EGF Signaling Axis Orchestrates Tissue Regeneration

status: Public on Oct 12 2020

first: Alvaro

last: Villarreal-Ponce

state: New York

address: Skirball Institute of Biomolecular Medicine Floor 2 Lab 3

department: Plastic Surgery

institute: NYU Langone Health

summary: Unveiling the molecular mechanisms underlying tissue regeneration provides new opportunities to develop treatments for diabetic ulcers and other chronic skin lesions. Here, to gain insight into critical wound response mechanisms regulated by epidermal Nrf2, we micro dissected a ~2.5mm concentric ring around the 10mm wound at 5 days-post-wounding (DPW) and used flow cytometry to isolate wound-associated keratinocytes from Nrf2 +/+ Ker and Nrf2 ∆/∆ Ker mice for bulk RNA-seq. Combining in vivo and ex vivo data with next generation sequencing, we show that Ccl2 secretion by epidermal keratinocytes is directly orchestrated by Nrf2, a prominent transcriptional regulator of tissue regeneration that is activated early after cutaneous injury. Through a unique feedback mechanism, we find that Ccl2 from epidermal keratinocytes not only drives chemotaxis of macrophages into the wound, but also triggers macrophage expression of EGF, which in turn activates basal epidermal keratinocyte proliferation. Notably, we find dysfunctional activation of Nrf2 in epidermal keratinocytes of diabetic mice after wounding, which in part, explains regenerative impairments associated with diabetes. These findings provide mechanistic insight into the critical relationship between keratinocyte-macrophage signaling during tissue regeneration, providing the basis for continued investigation of the therapeutic value of Nrf2.

overall_design: Examination of Nrf2 regulated genes in  keratinocytes during wound repair

first: Daniel

last: Ceradini


title: Impact of dioxin exposure and high fat diet feeding on adult mouse islets

status: Public on Oct 05 2020

city: Ottawa

first: Matthew

last: Meier

state: Ontario

address: 251 Sir Frederick Banting Driveway

department: Health Canada

institute: Government of Canada

summary: Purpose: to determine whether chronic low dose dioxin exposure in adult male and female mice impacts susceptibility to metabolic disease and islet dysfunction.

overall_design: Male and female C57Bl/6 mice were injected i.p. 2x/week with either corn oil or 20 ng/kg/d TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin). Each treatment group was further subdivided to receive either chow diet or 45% kcal high fat diet (HFD). Pancreatic islets were isolated 12 weeks after the beginning of treatment.

first: Jennifer

last: Bruin


title: Mapping the regulatory potential of fixed variants in modern humans

status: Public on Oct 09 2020

city: Jerusalem

last: Gokhman

address: Givat Ram, Safra Campus

country: Israel

institute: The Hebrew University of Jerusalem

summary: The Neanderthal and Denisovan genomes enabled the discovery of sequences that differ between modern and archaic humans, the majority of which are noncoding. However, our understanding of the regulatory consequences of these differences remains limited, in part due to the decay of regulatory marks in ancient samples. Here, we used a massively parallel reporter assay in embryonic stem cells, neural progenitor cells and bone osteoblasts to investigate the regulatory effects of the 14,042 single-nucleotide modern human-specific variants. Overall, 1,791 (13%) of sequences containing these variants showed active regulatory activity, and 407 (23%) of these drove differential expression between human groups. Differentially active sequences were associated with divergent transcription factor binding motifs, and with genes enriched for vocal tract and brain anatomy and function. This work provides insight into the regulatory function of variants that emerged along the modern human lineage and the recent evolution of human gene expression.

overall_design: Massively parallel reporter assay (MPRA) conducted on 14,042 pairs of modern and archaic human alleles in three cell types (ESCs, NPCs and osteoblasts) in three replicates each.

first: Carly

last: Weiss

first: Lana

last: Harshman

first: Fumitaka

last: Inoue

first: Hunter

middle: B

last: Fraser

first: Dmitri

middle: A

last: Petrov

first: Nadav

last: Ahituv


title: The mRNA derived MalH sRNA contributes to alternative carbon source utilization by tuning maltoporin expression in E. coli

city: Edinburgh

first: Sander

last: Granneman

address: Mayfield Road, Kings Buildings, Waddington building, room 3.06

department: Centre for Synthetic and Systems Biology

institute: University of Edinburgh

summary: Previous high-throughput studies in Gram-negative bacteria identified a large number of 3’UTR fragments that potentially function as sRNAs. Here we extensively characterise the MalH sRNA. We show that MalH is a stable degradation intermediate derived from the 3’ end of malG, which is part of the maltose uptake operon transcript malEFG. Unlike the majority of bacterial sRNAs, MalH is transiently expressed during the transition from the exponential to the stationary growth phase, suggesting that it contributes to adaptation to changes in nutrient availability. Over-expression of MalH reduces expression of general outer membrane porins and MicA, a repressor of the high-affinity maltose/maltodextrin transporter LamB. Disrupting MalH production and function significantly reduces lamB accumulation when maltose is the only available carbon source, presumably due to the accumulation of the MicA repressor. We propose that MalH is part of a regulatory network that, during the transition phase, directly or indirectly promotes accumulation of high-affinity maltose transporters in the outer membrane by dampening competing pathways.

overall_design: For pulse-overexpression studies overnight MG1655 cultures containing pBAD::sRNA and empty pBAD+1 control plasmids were inoculated in fresh LB-ampicillin medium at a starting OD600 of 0.05 and grown aerobically at 37°C to OD600 0.4. Pre-induction (0 min) and post-induction samples were harvested. For induction, cultures were supplemented with L-arabinose (Sigma, A3256) and rapidly collected by filtration and flash-frozen in liquid nitrogen at the indicated time-points. RNA was extracted from three biological replicate time-series, followed by RNA extraction using GTC-phenol. RNA-seq libraries were generated using an in-house protocol. Genomic DNA was removed by incubating 10 ug of total RNA with 2U Turbo DNase in a 50 ul final volume for 30 minutes at 37°C in the presence of 10 U superaseIn RNase Inhibitor. RNA was subsequently phenol-chloroform extracted and purified by ethanol-precipitation. Ribosomal RNA was removed with the Illumina Gram-negative Ribo-Zero rRNA Removal Kit according to the manufacturer's instructions. Successful rRNA depletion was verified on the Agilent 2100 Bioanalyzer. The RNA was fragmented for 5 min at 95°C in the presence of Superscript III buffer (Invitrogen) followed by a five-minute incubation on ice. Reverse-transcription (RT) was performed with Superscript III (Thermo Scientific) in 20 ul reactions according to manufacturer's procedures using 250 ng of ribosomal RNA depleted RNA and 2.5 uM random hexamers (PE_solexa_hexamer, 5'-CGTGTGCTCTTCCGATCTNNNNNN-3'). The RNA and free primers were degraded using 20U of Exonuclease I (NEB) and 50U RNaseIf (NEB) and the cDNA was purified with the DNA Clean & Concentrator 5 kit (Zymo Research). Ligation of the 5' P5_phospho_adapter (5'P-AGATCGGAAGAGCGTCGTGTAGGG-3SpC3) to the cDNA was performed using CircLigase II (Lucigen) for 6 hours at 60°C, followed by a 10-minute inactivation at 80°C. The cDNA was purified with the DNA Clean & Concentrator 5 kit. Half of the cDNA library was PCR amplified using Pfu polymerase (Promega) using the P5 forward PCR oligonucleotide (5'-AATGATACGGCGACCACCGAGATCTACACTCTTTCCCTACACGACGCTCTTCCGATCT-3') and barcoded BC reverse oligonucleotides (200 nM; 5'-CAAGCAGAAGACGGCATACGAGAT(BC 6nt barcode)GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT-3'; 95°C for 2 min, 95°C for 20s, 52°C for 30s and 72°C for 1 min, and a final extension of 72°C for 5 min. 20 cycles of amplification). The PCR products were treated with Exonuclease 1 (NEB) for 1 h at 37°C and purified by ethanol precipitation. Libraries were resolved on a 2% MetaPhor agarose gel 200-500 bp fragments were gel-extracted using the MinElute kit. All libraries were quantified on a 2100 Bionalyzer using the High-Sensitivity DNA assay (Agilent). Individual libraries were pooled in equimolar amounts. Paired-end sequencing (75 bp) was performed by Edinburgh Genomics on the Illumina HiSeq 4000 platform.

first: Ira

last: Iosub


title: 923 enhancer deletion newborn mouse

first: Cristina

last: de Guzman Strong


title: FYN-TRAF3IP2 activating fusion oncogene in peripheral T-cell lymphoma

status: Public on Oct 01 2020

city: New York

first: Stuart

middle: Aidan

last: Quinn

address: 1130 St Nicholas Ave

department: Institute for Cancer Genetics

institute: Columbia University

summary: FYN-TRAF3IP2 expression in hematopoietic progenitors induces T cell transformation in mice and cooperates with loss of the Tet2 tumor suppressor in PTCL development. To investigate the oncogenic activity of FYN-TRAF3IP2, we analyzed the lymphomagenic effects of expressing this gene fusion alone and in cooperation with loss of the Tet2 tumor suppressor gene in vivo. In these experiments, we first infected hematopoietic progenitors from CD4-specific tamoxifen-inducible Cre Tet2 conditional knockout mice (CD4 Cre-ERT2, Tet2fl/fl) with bicistronic retroviruses expressing wild type TRAF3IP2 and GFP, FYN-TRAF3IP2 and GFP, or GFP alone and injected these intravenously into isogenic recipients. Transplanted mice were then treated with vehicle only, to test the oncogenic effects of TRAF3IP2, FYN-TRAF3IP2 or GFP expression; or with tamoxifen, to evaluate the effects of TRAF3IP2, FYN-TRAF3IP2 or GFP expression in concert with genetic loss of Tet2 in CD4 T cells. In this setting, all animals transplanted with GFP-expressing progenitors remained lymphoma free at the end of follow up and one animal transplanted with wild type TRAF3IP2 GFP expressing cells developed a CD8+ T cell lymphoma. In contrast, 4/10 (40%) vehicle treated mice transplanted with FYN-TRAF3IP2-expressing cells and 5/10 (50%) tamoxifen treated mice transplanted with FYN-TRAF3IP2-expressing cells developed clonal CD4-restricted mature T cell lymphomas. To further analyze the lymphomagenic effect of the fusion gene, we performed transcriptomic profiling of FYN-TRAF3IP2-induced mouse CD4+ GFP+ PTCL tumor lymphocytes compared with normal mouse CD4+ T cells by RNAseq.

overall_design: We performed RNAseq on nine FYN-TRAF3IP2-induced mouse tumor samples of CD4+ GFP+ FACS-sorted lymphoma cells (six independent primary tumors and three independent secondary transplanted tumors) and three independent wild type normal mouse CD4+ T cell samples using Illumina HiSeq 4000. (average depth of 30 million, 150 base pair, paired-end reads per sample)

first: Christine

last: Kim

first: S

first: Adolfo

last: Ferrando


title: Transgenic Plasmodium falciparum parasites line 3D7/DDGFP-PfAP2-HC: transcriptional differences between 3D7/DDGFP-PfAP2-HC control parasites (cultured in presence of Shield-1; ON) vs PfAP2-HC-depleted parasites (cultured in absence of Shield-1; OFF)

status: Public on Oct 06 2020

city: Basel

first: Till

middle: Steffen

last: Voss

address: Socinstrasse 59

department: Medical Parasitology and Infection Biology

institute: Swiss Tropical and Public Health Institute

summary: Transcriptional profiling of transgenic P. falciparum asexual blood stage parasites of the transgenic strain 3D7/DDGFP-PfAP2-HC at five time points during intra-erythrocytic parasite development. The DD (FKBP destabilisation domain) allows for the conditional expression of fusion proteins: DD fusion proteins are rapidly degraded or stably expressed in absence or presence of the stabilising ligand Shield-1, respectively (Banaszynski LA, Chen LC, Maynard-Smith LA, Ooi AG, Wandless TJ. A rapid, reversible, and tunable method to regulate protein function in living cells using synthetic small molecules.Cell. 2006 Sep 8;126(5):995-1004). The goal of this experiment was to identify genes differentially expressed in DDGFP-PfAP2-HC-expressing compared to DDGFP-PfAP2-HC-depleted parasites during the intra-erythtrocytic cell cycle.

overall_design: P. falciparum 3D7/DDGFP-PfAP2-HC transgenic parasites expressing a GFP-DD-tagged version of the ApiAP2 factor PfAP2-HC (PF3D7_1456000) were grown in the presence of 675nM Shield-1 (DDGFP-PfAP2-HC “ON”) or in the absence of Shield-1 (DDGFP-PfAP2-HC “OFF”). RNA extracted from these samples at five consecutive time points during the intra-erythrocytic cell cycle was processed for microarray analysis.

first: Eilidh

last: Carrington

first: Roel

middle: H

last: Cooijmans

first: Dominique

last: Keller

first: Christa

last: Toenhake

first: Richard

last: Bartfai


title: Transcriptome analysis of Arabidopsis thaliana and Vibrio vulnificus MO6-24/O by RNA-seq

city: Cheongju-si

first: Yong-Soon

last: Park

state: Chungcheongbuk-do

address: Cheongju

department: Biotechnology Research  Institute

institute: Chungbuk National University

summary: In order to analyze the transcripts of Arabidopsis thaliana (Col-0) and Vibrio vulnificus MO6-24/O simultaneously, Vibrio vulnificus MO6-24/O was infiltrated onto Arabidopsis leaves and then leaves were harvested at 0, 3, 6, 12, 24 and 48 h post-infiltration. A total of 31, 128, 303, 219 and 130 differentially expressed genes (DEGs) of Vibrio were up- and down-regulated at 3, 6, 12, 24 and 48 h post-infiltration (hpi). Meanwhile, differentially expressed genes (DEGs) were monitored at 3, 6, 12, 24 and 48 h post-infiltration. A total of 2,097, 1,839, 1,220, 1,170 and 1,383 genes were characterized at each time points in Arabidopsis. Our data clearly indicate that total transcripts of the marine bacterial pathogen V. vulnificus MO6-24/O are detected and analyzed in plant Arabidopsis and two organisms were inter-communicated at the same time under favorable conditions.

overall_design: Vibrio vulnificus MO6-24/O was infiltrated in Arabidiosps (Col-0) and then samples were harvested at 0, 3, 6, 12, 24 and 48 h. Negative control treated plants were also harvested at the same time points. Two samples were replicated at each time point. Total RNA was isolated by RNeasy Mini Kit (Qiagen, CA, USA), according to the manufacturer's protocol. The quality and integrity of the RNA were confirmed by agarose gel electrophoresis and ethidium bromide staining, followed by visual examination under ultraviolet light. Sequencing library was prepared using TruSeq Stranded Total RNA LT (with Ribo-Zoro-Plant) Set A (Illumina, CA, USA) according to manufacturer’s protocols. Sequencing was performed in paired end reads (2x100 bp) using Hiseq-2000 (Illumina).

first: Jong-Seok

first: Choong-Min

last: Ryu


title: Expression data from rat lens epithelial cells after extracapsular lens extraction (ECLE)

status: Public on Sep 30 2020

city: Kahoku

first: Eri

last: Kubo

state: Ishikawa

address: 1-1 Daigaku, Uchinada

department: Ophthalmology

institute: Kanazawa Medical University

summary:
The epithelial-mesenchymal transition (EMT) of lens epithelial cells (LECs) has been proposed as a major cause of posterior capsule opacification (PCO) after cataract surgery. Molecular mechanism of PCO progression is still unclear.
Using a microarray-based approach, herein we studied the changes in gene expression pattern during rat PCO formation in vivo as a model.

overall_design: Rat LECs were collected at 0, 1 and 2 weeks (W) after ECLE  for RNA extraction and hybridization on Affymetrix microarrays.

first: Shinsuke

last: Shibata


title: ATAC-sequencing data for 923 enhancer deletion newborn mouse keratinocytes

address: 660 South Euclid

institute: Washington University in St Louis

summary: We examined the ATAC-seq chromatin profiles of epidermis from newborn CRISPR-edited mice that have the 923 enhancer located within the Epidermial Differentiation Complex deleted. We examined two independent deletion lines, 923del and 923large,  homozygous mutants, as well as wild type mice. We find overall there are not many differences in chromatin accessibility upon the loss of the 923 enhancer, though there are a few peaks of less accessible DNA within the EDC and more accessible DNA just outside the EDC.

overall_design: Examination of accessible chromatin profiles by ATAC-seq in two  923 enhancer deletion lines and WT  newborn mouse epidermis.

first: Mary

last: Mathyer


title: Expression data of dermal cells with and without small molecules treatment

status: Public on Sep 29 2020

city: Berkeley

first: Junren

last: Sia

state: California

address: 156 Stanley Hall

department: Bioengineering

institute: University of California, Berkeley

summary: Traumatic injury often results in muscle loss and impairment, which is worsened under aged and diseased conditions. Activation of resident stem cells or transplantation of myogenic stem cells can promote muscle regeneration. However, major challenges remain in harvesting sufficient autologous myogenic stem cells and expanding such cells efficiently for muscle regeneration therapies. Here, we identified a chemical cocktail that selectively induced a robust expansion of myogenic stem cells from readily-obtainable dermal cells and from muscle stromal cells. By differential plating and lineage tracing, we showed that Pax7+ cells were the major source for chemical-induced myogenic stem cells (CiMCs). We further performed single-cell RNA sequencing (scRNA-seq) analysis to characterize the transcriptomic profile of CiMCs and demonstrate a specific expansion of myogenic cells from heterogeneous dermal cell population. Upon transplantation into the injured muscle, CiMCs were efficiently engrafted and improved functional muscle regeneration in both adult and aged mice. In addition, CiMC transplantation rescued muscle function in mice with Duchenne muscular dystrophy (DMD). Furthermore, an in situ therapeutic modality using this cocktail was developed by loading the chemical cocktail into injectable nanoparticles, which enabled a sustained release of the cocktail in injured muscle and a local expansion of resident satellite cells for muscle regeneration in adult and aged mice. These findings will lead to the development of novel in vitro and in situ stem cell therapies for effective skeletal muscle repair.

overall_design: Neonatal dermal cells were cultured in medium with and without 20 μM Forskolin + 20 μM Repsox for 2 days before RNA is collected and prepared for microarray analysis.

first: Song

last: Li