title: Exposure of Eucalyptus grandis and E. globulus to varied temperature and elevated CO2 concentration alone or in combination have profound effect in the expression of genes controlling cell wall formation and remodeling

status: Public on Feb 02 2021

city: Campinas

first: Paulo

last: Mazzafera

state: Sao Paulo

address: Rua Monteiro Lobato

department: Plant Physiology

country: Brazil

institute: State University of Campinas

summary: Several reports have investigated the effects of future climate conditions on photosynthesis and biomass production in woody plants but none combining temperature and atmospheric CO2 concentration. We used grow chambers to expose Eucalyptus grandis and E. globulus seedlings to three temperature regimes (10-12°C - low temperature, 20-22°C - control temperature and  33-35°C - high temperature) combined with two CO2 concentration (390 ppm – normal and 700 ppm - high). After 35 days the stems were used for transcriptome (RNAseq) analysis aiming to understand  changes in the expression of genes related with biosynthesis and remodeling of the cell wall, as well as genes related with carbon partitioning and hormonal regulation. E. globulus, a species adapted to low temperature, was more responsive to the treatments compared to E. grandis. The most relevant biological processes affected by the treatments were related to stress, secondary metabolism, hormonal response, and signaling. We also analyzed in depth all cell wall biopolymer synthetic genes and their regulation by innumerous transcriptional factors. There was an upregulation of ethylene and auxin biosynthetic genes in both species, but it was stronger under high and low temperature in E.globulus. Temperature was more effective to change cell wall biosynthetic genes than CO2 and CO2 stimulated lignin biosynthetic pathway. We also analyzed lignin composition and structure. There was an increase in F5H and COMT expression and the S/G ratio also increased under high CO2 in E.globulus. Thus, the species responded differently to treatments and stem composition and development was highly affected by temperature and CO2 concentration.

sample_organism:
Eucalyptus globulus
Eucalyptus grandis

platform_organism:
Eucalyptus globulus
Eucalyptus grandis

overall_design: Total stem mRNA of 35-day-old  eucalyptus plants exposed to three temperatures (10°C - low, 20°C - controle and 30°C - high) and two concentrations of carbon dioxide (400 ppm - low and 700 ppm - high)

first: Daniela

last: Feltrim


title: Blood gene expression profiles following influenza vaccination in pregnant women

status: Public on Feb 11 2021

city: Memphis

first: Zhaohui

last: Xu

state: TN

address: 262 Danny Thomas Place

department: Infectious Diseases

country: USA

institute: St. Jude Children's Research Hospital

summary: Influenza immunization during pregnancy provides protection to the mother and the infant. Studies in adults and children with inactivated influenza vaccine (IIV) have identified changes in immune gene expression that correlated with antibody responses. We studied changes in transcriptional profiles induced by IIV in pregnant women and to identify correlates of antibody responses.

overall_design: Whole blood was collected in Tempus tubes from 44 pregnant women on day 0 (pre-vaccination), day 1 and day 7 post-vaccination. RNA was extracted and globin reduced. Labeled cRNA was hybridized to Illumina Human HT-12 Beadchips.

first: Raquel

last: Giacomelli Cao

first: Lisa

last: Christian

last: Jaramillo

first: Bennett

last: Smith

first: Erik

middle: A

last: Karlsson

first: Stacey

last: Schultz-Cherry

first: Asuncion

last: Mejias

first: Octavio

last: Ramilo


title: Enhancers predominantly control transcription initiation [ATAC-seq]

status: Public on Feb 03 2021

city: Oxford

first: Martin

middle: Stephen Charles

last: Larke

state: Oxfordshire

address: Headley Way

department: MRC Weatherall Institute of Molecular Medicine

country: United Kingdom

institute: University of Oxford

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Gene transcription occurs via a cycle of linked events including initiation, promoter proximal pausing and elongation of RNA polymerase II (Pol 2).  A key question is how do transcriptional enhancers influence these events to control gene expression? Here we have used a new approach to quantify transcriptional initiation and pausing in-vivo, while simultaneously identifying transcription start sites (TSSs) and pause-sites (TPSs). When analysed in parallel with nascent RNA-seq, these data show that differential gene expression is achieved predominantly via changes in transcription initiation rather than Pol 2 pausing or elongation. Using genetically engineered mouse models deleted for specific enhancers we show that these elements control gene expression via Pol 2 recruitment and/or initiation rather than via promoter proximal pause release. Using genome-wide analysis we show that enhancers, in general, control gene expression at the stage of Pol 2 recruitment and initiation rather than via pausing.

overall_design:
To characterise accessible chromatin genome wide we performed ATAC-seq on primary mouse erythroid cells from Wildtpye or R1R2, HS2HS3 lines

HS2HS3 genotype: HS2 and HS3 enhancers of beta globin are homozygously deleted in combination
R1R2 genotype: A compound homozygous deletion of the R1 and R2 (a.k.a -26 and -31) transcriptional enhancers of the alpha globin gene (first reported in Hay et al 2016, Nature Genetics).

first: Jacqueline

last: Sharpe

last: Sloane-Stanley

first: Sue

last: Butler

first: Jim

middle: R

last: Hughes

first: Douglas

last: Higgs


title: alpha/beta/gamma/delta T cells play a vital role in fetal human skin development and immunity

city: Vienna

first: Christoph

last: Bock

address: Lazarettgasse 14

country: Austria

institute: CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences

type: Expression profiling by high throughput sequencing

summary: T cells in adult human skin play a pivotal role in the defense against pathogens and tumors. While T cells can also be found in fetal human skin, little is known about their phenotype and function. Using single cell analyses, here we identify an unconventional naive T cell population expressing both alpha/beta and gamma/delta T cell receptors (TCRs) that was markedly enriched within early fetal skin and intestine but was untraceable in other fetal organs and peripheral blood. TCR sequencing data revealed that double-positive (DP) T cells displayed marginal or no overlap of CDR3 sequences with single-positive alpha/beta T cells. Gene signature, cytokine profile and in silico receptor-ligand interaction studies indicated that DP T cells contribute to skin development early in life. Finally, DP T cells were phosphoantigen-responsive suggesting that these cells participate in the protection of the fetus against pathogens in case of intrauterine infections. Together, our analyses unveil a unique and hitherto unrecognized cutaneous T cell type within the native skin microenvironment and point to fundamental differences in the adaptive immune surveillance of developing and adult human skin.

overall_design:
Three samples of human embryonic skin.

**Submitters declare that they can not provide raw sequencing data, as the data contain identifiable information.

first: R

last: Reitermaier

first: T

last: Krausgruber

first: N

last: Fortelny

last: Ayub

first: PA

last: Vieyra-Garcia

first: P

last: Kienzl

last: Wolf

first: A

last: Scharrer

first: C

last: Fiala

first: M

last: Kölz

last: Hiess

last: Vierhapper

last: Schuster

last: Spittler

last: Worda

first: WP

last: Weninger

first: W

last: Eppel

last: Elbe-Bürger


title: Enhancers predominantly control transcription initiation [RNA-seq]

last: Larke

overall_design: To characterise intronic RNA expression levels genome wide we performed RNA-seq on poly adenylated, globin RNA and rRNA depleted RNA from primary mouse erythroid cells.

last: Marieke Oudelaar

first: Robert

last: Beagrie

last: Butler

last: Sharpe

last: Sloane-Stanley

last: Hughes

last: Higgs


title: BisMapR: a strand-specific, nuclease-based method for genome-wide R-loop detection

status: Public on Jan 29 2021

city: Philadelphia

first: Phillip

last: Wulfridge

state: Pennsylvania

address: 3601 Spruce St

institute: The Wistar Institute

summary: We have developed an antibody-independent and strand-specific R-loop detection strategy, BisMapR, that combines nuclease-based R-loop isolation with non-denaturing bisulfite chemistry to produce high-resolution, genome-wide R-loop sequencing maps.

overall_design: BisMapR and MapR was performed on E14 mouse embryonic stem cells, with two replicates per method. As controls, BisMapR was performed on RNase A-treated R-loop samples, MapR was performed with a nonspecific MNase protein, and MapR was performed with second-strand synthesis but without bisulfite conversion.

first: Kavitha

last: Sarma


title: Gene expression patterns of a novel breast cancer cell line derived from a bone metastatic lesion of a breast cancer patient

status: Public on Jan 31 2021

city: Herston

first: Georgia

last: Trench

state: QLD

address: 300 Herston Rd

department: Cancer Genetics

country: Australia

institute: QIMR Berghofer Medical Research Institute

summary: As organ-specific models of breast cancer bone metastasis do not exist, we established a novel breast cancer line was established from the ER-/PR-/HER2- bone metastasis from a breast cancer patient.  This cell line was characterized and compared to the primary tumour and/or normal mammary epithelial cells with regards to phenotypes (i.e. marker expression), gene expression and copy number variation.

overall_design: Total RNA was isolated from the P7731 cell line (3 replicates) and compared to that isolated from the primary tumour (2 replicates) or normal mammary epithelial cells (3 replicates)

first: Julie

last: Johnson

first: Darrell

middle: C

last: Bessette

first: Jodi

middle: M

last: Saunus

first: Chanel

middle: E

last: Smart

first: Sarah

last: Song

first: Rebecca

last: Johnston

first: Fares

last: Al-Ejeh

first: Sibylle

last: Cocciardi

first: Cameron

middle: N

last: Johnstone

first: Peter

last: MacCallum

first: Kum

middle: K

last: Khanna

first: Sunil

last: Lakhani

last: Chenevix-Trench

middle: T

last: Simpson

first: Nic

last: Waddell


title: Enhancers predominantly control transcription initiation

first: Martin

last: Larke

type: Non-coding RNA profiling by high throughput sequencing

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

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Refer to individual Series


title: Enhancers predominantly control transcription initiation [scaRNA-seq]

overall_design:
To characterise promoter proximal transciption levels genome wide we performed scaRNA-seq on Total RNA from primary mouse erythroid cells and the human K562 cell line.

HS2HS3 genotype: HS2 and HS3 enhancers of beta globin are homozygously deleted in combination
R1R2 genotype: A compound homozygous deletion of the R1 and R2 (a.k.a -26 and -31) transcriptional enhancers of the alpha globin gene (first reported in Hay et al 2016, Nature Genetics).

first: Bryony

last: Crompton

first: Sue

last: Butler

first: Jacqueline

last: Sharpe

last: Sloane-Stanley

first: Jim

last: Hughes

first: Douglas

last: Higgs


title: Enhancers predominantly control transcription initiation [ChIP-seq]

overall_design:
To characterise Pol2 density genome wide we performed ChIP-seq on primary mouse erythroid cells. A 10% input sample was taken from a wildtype sample prior to immunoprecipitation as a control and 5ng of this material was subjected to library prep alongside the immunoprecipitated samples.

HS2HS3 genotype: HS2 and HS3 enhancers of beta globin are homozygously deleted in combination
R1R2 genotype: A compound homozygous deletion of the R1 and R2 (a.k.a -26 and -31) transcriptional enhancers of the alpha globin gene (first reported in Hay et al 2016, Nature Genetics).


title: ChIP-seq analysis of H3K9me3 signal from T-ALL cell line (Jurkat) upon fast CXCL12 stimulation

city: Madrid

first: Javier

last: Redondo-Muñoz

state: Madrid

address: Ramiro de Maeztu,9

department: Molecular Biomedicine

country: Spain

institute: CIB-CSIC

summary: T-acute lymphoblastic leukaemia (T-ALL) is an aggressive haematological malignancy, which comprises the accumulation of malignant T-cell precursors. Despite current therapies, up to 20% of children with failure to respond to treatments and relapse. Our understanding of T-ALL infiltration and how leukaemia cells-microenvironment interactions play a role in the clinical outcome is still vague. In the present study, we showed a novel function of the chemokine CXCL12, which induced fast epigenetic changes within minutes in cell lines and primary T-ALL cells. Our results identified that CXCL12-mediated H3K9 methylation impacted on the global chromatin configuration and the nuclear mechanics of T-ALL cells. We characterised changes in the transcriptional profile of T-ALL cells associated with rapid CXCL12 stimulation. We demonstrated that cytoskeletal changes and protein kinase-C (PKC) activity were the molecular mechanisms by which CXCL12 promoted H3K9 methylation in T-ALL cells. Furthermore, targeting H3K9 methyltransferases reduced the migration and the nuclear deformability in both cell line and primary human ALL cells. Together, our data indicate a novel funcion of H3K9 methylation induced by CXCL12 in T-ALL cells, that reveals the significance of nuclear changes to mechanobiology and function of leukemia cells and emerges as a promising pharmacological target against T-ALL infiltration and dissemination.

overall_design: Jurkat cells were culture in growth medium without  FBS (starving condition) for 90 min, then cells were stimulated or not with CXCL12 (100ng/ml) for 15 min. Cells were collected, and chromatin crosslinked with PFA. Cell lysates were anaysed for H3K9me3 ChIP-seq according to Active Motif protocol.

first: Elena

last: Madrazo

first: Raquel

last: González-Novo

first: Mario

last: García de Lacoba

first: Lorea

last: Abad

first: África

last: González-Murillo

first: Manuel

last: Ramírez


title: Expression data changes induced by CXCL12 in T-ALL cells

summary: Our results identified that CXCL12-mediated H3K9 methylation impacted on the global chromatin in the transcriptional profile of T-ALL cells.

overall_design: Jurkat  cells  (T-ALL cell line) were used for RNA extraction and hybridization on Affymetrix microarrays. Untreated or CXCL12 (100ng/ml, 15 min) stimulated Jurkat cells were lised and mRNA was extracted to compare expressions profiles. We used three replicates of each sample.


title: Spatial Transcriptomics to define transcriptional patterns of zonation and structural components in the liver

status: Public on Jan 20 2021

city: Stockholm

first: Franziska

middle: Frederike Annalena

last: Hildebrandt

state: Stockholms Lan

address: Svante Arrhenius Väg 20c, MBW, Stockholms University

department: Department of Molecular Biosciences, the Wenner-Gren Institute

country: Sweden

institute: Stockholm University

summary: Reconstruction of heterogeneity through single-cell transcriptional profiling has greatly advanced our understanding of the spatial liver transcriptome in recent years. However, global transcriptional differences across lobular units remain elusive in physical space. Here, we implement Spatial Transcriptomics to perform transcriptomic analysis across sectioned liver tissue. We confirm that the heterogeneity in this complex tissue is predominantly determined by lobular zonation. By introducing novel computational approaches, we enable transcriptional gradient measurements between tissue structures, including several lobules in a variety of orientations. Further, our data suggests the presence of previously transcriptionally uncharacterized structures within liver tissue, contributing to the overall spatial heterogeneity of the organ. This study demonstrates how comprehensive spatial transcriptomic technologies can be used to delineate extensive spatial gene expression patterns in the liver, indicating its future impact for studies of liver function, development and regeneration as well as its potential in pre-clinical and clinical pathology. Additional data to reproduce the data presented in this study with instrctions can be found at https://github.com/almaan/ST-mLiver and https://zenodo.org/record/4399655

overall_design: Spatial Transcriptomics analysis of 8 wildtype murine liver sections from 2 lobes

first: Johan

last: Ankarklev


title: High throughput error correction using dual nucleotide dimer blocks allows direct single-cell nanopore transcriptome sequencing

first: Adam

last: Cribbs

address: Windmill Road

department: NDORMS

summary: Droplet-based single-cell sequencing techniques have provided unprecedented insight into cellular heterogeneities within tissues. However, these approaches only allow for the measurement of the distal parts of a transcript following short-read sequencing. Therefore, splicing and sequence diversity information is lost for the majority of the transcript. The application of long-read Nanopore sequencing to droplet-based methods is challenging because of the low base-calling accuracy currently associated with Nanopore sequencing. Although several approaches that use additional short-read sequencing to error-correct the barcode and UMI sequences have been developed, these techniques are limited by the requirement to sequence a library using both short- and long-read sequencing. Here we introduce a novel approach termed single-cell Barcode UMI Correction sequencing (scBUC-seq) to efficiently error-correct barcode and UMI oligonucleotide sequences synthesized by using blocks of dimeric nucleotides. The method can be applied to correct both short-read and long-read sequencing, thereby allowing users to recover more reads per cell that permits direct single-cell Nanopore sequencing for the first time. We illustrate our method by using species-mixing experiments to evaluate barcode assignment accuracy and multiple myeloma cell lines to evaluate differential isoform usage and Ewing’s sarcoma cells to demonstrate Ig fusion transcript analysis.

overall_design: Mouse (3T3) and human (HEK293T) cells were mixed at a ratio of 1:1 and individual cells were encapsulated using a droplet based single-cell sequencing encapsulator device (DolomiteBio Nadia system). Libraries were then sequenced using  a MinION Oxford nanopore flow cell (FLO-MIN106D) or PrimethION flow cell (FLO-PRO002).

middle: P


title: Genome-wide perturbations of Alu expression and Alu-associated post-transcriptional regulations find a uniqueness in oligodendroglioma

status: Public on Jan 27 2021

city: Seoul

first: Chul-Kee

last: Park

address: 101 Daehak-ro, Jongno-gu

department: Department of Neurosurgery

country: South Korea

institute: Seoul National University College of Medicine

summary: Alu is a primate-specific repeat element in the human genome and has been increasingly appreciated as a regulatory element in many biological processes. But the role of Alu has not been studied comprehensively in brain tumor because an evolutionary perspective has been the subject of little research in brain tumor. We aim to investigate the relevance of Alu to the gliomagenesis.

overall_design: Using a total of 41 pairs of neurotypicial brain tissue samples and samples of diverse gliomas, we performed strand-specific RNA-seq and analyzed two Alu-associated post-transcriptional regulations, A-to-I editing and circular RNAs, and Alu expression in a genome-wide way.

first: Taeyoung

last: Hwang


title: Reiterative infusions of MSCs improve pediatric Osteogenesis Imperfecta eliciting a pro-osteogenic paracrine response: TERCELOI clinical trial

status: Public on Jan 22 2021

city: Muenster

first: Marcos

middle: J.

last: Araúzo-Bravo

address: Rogentstrasse

department: Cell and Developmental Biology

country: Germany

institute: Max Planck Institute for Molecular Biomedicine

summary:
Osteogenesis Imperfecta (OI) is a genetic,
rare disease characterized by bone fragility, with a wide range in the severity of clinical manifestations.
The majority of the cases are due to mutations in COL1A1 or COL1A2 genes which code for the type I collagen molecule.
Mesenchymal stem cells (MSCs), as the progenitors of the osteoblasts, the main type I collagen secreting cell type in the bone,
have been proposed and tested as an innovative therapy for OI with promising but transient outcomes.
In this study, we performed a phase I clinical trial based on reiterative infusions of histocompatible MSCs
in two pediatric patients affected by severe and moderate OI.
The aim of this study was to assess the safety and effectiveness of this cell therapy in non-immunosuppressed OI patients.
The host response to MSCs was also evaluated by collecting the sera from OI patients before, along and after the cell therapy.
The results of this clinical trial are twofold:
the sequential administration of MSCs is safe and improve the bone parameters and quality of life of OI patients.
Moreover, MSCs therapy elicits a pro-osteogenic paracrine response in patients, especially noticeable in that affected by severe OI.
These results indicate the feasibility and potential of reiterative MSCs infusion
for OI and highlight the paracrine response shown by OI patients as a consequence of MSCs treatment.
Study design and patients:
The Mesenchymal Stem Cell Therapy for the Treatment of Osteogenesis Imperfecta (TERCELOI) study
(www.clinical trials.gov: # NCT02172885, EudraCT number: 2012-002553-38) is an independent multi-center Phase I clinical trial to evaluate the feasibility,
safety and potential efficacy of infused sibling HLA-matched MSCs in non-immunosuppressed children with OI.
PBMCs isolation and serum collection:
Venous blood samples were collected before the cell therapy (basal serum), along the cell therapy (1 week, 1 month and 4 months after each MSCs infusion)
and during the follow-up period (1 and 2 years after the fifth and last MSCs infusion).
MLR assay:
PBMCs isolated from patients were resuspended in PBS+FBS (5%) and stained with carboxyfluoroscein succinate-ester (CFSE) (Molecular Probes, USA).
Bone mineral density:
Bone mineral density (BMD) at the lumbar spine (LS) from L1 to L4, was measured by dual energy x-ray absorptiometry (DXA)
on a whole-body scanner within a pediatric platform (Hologic QDR densitometer).
Raw measurements were converted to Z-scores for analysis using reference standards for age and pubertal status.
OI-MSCs isolation and characterization and culture:
OI-MSCs were derived from discarded and donated bone fragments of OI pediatric patients undergoing corrective surgery.
All patients suffered Type III-IV OI, with mutations in either COL1A1 or COL1A2 genes.
The donation was approved by the Basque Clinical Research Ethics Committee. Briefly, bone chips were mechanically flushed with PBS
and then cut into small pieces to extrude cells without the use of enzymes.
The cut bone pieces were let undisturbed during 14 days until OI-MSCs migrated to cell culture plate.
OI-MSCs were assessed for the expression of CD105, CD90 and CD73 and the absence of CD14, CD34, CD45, CD19 and HLA-DR.
MSCs were also tested for their potential to differentiate to osteoblasts and adipocytes using specific cell culture media.
OI-MSCs were cultured under standard growth medium (DMEM low glucose with glutamax (Gibco),
penicillin/streptomycin (Gibco) and fetal bovine serum at 10% (Sigma-Aldrich, USA).
Osteogenesis induction medium (OIM) was composed of standard medium plus ascorbic acid 0.2 mM, beta-glycerophosphate 10 mM, and dexamethasone 10 nM
(all from Sigma-Aldrich). When specified, FBS was replaced by P01 and P02 serum samples at 2.5%.
RNAseq and Q-PCR validation:
OI-MSCs from 10 OI pediatric patients were seeded in 96-well plates (1 000 cells/well) and the following day cultured in the presence of OIM.
Next, total RNA was isolated with the AllPrep kit (QIAGEN, USA). cDNA library (TrueSeq stranded Total cDNA library, Illumina, USA) and sequencing at
HiSeq 4000 (PE100nt, 50 million reads/sample).

overall_design:
RNAseq and Q-PCR validation:
OI-MSCs from 10 OI pediatric patients were seeded in 96-well plates (1 000 cells/well) and the following day cultured in the presence of OIM.
Next, total RNA was isolated with the AllPrep  kit (QIAGEN, USA). cDNA library (TrueSeq stranded Total cDNA library, Illumina, USA) and sequencing at
HiSeq 4000 (PE100nt, 50 million reads/sample).
TNAP activity measurement:
OI-MSCs from 8 pediatric patients were seeded in 96-well plates (1 000 cells/well) and the following day induced to differentiate in the presence of OIM
and serum from P01 or P02 patients (2.5%). After 4 days of differentiation,
TNAP activity was determined by adding to each well a chromogenic substrate of alkaline phosphatase, 1-stepTM PNPP substrate solution
(Thermo Fisher Scientific, USA) and absorbance was measured at 405 nm. To normalize TNAP activity,
cell number was determined by Cell Counting Kit-8 (CCK-8, Bimake, USA) and absorbance at 415 nm measured in a parallel plate.
As negative controls, OI-MSCs cultured under non differentiation conditions were used.
ARS staining:
OI-MSCs were seeded in triplicates in 12-well plates (15 000 cells/well) and the following day induced to differentiate in the presence of OIM
supplemented with serum samples obtained from P01 and P02 at 2.5%. Medium was changed twice per week.
After 14 days of mineralization was assessed by staining the extracellular calcium deposits.
For this, cells were fixed with paraformaldehyde (4%) and stained with ARS (Thermo Fisher Scientific) (2%) for 45 minutes.
Then, to quantify the mineralization, the wells were washed twice and the dye was extracted with cetylpyridinium chloride (Sigma-Aldrich) at 10%
in sodium phosphate buffer (Sigma-Aldrich). Then absorbance at 560 nm was measured in a plate reader.

first: Arantza

last: Infante

first: Blanca

last: Gener

first: Miguel

last: Vazquez

first: Nerea

last: Olivares

last: Arrieta

first: Gema

last: Grau

first: Isabel

last: Llano

first: Luis

last: Madero

first: Ana

last: Bueno

first: Belén

last: Sagastizabal

first: Daniela

last: Gerovska

middle: J

last: Arauzo-Bravo

first: Itziar

last: Astigarraga

first: Clara

middle: I

last: Rodríguez


title: Gene expression profiling in periodontal ligament cells derived from Down syndrome

status: Public on Jan 17 2021

city: Tokyo

first: Asakawa

last: Takeyoshi

state: Ota-ku

address: 2-1-1 Kitasenzoku

department: Pediatric

country: Japan

institute: Showa university

summary: We performed the microarray analysis for gene expression profiling in periodontal ligament cells derived from Down syndrome.

overall_design: Human periodontal ligament cells derived from control (sample 1; PDLGO) and Down syndrome (sample 2; PDLDS) .

first: Takeyoshi

last: Asakawa

first: Atsushi

last: Yamada

first: Yoichi

last: Miyamoto

first: Kentaro

last: Yoshimura

first: Kiyohito

last: Sasa

first: Misumi

last: Kugino

first: Ryutaro

last: Kamijo


title: EMSY inhibits homologous recombination repair and the interferon response promoting lung cancer immune evasion

status: Public on Jan 15 2021

city: New York

first: Genome Technology Center

last: (GTC)

address: 550 First Avenue

institute: NYU Langone Health

summary: Purpose: The goal of this study is to analyze the transcriptional pathways regulated by Emsy and Keap1 in subcutaneous mouse lung adenocarcinoma tumors.  Methods: Keap1 wild type (KP) or mutant (KPK) cells expressing inducible shEmsy or shCTRL were implanted subcutaneously into the flanks of 6- to 8-week-old female syngeneic mice. Once tumors were measurable mice were treated with doxycycline to induce the shRNA expression. Knock down efficiency has been evaluated by western blot (using specific antibody for Emsy) and qPCR (using specific oligos for Emsy).   Results: The transcriptomic analysis helps us to support our finding that Keap1-null tumors are characterized by suppression of the IFN response resulting from the stabilization of Emsy.

overall_design: All 12 samples generated by deep sequencing

first: Michele

last: Pagano

first: Antonio

last: Marzio


title: Transcriptome changes of Hela cells with blocking classical and alternative non-homologous end-joining (NHEJ) pathways

status: Public on Dec 31 2020

city: Las Vegas

first: Martin

last: Schiller

state: NV

address: 4505 S Maryland Pkwy

department: Nevada Institute of Personalized Medicine

institute: University of Nevada, Las Vegas

summary:
Purpose: The recombinant endonucleases introduce double-strand breaks (DSBs) in genomic DNA at targeted sites and non-homologous end-joining (NHEJ) pathways repair the breaks introducing indels into the genome. To investigate whether there is another NHEJ pathway or if A-NHEJ and C-NHEJ are redundant, we examined NHEJ edits in HeLa cells.
Methods: To test among these possibilities, we engineered a NHEJ reporter system using targeted TALEN induced double strand DNA breaks. C-NHEJ was blocked in XRCC4(-/-) cells and A-NHEJ was blocked pharmacologically with mirin. To assess how NHEJ was maintained, differentially expressed genes were determined by RNA-Seq.
Results: When both C-NHEJ and A-NHEJ are blocked, edits were still observed. No changes in expression of DNA repair genes was observed. However, when both NHEJ pathways were blocked, genes from several pathways including DNA damage response, hypoxic response, chromatin packaging, p53 response and two genes that stimulate homologous recombination were differentially expressed.
Conclusions: NHEJ is maintained when both NHEJ pathways are blocked through a transcriptional response that must compensate for the inhibited steps in both pathways.

overall_design: Overall design: Total eight samples separated in two experiments (biological duplicates) were sequenced, for profiling transcriptomes of Hela cells under four conditions – 1) Hela cells, 2) Hela cells with only blocking C-NHEJ pathway by XRCC4 knockout, 3) Hela cells with only blocking A-NHEJ pathway by mirin treatment, and 4) Hela cells with blocking both C-NHEJ and A-NHEJ pathways.

first: Ronald

last: Benjamin


title: Chromatin status and transcriptomics of HEK293, U2OS, HepG2 and K562

city: Charlestown

first: Keith

last: Joung

state: MA

institute: MGH

summary: RNA-seq, H3K27Ac ChIP-seq, and ATAC-seq data of U2OS, HEK293, HepG2, and K562 cell lines

overall_design: Two biological replicates for each samples were prepared.  Cells were native cells with no treatment.

first: Y

last: Tak


title: Transcriptomic profiling of S. cerevisiae and S. boulardii during growth in host-related conditions

status: Public on Jan 12 2021

city: Lisbon

first: Miguel

middle: Cacho

last: Teixeira

address: Avenida Rovisco Pais 1

department: Institute for Bioengineering and Biosciences

country: Portugal

institute: Instituto Superior Tecnico

summary: In this study, the hypothesis that the different phenotypes exhibited by S. cerevisiae and S. boulardii lay on differential gene expression, based on promoter sequencing variability, is approached. To test this possibility, S. boulardii and S. cerevisiae were grown in intestinal like medium (ILM), optimized to enable the growth of both strains, and their transcriptome-wide expression patterns were evaluated through RNA-sequencing. mRNA profiles were generated by deep sequencing, in triplicate, using Illumina NextSeq. The sequence reads that passed quality filters were analyzed with TopHat followed by HTSeq. RNA-seq data allowed to identify genes whose expression is differentially regulated in both organisms with a log2fold change ≥1 and p value <0.01. The RNA-Seq data was also used to refine the current S. boulardii annotation.

overall_design: Yeast mRNA profiles from cells grown during 6h in ILM were generated by RNA-Seq, in triplicate using Illumina NextSeq

first: Pedro

last: Pais

first: Oliveira

last: Jorge

first: Almeida

last: Vanda

first: Yilmaz

last: Melike

last: Monteir


title: mRNA expression data in epididymal fat tissues derived from mir221/222 adipose tissue specific knockout (AdipoKO) and mir221/222 flox/y mice

status: Public on Dec 29 2020

city: Okayama

first: Jun

last: Wada

address: 2-5-1 Shikata-cho, Kita-ku

department: Department of Nephrology, Rheumatology, Endocrinology and Metabolism

institute: Okayama University

summary:
mir221/222 AdipoKO and mir221/222 flox/y C57BL/6J mice were fed with standard diet (STD) and high fat-high sucrose diet (HFHS).
We used microarrays to investigate the mRNA expression profile in the epididymal adipose tissues.

overall_design: mir221/222 AdipoKO and mir221/222 flox/y C57BL/6J mice were housed in cages and maintained on a 12-hour light-dark cycle. The were fed with STD (NMF; Oriental Yeast) and HFHS chow (D12331; Research Diet, New Brunswick, NJ). The mice were sacrificed at 16 weeks of age and epididymal adipose tissues were used for RNA extraction and applied to GeneChip Mouse Gene 2.0 ST Array.

first: Satoshi

last: Yamaguchi

first: Dongxiao

last: Zhang


title: Gene expression profiling of mouse pancreatic tumors derived from acinar or ductal cells

status: Public on Jan 05 2021

city: Stanford

first: Brittany

middle: Maria

last: Flowers

state: CA

address: 269 Campus Drive

department: Department of Radiation Oncology

institute: Stanford University

summary: Acinar and ductal cells are the major epithelial cell types in the exocrine pancreas, but which of these serves as the cell-of-origin in human PDAC has been debated. Our mouse models have demonstrated that oncogenic Kras expression and Trp53 loss in pancreatic acinar or ductal cells can lead to pancreatic cancer. Here, we performed gene expression profiling of bulk acinar cell-derived and ductal cell-derived mouse pancreatic tumors to identify the transcriptomic profiles.

overall_design: RNA-Seq was performed on 7 tumors originating from mouse models of acinar cell-derived (KT;Ptf1aCreER;Trp53fl/fl) or ductal cell-derived (KT;Sox9CreER;Trp53fl/fl) pancreatic cancer. KT = KrasLSL-G12D/+;R26LSL-tdTom/LSL-tdTom

first: Laura

middle: D

last: Attardi


title: Gene expression signatures for GP73-induced non-obese nonalcoholic fatty liver disease

city: Beijing

first: Luming

last: Wan

address: Dongda Street No.20

country: China

institute: Beijing Institute of Biotechnology

summary: In searching for common and specific features of transcriptional changes associated with non-obese NAFLD induced by GP73 and obese NAFLD induced by high-fat diet (HFD), we used Agilent SurePrint G3 Mouse Gene Expression Microarray 8x60K to analyze gene expression in the livers isolated from mice 12 months after AAV-V or AAV-GP73 injection and in the livers isolated from AAV-V control mice fed a HFD for 12 months.

overall_design: Mice were injected with AAV-V and AAV-GP73 viruses intravenally, given a normal diet and a high-fat diet, and their liver gene expression was measured.

first: Hui

last: Zhong

first: Congwen

last: Wei


title: H2AK119ub1 ChIP-seq in rat peripheral nerve

status: Public on Dec 24 2020

city: Madison

first: John

last: Svaren

state: WI

address: 1500 Highland Ave.

department: Comparative Biosciences

institute: University of Wisconsin

summary: ChIP-seq of H2AK119ub1 in sham and injured rat peripheral nerve.

overall_design: H2AK119ub1 ChIP samples were prepared from rat sciatic nerve at 1 day post-transection using both the distal stump of the injured nerve and the contralateral (sham) nerve.

first: Ki

middle: H

last: Ma


title: Differentially expressed genes after miRNA transfection in human cancer cell lines

status: Public on Dec 15 2020

city: Kagoshima

first: Shunsuke

last: Misono

address: 8-35-1, Sakuragaoka

department: Pulmonary Medicine

institute: Kagoshima University Hospital

summary: To identify differentially expressed genes by miRNAs transfection in human cancer, several cell lines (lung adenocarcinoma, lung squamous cell carcinoma and small cell lung cancer) were subjected to Agilent whole genome microarrays.

overall_design: Human cell lines (EBC-1, SK-MES-1 and SBC-3) were treated with miRNAs (miR-150-5p, miR-150-3p) and siRNAs (siDONSON-1, 2, 3). A549 knocked down LOXL2 by genome editing with the CRISPR/Cas9 system.

first: Naohiko

last: Seki

first: Kengo

last: Tanigawa


title: Enhancers predominantly control transcription initiation [Capture-C]

city: Oxford

first: Martin

middle: Stephen Charles

last: Larke

address: Headley Way

institute: University of Oxford

summary: Gene transcription occurs via a cycle of linked events including initiation, promoter proximal pausing and elongation of RNA polymerase II (Pol 2).  A key question is how do transcriptional enhancers influence these events to control gene expression? Here we have used a new approach to quantify transcriptional initiation and pausing in-vivo, while simultaneously identifying transcription start sites (TSSs) and pause-sites (TPSs). When analysed in parallel with nascent RNA-seq, these data show that differential gene expression is achieved predominantly via changes in transcription initiation rather than Pol 2 pausing or elongation. Using genetically engineered mouse models deleted for specific enhancers we show that these elements control gene expression via Pol 2 recruitment and/or initiation rather than via promoter proximal pause release. Using genome-wide analysis we show that enhancers, in general, control gene expression at the stage of Pol 2 recruitment and initiation rather than via pausing.

overall_design: NG Capture-C combines 3C library preparation with oligonucleotide capture for the desired viewpoint restriction fragments; in this case haemoglobin alpha (hba-a1)  haemoglobin beta (Hbb-b1) and mitoferrin (slc25a37), were used as viewpoints. The method was applied to primary mouse fetal liver erythroid cells in which the HS2 and HS3 enhancers of beta globin are homozygously deleted in combination.  Wildtype and R1R2 KO have been published and data were deposited under GSE78803.

first: Damien

last: Downes

first: A

last: Marieke Oudelaar

first: Jim

last: Hughes

first: Douglas

last: Higgs


title: CircCDKN2B-AS1 interacts with IMP3 to stabilize Hexokinase 2 mRNA and facilitate cervical cancer aerobic glycolysis progression

status: Public on Dec 21 2020

city: Hangzhou

first: Yanan

state: Zhejiang

address: No. 1 Xueshi Road - Shangcheng

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

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

overall_design: Refer to individual Series


title: circularRNA Sequencing of HPV16 positive cervical cancer tissues, HSIL tissues and normal cervical epithelial tissues

summary: To explore the circRNA expression profiles during the development and progression of cervical cancer, we performed RNA sequencing analysis with ribosomal RNA-depleted in HPV negative normal cervical epithelium, HPV16 positive normal cervical epithelium, HPV16 positive high-grade squamous intraepithelial lesion (HSIL), and HPV16 positive cervical squamous cell carcinoma tissues,6 cases in each group.Totally 66868 circRNAs were identified (Back-spliced junctions reads≥1)

overall_design: RNA sequencing with ribosomal RNA-depleted of  HPV negative normal cervical epithelium, HPV16 positive normal cervical epithelium, HPV16 positive high-grade squamous intraepithelial lesion (HSIL), and HPV16 positive cervical squamous cell carcinoma tissues,6 cases in each group  were generated by deep sequencing, using Illumina Hiseq4000

first: Lu

last: Zhao


title: RNA sequencing data of Ssu72 Wild Type and Knock-Out MEFs, hepatocytes and ES cells

status: Public on Dec 28 2020

city: Suwon

first: Hyun-Soo

last: Kim

address: 2066, SEOBU-RO, JANGAN-GU, SUWON-SI, GYEONGGI-DO

department: School of medicine

institute: SUNGKYUNKWAN UNIVERSITY

summary: To investigate how mammalian Ssu72 regulated gene expression, we performed RNA-Seq analyses of Ssu72 in three different cell types depending on their proliferation capabilities: embryonic stem (ES) cells (highly proliferating), primary murine embryonic fibroblasts (MEFs) (moderate proliferating), and hepatocytes (almost quiescent). Ssu72 deletion was induced by infection with Cre recombinase adenovius (Ad-Cre). Infection of ESf/f, MEFf/f, and hepatocytef/f with Ad-Cre resulted in efficient reduction of Ssu72 levels. Depletion of Ssu72 resulted in downregulation or upregulation of large subsets of genes in three different cell types.

overall_design: MEFs, hepatocytes and ES cells mRNA profiles of 6-weeks old wild type (WT) and Ssu72 knock-out (KO) mice were generated by deep sequencing, in duplicate or triplicate, using Illumina GAIIx.

first: Chang-Woo

last: Lee


title: RNA-seq and ChIP-seq of Ssu72 Wild Type and Knock-Out MEFs, hepatocytes and ES cells


title: Genome-wide association studies reveal susceptibility loci for digital dermatitis in Holstein cattle

status: Public on Dec 07 2020

city: Davis

first: Anita

last: Oberbauer

address: One Shields Ave.

department: Animal Science Department

institute: University of California, Davis

summary: Digital dermatitis (DD) causes lameness in dairy cattle. To detect quantitative trait loci (QTL) associated with DD, genome-wide association studies (GWAS) were performed using high density single nucleotide polymorphism (SNP) genotypes and binary case/control, quantitative (average number of FW per hoof trimming record), and recurrent (cases with  2 DD episodes vs. controls) phenotypes from cows across four dairies (controls n = 129 vs. FW n = 85). Linear mixed model (LMM) and random forest (RF) approaches identified top SNPs, which were used as predictors in Bayesian regression models to assess SNP predictive value. The LMM and RF analyses identified a QTL on BTA1 for the binary phenotype and on BTA2 for the quantitative phenotype. The binary and recurrent LMM GWASs also detected a QTL on BTA2 that was at a different position than the QTL from the quantitative phenotype analysis. The QTL on BTA1 lacked apparent candidate genes, whereas both QTL on BTA2 included genes associated with immune response and wound healing. Although larger sample sizes are necessary to reaffirm these small effect loci amidst a strong environmental effect, the sample cohort used in this study was sufficient for estimating SNP effects with high predictive value.

overall_design: Whole blood was collected from 222 cows (cases n = 90, controls n = 132) from the coccygeal vein in the tail. Blood samples were centrifuged, and genomic DNA was extracted from the resulting buffy coat layer using the QIAGEN QIAamp DNA Blood Mini Kit . DNA samples were genotyped on the Illumina BovineHD BeadChip, and Illumina’s GenCall algorithm was used to call genotypes. Of the 222 samples, six were removed during quality filtering (cases n = 3, controls n = 3), leaving 216 cows for analysis (cases n = 87, controls n = 129). Of the 87 cases, 24 had recurrent FW episodes and were used in the GWAS of controls vs. recurrent FW cases.

first: Ellen

last: Lai

first: Alexa

middle: L

last: Danner

first: Thomas

middle: R

last: Famula


title: Single-cell profiling of myeloid cells in glioblastoma across species and disease stage reveals macrophage competition and specialization

city: Brussels

first: Daliya

last: Kancheva

address: Pleinlaan 2

country: Belgium

institute: Vrije Universiteit Brussel

summary: Glioblastomas are aggressive primary brain cancers that invariably recur as therapy-resistant tumors. Myeloid cells control glioblastoma malignancy, but their dynamics during disease progression remains poorly understood. By relying on single-cell RNA and protein profiling, we mapped the glioblastoma immune landscape in newly diagnosed and recurrent patients and in mouse tumors. This revealed a large and diverse myeloid compartment, with dendritic cell and macrophage populations that were conserved across species and were dynamic across disease stages. Tumor-associated macrophages (TAMs) consisted of microglia- or monocyte-derived populations, with both exhibiting additional heterogeneity, including subsets with conserved lipid and hypoxic signatures. Microglia- and monocyte-derived TAMs (Mo-TAMs) were self-renewing populations that competed for space and could be depleted via CSF1R blockade. Microglia-derived TAMs were predominant in newly diagnosed tumors but were outnumbered by Mo-TAMs upon recurrence, especially in hypoxic tumor environments. Our results unravel the glioblastoma myeloid landscape and provide a framework for future therapeutic interventions.

overall_design:
samples of WT mouse GL261 glioblastoma (GBM), samples of CCR2-KO mouse GL261 GBM, samples of WT mouse GL261 tumor-bearing whole brain, samples of human recurrent GBM, samples of human newly diagnosed GBM

***Please note that the human raw data will be deposited to the European Genome-phenome Archive (EGA) with study ID EGAS00001004871, as this is a dataset with restricted access

first: Ana

last: Antunes

first: Isabelle

last: Scheyltjens

first: Francesca

last: Lodi

first: Julie

last: Messiaen

first: Asier

last: Antoranz

first: Johnny

last: Duerinck

first: Liesbet

last: Martens

first: Karen

last: De Vlaminck

first: Hannah

last: Van Hove

first: Signe

middle: S

last: Hansen

middle: M

last: Bosisio

first: Koen

last: Van der Borght

first: Steven

last: De Vleeschouwer

first: Raf

last: Sciot

first: Luc

last: Bouwens

first: Michiel

last: Verfaillie

first: Niels

last: Vandamme

first: Roosmarijn

last: Vandenbroucke

first: Olivier

last: De Wever

first: Yvan

last: Saeys

last: Guilliams

first: Conny

last: Gysemans

first: Bart

last: Neyns

first: Frederik

last: De Smet

first: Diether

last: Lambrecht

first: Jo

middle: A

last: Van Ginderachter

first: Kiavash

last: Movahedi


title: Gene expression data of organoids derived from neuroendocrine carcinomas

first: Kohta

last: Toshimitsu

address: 35, Shinanomachi, Shinjuku-ku

institute: Keio University

summary: Gastroentero-pancreatic neuroendocrine neoplasm(GEP-NEN) is consisted of neuroendocrine tumor and neuroendocrine carcinoma, which is a lethal, but under-investigated disease owing to its rarity. We established an organoid library of GEP-NEN and added their comprehensive molecular characterization.

overall_design: We analyzed gene expression of 7 organoids derived from neuroendocrine carcinoma patients. Total RNA were extracted from organoids and hybridized to GeneChip PrimeView Human Gene Expression Array, and scanned with GeneChip Scanner 3000 7G.

first: Kenta

last: Kawasaki


title: Capture of mouse and human stem cells with features of formative pluripotency [mouse, RNA-Seq 2]

status: Public on Dec 04 2020

city: St Louis

first: Sabine

last: Dietmann

state: MO

address: 4565 McKinley Avenue

institute: Washington University School of Medicine

summary: Pluripotent cells emerge as a naïve founder population in the blastocyst, acquire capacity for germline and soma formation, and then undergo lineage priming. Mouse embryonic stem (ES) cells and epiblast stem cells (EpiSCs) respectively represent the initial naïve and final primed phases of pluripotency. Here we investigated the intermediate formative stage. Using minimal exposure to specification cues, we derived stem cells from formative mouse epiblast. Unlike ES cells or EpiSCs, formative stem (FS) cells responded directly to germ cell induction. They colonised somatic tissues and germline in chimaeras. Whole transcriptome analyses showed similarity to pre-gastrulation formative epiblast. Signal responsiveness and chromatin accessibility features reflect lineage capacitation. Furthermore, FS cells showed distinct transcription factor dependencies, relying critically on Otx2. Finally, FS cell culture conditions applied to human naïve cells or embryos supported expansion of similar stem cells, consistent with a conserved staging post on the trajectory of mammalian pluripotency.

overall_design: Transriptome analysis of different type of pluripotent stem cell

first: Masaki

last: Kinoshita

first: Michael

last: Barber

first: Austin

middle: G

last: Smith


title: Capture of mouse and human stem cells with features of formative pluripotency [mouse, RNA-Seq 1]

overall_design: Transcriptome of mouse embryonic epiblast and pluripotent cells in culture


title: Capture of mouse and human stem cells with features of formative pluripotency [human, RNA-seq]

overall_design: Transcriptome of human formative stem cells


title: Targeting liver sinusoidal endothelial cells with miR-20a-loaded nanoparticles reduces murine colon cancer metastasis to the liver

status: Public on Jan 26 2021

city: Muenster

first: Marcos

last: Araúzo-Bravo

address: Rogentstrasse

institute: Max Planck Institute for Molecular Biomedicine

summary:
Phenotypic transformation of liver sinusoidal endothelial cells is one of the most important stages of liver metastasis progression.
The miRNA effects on liver sinusoidal endothelial cells during liver metastasis have not yet been analysed.
Herein, analysis of miRNA expression in these cells during colorectal liver metastasis revealed repressed expression of microRNA-20a.
Importantly, downregulation of miR-20a occurs in parallel with upregulation of its known protein targets.
To restore normal miR-20a levels in liver sinusoidal endothelial cells,
we developed chondroitin sulfate-sorbitan ester nanoparticles conjugated with miR-20a in a delivery system that specifically targets liver sinusoidal endothelial cells.
The restoration of normal mir-20a levels in these cells induced downregulation of the expression of its protein targets,
and this also resulted in a reduction of in vitro LSEC migration and a reduction of in vivo activation and tumor-infiltrating capacity and ability of the tumor decreased by approximately 80% in a murine liver metastasis model.
Animals:
Balb/c mice (6- to 8-week-old males) were obtained from Charles River Laboratories Spain SA (Barcelona, Spain).
All the procedures were approved by the Ethical Committee for Animal Experimentation (CEEA) of the
University of the Basque Country (EHU/UPV) in accordance with institutional,
national and international guidelines regarding the protection and care of animal use for scientific purposes.
Mice were kept in the animal facility of EHU/UPV and had access to standard chow and water ad libitum.
Colorectal cancer cells:
Murine colorectal cancer C26 cells (ATCC, Manassas, VA) syngeneic with Balb/c mice were used.
The cells were grown under standard conditions in RPMI medium (Sigma-Aldrich, St. Louis, MO, USA)
supplemented with 10% fetal bovine serum (FBS), penicillin (100 U/ml), streptomycin (100 microg/ml) and amphotericin B (0.25 microg/ml),
all purchased from Life technologies, Carlsbad, CA.
Control and tumor-activated LSECs culture:
Control and tumour activated primary cultures of LSECs were isolated form livers with colorectal cancer metastasis or from healthy ones.
To do so, Balb/c mice were anesthetized with isofluorano and small cut done in their left broadside.
Then, 50 microl for each mice of C26 colon carcinoma cells were inoculated in the spleen at 2 x106 cells/ml concentration
and the peritoneum and skin were sutured.
The control mice were inoculated with PBS 1x buffer. Fourteen days later,
all mice were sacrificed and the tumour activated LSECs and control LSECs were isolated by differential centrifugation.
To do so, mice were perfused with collagenase P from Clostridium histolyticum (Sigma-Aldrich, St.Louis, MO, USA) through cava vein.
The obtained cell suspension was twice centrifuged resulting in a parenchymal (PC)-enriched pellet and a non PC-enriched supernatant.
Then, the non PC-enriched supernatant was layered on Percoll gradients (25% on top of 50%) to obtain LSEC.
After a centrifugation the interphase between the two density cushions was recollected with purified non-PC enriched LSECs and
Kupffer cells (KC).
Then, the KCs were separated by adherence and isolated LSECs were washed and cultured with RPMI 1640 medium
(Sigma-Aldrich, St.Louis, MO, USA) supplemented with 10% FBS and used in different experiments 24 hours after the isolation.
(Life technologies, Carlsbad, CA).
miRNA microarray analysis :
The purified total RNA was analysed using Agilent mouse miRNA microarrays.
Release 18.0, 8x60K (v18) microarray slides (Agilent Technologies, USA), with 1,200 mouse miRNAs represented, were used.
Briefly, 100 ng of total RNA was labelled and hybridized following the standard Agilent Protocol for the miRNA Microarray System
with a miRNA Complete Labelling and Hybridization Kit, including Agilent miRNA Spike-ins,
and the results were scanned using an Agilent Microarray Scanner G2565CA (Agilent Technologies, USA).
The scanned TIFF image files were processed using Agilent Feature Extraction Software vs 10.7.3.1 to extract raw data and obtain QC reports.
The microarray raw data were normalized using the quantile method.
To find the statistically significant differentially expressed miRNAs (DEMs) between two groups of samples,
we calculated the mean of each probe across all the samples of the group.
Next, we filtered out all the probes whose absolute value of the difference of mean values between the two groups was less than a
selection threshold thetaDEM = 4 (that corresponds to a fold change of 2 in log2 scale),
and we applied a Student's t-test with a significance threshold alphaDEM = 0.05.
The PCA and the hierarchical clustering of genes and samples were performed with one minus the correlation metric and the unweighted
average distance (UPGMA) linkage method.
All the analysis algorithms and graphics were implemented functions in Matlab.

overall_design:
5 samples were analyzed
ET, Tumour, 3 replicates
EC, No tumour, 2 replicates

first: Joana

last: Marquez

first: Ines

last: Fernandez-Piñeiro

first: Gereon

last: Poschmann

first: Kai

last: Stühler

first: Abdel-Majid

last: Khatib

first: Alejandro

last: Sanchez

first: Fernando

last: Unda

first: Gaskon

last: Ibarretxe

first: Irantzu

last: Bernales

first: Iker

last: Badiola


title: Capture of mouse and human stem cells with features of formative pluripotency [ATAC-Seq]

overall_design: ATAC-seq analysis of different pluripotent cell types


title: Phased gene expression and chromatin accessibility during progressive EMT and MET linked to dynamic CTCF engagement (RNA-Seq)

status: Public on Dec 01 2020

city: Waco

first: Joe

last: Taube

state: Texas

address: One Bear Place #97388

institute: Baylor University

summary: We report the chromatin accessibility alterations and gene expression changes in TGFB-induced and -withdrawn reversible EMT in MCF10A mammary epithelial cells

overall_design: MCF10A cells were treated and withdrawn from TGFB for various durations and subjected to ATAC- and RNA-seq

first: Kelsey

last: Johnson

first: Joseph


title: The Human Accelerated Region HACNS1 modifies developmental gene expression in humanized mice

city: New Haven

first: James

middle: P.

last: Noonan

state: CT

address: 333 Cedar St

department: Genetics

institute: Yale University

summary: Understanding how uniquely human genetic changes altered developmental processes is essential to understanding human evolution. We investigated the role of the Human Accelerated Region HACNS1 in human limb evolution by directly interrogating its biological functions in a humanized mouse model. Using genome-wide epigenetic profiling, we found that HACNS1 maintains its human-specific transcriptional enhancer activity compared to its chimpanzee ortholog in the mouse embryonic limb, and that it alters promoter activity of the transcription factor gene Gbx2. Using single-cell transcriptional profiling, we demonstrate that Gbx2 is upregulated in humanized limb bud chondrogenic mesenchyme, providing insight into the developmental processes altered due to human-specific alterations in enhancer activity. Our findings establish that HACNS1 directs changes in the level and distribution of gene expression during development, and illustrate how humanized mouse models provide insight into regulatory pathways altered in human evolution.

overall_design: For ChIP-seq, tissue was collected from E11.5 forelimb and hindlimb bud pairs or pharyngeal arch tissue from HACNS1 and chimpanzee ortholog line heterozygous crosses to obtain two biological replicates each of pooled, litter matched limb bud or pharyngeal arch samples for all three genotypes (HACNS1+/+, chimpanzee ortholog line, and wild type). Each replicate is comprised of H3K27ac ChIP, H3K4me2 ChIP, and input control data. For scRNA-seq, tissue was collected from E11.5 hindlimb from HACNS1+/+, chimpanzee ortholog line, and wild type litters from homozygous crosses. Left hindlimb buds from three embryos per genotype per replicate were pooled.

first: E

middle: V

last: Dutrow

first: J


title: Drug Combination Signatures for Prediction and Mitigation of Toxicity

status: Public on Nov 30 2020

city: New York

first: Ravi

last: Iyengar

state: NY

address: 1425 Madison Avenue, Room 12-70C

department: Pharmacological Sciences

institute: Icahn School of Medicine at Mount Sinai

summary: Generation and extensive characterization of a gender-balanced, racially/ethnically diverse library of iPSCs from forty clinically healthy human individuals who range in age from 22-61. Specifically,  here we provide  mRNAseq data of duplicate samples of one clone from each of these forty iPSC lines.

overall_design:
bulk mRNA-seq data files for 80 human iPSC samples at the control condition generated by random-primed mRNA-sequencing method
The overall goal of the drug signatures for prediction and mitigation of toxicity study is to use genomic and proteomic high-throughput measurements as the basis for computational analysis that integrates network analyses with structural constraints and dynamical models in multiple cell types to identify signatures that predict toxicity induced by 55 FDA-approved chemotherapeutic drugs and potential mitigation of this toxicity.  For this purpose, we have recruited and pre-screend healthy individuals to be included in our study. Ninty six male and female individuals that satisfied the initial pre-screening for the study and their sex, age, race/ethnicity recorded through an enrollment questionnaire were consented. Eighty five underwent a formal and thorough medical health screening. The evlaution involved assessment of all inclusion and exclusion criteria, a full medical history, measurement of weight, height, waist and hip circumference, heart rate, blood pressure, respiratory rate, and oxygen saturation, a physical exam and an electrocardiograph (ECG). Blood was drawn for analysis of clinical relevant parameters. A pregnancy test was included for female participants. All blood draws and pregnancy tests were sent to a certified clinical laboratory for clinical analysis. Of the 85 subjects that were screened 42 (48.3%) were deemed eligible for final inclusion. Of these 42, one declined to undergo skin biopsy and/or venesection and one suject didn’t show up for the scheduled biopsy, leaving 40 clinically healthy subjects on which a skin biopsy and venesection was performed. Fibroblast lines were established from the skin biopsies. Induced pluripotent cells (iPSC) were generated from the established fibroblast lines of these forty clinically healthy racially diverse male and female study participants who ranged in age from 22 and 61 years through reprogramming using either the mRNA or Sendai-virus methods. One clone from each of the forty iPSC line was fully characterized for normal karyotype, short-tandem repeat matching to the original fibroblast line (authentication), pluripotency by select pluripotent marker expression via immunocytochemistry and by mRNAseq-based PluriTest analysis, and whole-genome sequencing. Specifically, mRNAseq was performed in duplicate samples of one clone from each of the 40 iPSC lines.


title: Capture of mouse and human stem cells with features of formative pluripotency

first: Sabine

last: Dietmann


title: Characterization of the cellular adaptation during biofilm formation in Gram positive bacteria

status: Public on Dec 05 2020

city: Sherbrooke

first: Pascale

middle: B.

last: Beauregard

state: Quebec

address: 2500 Boulevard de l'Université

department: Département de Biologie

country: Canada

institute: Université de Sherbrooke

summary: Cellular adaptation during biofilm formation in Gram positive bacteria

sample_organism:
Staphylococcus aureus
Enterococcus faecalis

overall_design: Biofilm formation mRNA profiles of S. aureus SH1000, S. aureus USA300, E. faecalis V583 and E. faecalis 29212 compared to their planktonic state, in either TSBg(s) or BHIg(s) medium, in 3 biological replicates.

first: Gélinas

last: Martin

first: Léa

last: Museau

first: Milot

last: Arielle

first: Beauregard

middle: B

last: Pascale


title: Human PBMCs: Healthy vs Diabetic nephropathy vs ESRD

city: San Francisco

first: Swastika

last: Sur

address: 513 Parnassus Ave

department: Surgery

institute: UCSF

summary: Transcriptional profiling of human PBMCs comparing healthy controls, patients with diabetic nephropathy and patients with ESRD. PBMCs were analyzed as they mediate inflammatory injury. Goal was to determine effects of increasing severity of diabetic nephropathy on global PBMC gene expression. Microarray analysis of PBMCs taken from patients with varying degrees of diabetic nephropathy.

overall_design: 3 condition experiment - Healthy control (10) vs diabetic nephropathy (23) vs ESRD (7)

first: Minnie

last: Sarwal


title: Phased gene expression and chromatin accessibility during progressive EMT and MET linked to dynamic CTCF engagement


title: Targeting transcription-replication conflicts is an effective therapeutic principle for MYCN-driven neuroblastoma [RNA-seq mouse GB]

status: Public on Nov 27 2020

city: Wuerzburg

first: Martin

last: Eilers

address: Am Hubland

department: Chair for Biochemistry and Molecular Biology

institute: University of Wuerzburg

summary:
Targeting the function of MYC proteins for therapy is a major challenge in tumor biology. MYC proteins are transcription factors that can globally release RNA polymerase II (RNAPII) from core promoters. In neuroblastoma, the MYC paralogue, MYCN, and the Aurora-A kinase form a complex during S phase that stabilizes MYCN and enhances Aurora-A kinase activity. Here we show that Aurora-A phosphorylates histone H3 at serine 10 in S phase and suppresses transcription-dependent R-loop formation.
Inhibition of Aurora-A causes global stalling of RNAPII at pause sites and exon/intron boundaries and induces transcription/replication conflicts, activating the Ataxia telangiectasia and Rad3 related (ATR) kinase. Activation of ATR is required to prevent the accumulation of double-strand breaks. In genetically-engineered mice with MYCN-driven neuroblastoma, combined inhibition of Aurora-A and ATR induces rampant tumor-specific apoptosis and tumor regression, leading to permanent eradication of tumors in a subset of mice.
The therapeutic efficacy is not only due to tumor cell-intrinsic mechanisms, but also engages the host immune system for tumor eradication. Since stabilization of MYCN promotes promoter-proximal transcription termination, we propose that MYCN/Aurora-A complex formation enables tumor cells to prevent transcription-replication conflicts and that targeting this mechanism is an effective therapy for MYCN-driven neuroblastoma.

overall_design: THMYCN mice (a neuroblastoma model) were untreated, treated for 24 h, or 5 days with the combination of MLN8237 or AZD6738

first: Gabriele

last: Büchel


title: Targeting transcription-replication conflicts is an effective therapeutic principle for MYCN-driven neuroblastoma [4SU-seq]

overall_design: S phase synchronized IMR-5 cells treated for 4 h with 1 µM MLN8237, 1 µM MK5108, 1 µM Pladienolide B or DMSO, 2 h before harvest cells were incubated for 15 min with 500 µM 4sU

first: Isabelle

last: Roeschert


title: Targeting transcription-replication conflicts is an effective therapeutic principle for MYCN-driven neuroblastoma [ChIP-seq IMR-5]

overall_design: S phase synchronized IMR-5 cells treated for 4 h with 1 µM MLN8237 or 1 µM MK5108 or DMSO


title: Targeting transcription-replication conflicts is an effective therapeutic principle for MYCN-driven neuroblastoma [ChIPseq_IMR5_RNAseqH1]

overall_design:
S phase synchronized IMR-5 cells with inducible RNaseH1 treated for 24 h with 1 µg/ml Doxycycline; ChIP-Sequencing of total RNA Polymerase II and Serine 2 phosphorylated RNA Polymerase II

A combined 'Input.fastq' file was generated by merging the files IR166_INPUT_IMR5_RnaseH_Dox_DMSO.fastq, IR166_INPUT_IMR5_RnaseH_Dox_MK.fastq, IR166_INPUT_IMR5_RnaseH_Dox_MLN.fastq, IR166_INPUT_IMR5_RnaseH_EtOH_DMSO.fastq, IR166_INPUT_IMR5_RnaseH_EtOH_MK.fastq, IR166_INPUT_IMR5_RnaseH_EtOH_MLN.fastq, reseq_IR166_INPUT_IMR5_RnaseH_Dox_DMSO.fastq, reseq_IR166_INPUT_IMR5_RnaseH_Dox_MK.fastq, reseq_IR166_INPUT_IMR5_RnaseH_Dox_MLN.fastq, reseq_IR166_INPUT_IMR5_RnaseH_EtOH_DMSO.fastq, reseq_IR166_INPUT_IMR5_RnaseH_EtOH_MK.fastq & reseq_IR166_INPUT_IMR5_RnaseH_EtOH_MLN.fastq.
The file Input.fastq was then randomly subsampled to 6068151 and 7845813 reads, resulting in the files Input_sorted6068151.bedgraph and Input_sorted7845813.bedgraph, which were used as input controls for the A10- and the S2P-ChIPseq samples, respectively.

A combined Input_H3.3.fastq file was generated by merging the files IR181_Input_Dox_DMSO.fastq, IR181_Input_Dox_MLN.fastq, IR181_Input_EtOH_DMSO.fastq & IR181_Input_EtOH_MLN.fastq.
The file 'Input_H3.3.fastq' was then randomly subsampled to 11604734 reads, resulting in the file Input_sorted11604734.bedgraph, which was used as input control for the H3.3-ChIPseq samples.


title: Targeting transcription-replication conflicts is an effective therapeutic principle for MYCN-driven neuroblastoma [ChIPseq_IMR5]

overall_design:
S phase synchronized IMR-5 cells treated for 4 h with 1 µM MLN8237 or DMSO; ChIP-Sequencing of H3.3

A combined Input.fastq file was generated by merging the files IR180_Input_DMSO.fastq & IR180_Input_MLN.fastq.
The file 'Input.fastq' was then randomly subsampled to 15603418 reads, resulting in the file Input_sorted15603418.bedgraph, which was used as input control for the H3.3-ChIPseq samples.


title: Gene cascade analysis in human granulosa tumor cells (KGN) following exposure to high levels of free fatty acids and insulin

status: Public on Nov 21 2020

city: Québec

first: Patricia

last: Tremblay

state: Québec

address: 2440 Hochelaga Blvd

department: Sciences animales

institute: Laval University

summary: Maternal metabolic disorders such as obesity and diabetes are detrimental factors that compromise fertility and the success rates of medically assisted procreation (MAP) procedures. During metabolic stress, adipose tissue is more likely to release free fatty acids (FFA) in the serum resulting in an increase of FFA levels not only in blood, but also in follicular fluid (FF). In humans, high concentrations of palmitic acid (PA) and stearic acid (SA) reduced granulosa cell survival and were associated with poor cumulus-oocyte complex (COC) morphology. Obesity and high levels of circulating FFA were also causatively linked to hampered insulin sensitivity in cells and compensatory hyperinsulinemia. To provide a global picture of the principal upstream signaling pathways and genomic mechanisms involved in this metabolic context, human granulosa-like tumor cells (KGN) were treated with a combination of PA, oleic acid (OA), and SA at the higher physiological concentrations found in the follicular fluid of women with a higher body mass index (BMI) (≥ 30.0 kg/m2). We also tested a high concentration of insulin alone and in combination with high concentrations of fatty acids. Transcription analysis by RNA-seq with a cut off for fold change of 1.5 and p-value 0.05 resulted in thousands of differentially expressed genes for each treatment. Using analysis software such as Ingenuity Pathway Analysis (IPA), we were able to establish that high concentrations of FFA affected the expression of genes mainly related to glucose and insulin homoeostasis, fatty acid metabolism, as well as steroidogenesis and granulosa cell differentiation processes. The combination of insulin and high concentrations of FFA affected signaling pathways related to apoptosis, inflammation, and oxidative stress. Taken together, our results provided new information on the mechanisms that might be involved in human granulosa cells exposed to high concentrations of FFA and insulin in the contexts of metabolism disorders.

overall_design: Cells treated with differents concentations of free fatty acids and insulin alone or in combination. 3 reps per treatment were pooled for RNAseq analysis.

first: Marc-André

last: Sirard

first: Chloé

last: Fortin


title: Targeting transcription-replication conflicts is an effective therapeutic principle for MYCN-driven neuroblastoma [PDX]

overall_design: PDX mouse model was treated for 24 h or 14 days with vehicle or the combination of MLN8237 & AZD6738


title: Targeting transcription-replication conflicts is an effective therapeutic principle for MYCN-driven neuroblastoma [ChIP-seq GB]

overall_design: double thymidin block of IMR-5 cells. Release, Treatment and harvest after 4 h (S)


title: Ubiquitin-specific peptidase 39 promotes human glioma cells migration and invasion by facilitating ADAM9 mRNA maturation

status: Public on Oct 23 2020

city: Shenzhen

first: Yue

last: Xiao

address: xueyuan road

institute: Shenzhen University

summary: Glioma is characterized by high migration and invasion, and the relative molecular mechanism is still poor. Accumulating studies demonstrated that ubiquitin specific protease 39 (USP39) played an oncogenic role in several cancers. Here, we investigate USP39 expression and function in human glioma. Oncomine database analysis revealed high USP39 expression in glioma and elevated USP39 expression correlated significantly with poor overall survival. Knockdown of USP39 significantly inhibited the migration and invasion of U251 and U87 cells. The gene expression profile was executed to screen the target molecules of USP39. The result showed that ADAM9, a molecule involved in migration and invasion of various human tumors, was significantly downregulated in the U251 and U87 cells with shRNA-mediated USP39 knockdown. Mechanistically, USP39 directly interacted with the ADAM9 mRNA and induced ADAM9 mRNA maturation, following the decreased expression of integrin β1. Besides, overexpressed ADAM9 rescued the inhibited migration and invasion of glioma cells causing by USP39 depletion. USP39 promoted invasion in vivo and reduced the overall survival of the mice. Collectively, USP39 may have oncogenic role that increase ADAM9 protein levels by inducing maturation of ADAM9 mRNA in glioma. USP39 could be considered a new potential therapeutic target for glioma.

overall_design: Human glioma cell line U87 were infected with  lentiviruses encoding USP39-targeting or negative control shRNA (shUSP39 or shNA) ,  cells with stable knockdown USP39 or negative control were screened by GFP and puromycin.

first: Weilin

last: Chen


title: Transcriptome of murine gastric cancer treated with anti-IL-17, PD-1 or CTLA-4 antibodies.

status: Public on Oct 28 2020

city: Osaka

first: Koji

last: Nagaoka

address: 377-2 Ohno-Higashi, Osaka-Sayama

department: Department of Immunology

institute: Kindai University Faculty of Medicine

summary: YTN2 and YTN16 cells are chemically induced gastric cancer cell lines. YTN2 is spontaneously regressed in C57BL/6 mice in CD8+ T cell-dependent manner, while YTN16 grows progressively.  To investigate the interaction of tumor microenvironment and T cells, bulk RNA-Seq of total tumor RNA were performed in YTN2 and YTN16 gastric tumor-bearing mice.

overall_design: Mice were inoculated with 5 ×106 YTN2 or YTN16. Anti-IL-17(clone 17F3), anti-PD-1(RMP1-14) or anti-CTLA-4 (9H10) mAbs were injected on days 3, 6, 9, and 12, days 6, 9, and 12 or days 0, 4 and 8, respectively. Tumors were harvested on day 7, 10, 14 or 21. Expression profiles were analyzed by RNA-sequencing.

first: Kazuhiro

last: Kakimi


title: UVDE-seq mapping of UV-induced non-CPD lesions

status: Public on Nov 19 2020

city: Pullman

first: John

middle: J.

last: Wyrick

state: WA

address: Biotechnology Life Sciences 241

department: School of Molecular Biosciences

institute: Washington State University

summary: Here we have used a novel high-throughput sequencing method known as UVDE-seq to map the formation of non-CPD lesions at single nucleotide resolution across the yeast genome in wild-type and rad16 mutant cells, which are deficient in GG-NER.

overall_design: UV damage mapping data was analyzed for wild-type BY4741 yeast cells and mutant yeast cells deleted for the RAD16 gene. (i.e., rad16∆). Yeast cells were irradiated with 600J/m2.

first: Kaitlynne

last: Bohm


title: Environment-driven reprogramming of gamete DNA methylation occurs during maturation and influences offspring fitness in salmon

status: Public on Nov 26 2020

first: Kyle

last: Wellband

state: QC

address: 1030 avenue de la Médecine

department: Institut de biologie intégrative et des systèmes

institute: Université Laval

type: Methylation profiling by high throughput sequencing

summary: We investigated whether exposure to a captive environment during maturation influenced gamete DNA methylation for wild Atlantic Salmon individuals. We then investigated whether these parental effects were detectable in an F1 generation reared in a common environment. We associated DNA methylation with growth and fitness-related phenotypes and demonstrated that intergenerational effects of hatchery exposure during maturation of the parental generation influence fitness-related methylation patterns in the F1 generation.

overall_design: We compared DNA methylation of sperm for 6 wild males and 6 males that had been reared in captivity during maturation (SAS). Pure-type crosses of wild and SAS fish were created and we compared DNA methylation in liver tissue of F1 offspring (approximately 10 month old juveniles). For juvenile comparisons we used two offspring from each of four families for both wild and SAS groups (N total = 16).

first: Louis

last: Bernatchez


title: Mouse Olfactory Epithelium, whole olfactory mucosa, mouse

status: Public on Nov 16 2020

city: Berkeley

first: Rebecca

middle: K.

last: Chance

department: Molecular and Cell Biology

institute: UC Berkeley

summary: The lineage of wild-type horizontal basal cells (HBC) stem cells  from the olfactory epithelium were profiled by single-cell RNA-Seq (10X v3 chemistry) to identify differences in aged versus not-aged adult stem cells mRNA expression profiles

overall_design: Whole olfactory epithelium cells are FACS-purified via live cell repoter in young and old adult mice. A minimum of two biological replicates were collected for each time-point/experimental condition.

middle: K

first: Hsin-Jung

last: Chou


title: Adherent Intestinal Cells from Atlantic Salmon Show Phagocytic Ability and Express Macrophage Markers

status: Public on Nov 10 2020

city: Bodø

first: Jorge

last: Fernandes

address: Universitetsalleen 11

department: Faculty of Biosciences and Aquaculture

country: Norway

institute: Nord University

summary: Atlantic salmon (average 550g) were used in this study. The fish were starved for 24 h and were sacrificed with an overdose of MS-222. Then head kidney (HK) and distal intestine (DI) samples were collected. Immune cells from HK and DI were isolated and cultured at 12°C in Leibovitz’s L-15 Medium (L-15). Both HK and DI leukocytes were allowed to adhere on a petri dish with 2 mL L-15+ for 2 days at 12°C. After removing the supernatant containing non-adherent cells, the petri dish with the adherent cells was placed on ice for 10 min, and the cells were detached by washing three times with 1.5 mL ice-cold PBS supplemented with 5 mM EDTA. The cells obtained were centrifuged (500 × g, 5 min, 4°C) and re-suspended in 1 mL L-15+. Then, the cells were counted using a portable cell counter. Total RNA was extracted from adherent cells from HK or DI (< 500,000 cells). To compare the expression of marker genes between adherent cells from HK and DI, 12 libraries were prepared (6 replicates for each). Transcriptomic analysis revealed that adherent intestinal cells (AICs) expressed genes associated with macrophages, T cells, and endothelial cells. In addition, AIC had higher expression of TNF and TGFβ-related genes that can be linked to macrophages and T cells. Heatmap analysis of selected genes indicated that the adherent cells from the two organs had apparently higher expression of macrophage-related markers.

overall_design: Twelve samples were prepared including 6 from distal intestine and 6 from head kidney.

first: Youngjin

last: Park

first: Qirui

last: Zhang

first: Geert

last: Wiegertjes

first: Viswanath

last: Kiron


title: Age-related changes in gene expression in mouse cochlea

status: Public on Oct 31 2020

city: Charleston

first: Jeremy

last: Barth

state: SC

address: 173 Ashley Ave, BSB 627

institute: MUSC

summary: Age-related hearing loss is a multifactorial and progressive process, which negatively impacts quality of life in many senior adults as the number one chronic neurodegenerative condition. This study was done to examine gene expression changes occurring in mouse auditory nerve and cochlear lateral wall tissues that may contribute to age-related hearing loss. In addition to conducting general differential expression analysis, a focused analysis of genes linked to neural cells was done.

overall_design: Young adult (1-3 months) and aged (1.5 year) CBA/CaJ mice were used in the study. Cochlea (2) were used to prepare individual experimental replicates, with three samples for young adults and for aged conditions (n=3).

first: Hainan

last: Lang


title: BT20_24hr_DMSOvERLOTINIB_RNAseq and WHOLE GENOME CRISPR screen

status: Public on Nov 01 2020

city: Worcester

first: Alper

last: Kucukural

address: 364 Plantation Street

department: Program in Molecular Medicine

institute: UMass Medical School

summary:
BT20 (TNBC) cells were cultured with either DMSO (v/v%) or Erlotinib (10uM) for 24 hours to observe differential expression changes between conditions.

CAS-9 expressing BT20 (TNBC) cells were transduced with a whole-genome library of CRISPR sgRNAs; then, cultured them with either DMSO (v/v%) or Erlotinib (10uM) and collected samples to observe differential sgRNA abundances between conditions.

sample_organism:
Homo sapiens
synthetic construct

platform_organism:
Homo sapiens
synthetic construct

overall_design:
BT20 cells were cultured appropriately and treated for 24 hours --> Phenol-Chloroform extraction of RNA --> TruSeq HT Stranded (mRNA) library preparation --> Whole library sequenced on Illumina Nextseq platform

BT20 cells were cultured appropriately and treated for 3-4 population doublings  --> gDNA was isolated using phenol-chloroform extraction --> 2 PCRs were done to a) amplify sgRNAs and b) to attach barcodes for library preparation --> Whole library sequenced on Illumina Nextseq platform

first: Peter

last: Cruz-Gordillo

first: Michael

middle: J

last: Lee


title: Expression of circular RNAs in human HT29, 293FT and HeLa cells

status: Public on Oct 09 2020

city: Shanghai

first: Li

last: Yang

address: 131 Dong-An Road

department: Institutes of Biological Sciences

institute: Fudan University

summary: Deep sequencing datasets are used for circRNA expression analysis in  human HT29, 293FT and HeLa cells.

overall_design: Ribo– RNA-seq for circRNA expression analysis

first: Ling-Ling

last: Chen

first: Siqi

last: Li

first: Xiang

first: Wei

last: Xue


title: An Integrative Omics Approach Reveals Involvement of BRCA1 in Hepatic Metastatic Progression of Colorectal Cancer (microRNA)

status: Public on Nov 02 2020

first: Marcos

last: Araúzo-Bravo

department: Cell and Developmental Biology

institute: Max Planck Institute for Molecular Biomedicine

summary:
The role of different cells in the tumor microenvironment (TME) is critical to the metastatic process.
Phenotypic transformation of the liver cells is one of the most important stages of hepatic metastasis progression of colorectal cancer (CRC).
Our aim was to identify the major molecules (i.e. genes, miRNAs and proteins) involved in this process.
We isolated and performed whole genome analysis of gene, miRNA and proteins expression in three types of liver cells
(Ito cells, Kupffer cells and liver sinusoidal endothelial cells) from the TME of a murine model of CRC liver metastasis.
We selected the statistically significant differentially expressed molecules using the Student's t-test with Benjamini-Hochberg correction.
and performed functional statistically-significant enrichment analysis of differentially expressed molecules with hypergeometric distribution using
the curated collection of molecular signatures, MSigDB.
To build a gene-miRNA-protein networks centered in Brca1, we developed software that collects miRNA targets from the union of the TargetScan, MicroCosm, mirTarBase and miRWalk databases.
This was used to select miRNAs targeting Brca1. We validated the most relevant miRNAs with real-time quantitative PCR. To investigate BRCA1 protein expression,
we built tissue microarrays (TMAs) from hepatic metastases of 34 CRC patients.
Using integrated omics analyses we observed that Brca1 gene is among the twenty transcripts simultaneously up-regulated in all three types of TME liver cells during metastasis.
Further analysis revealed that Brca1 is the last BRCA1-associated genome surveillance complex (BASC) gene activated in the TME.
We confirmed this finding in human reanalyzing transcriptomics datasets from 184 patients from non-tumor colorectal tissue,
primary colorectal tumor and colorectal liver metastasis of the GEO database. We found that the sequence of cell activation during metastasis is Endothelial->Ito->Kupffer.
Immunohistochemical analysis of human liver metastases showed BRCA1 protein was co-localized in Ito, Kupffer and endothelial cells in 81.8% of early or synchronous metastases.
However, in the greater part of the metachronous liver metastases this protein was not expressed in any of these TME cells.
These results suggest a possible role of the co-expression of BRCA1 in Ito, Kupffer and sinusoidal endothelial cells in the early occurrence of CRC liver metastases,
and point to BRCA1 as a potential TME biomarker.
Animals:
Balb/c mice (6- to 8-week-old males) were obtained from Charles River Laboratories Spain SA (Barcelona, Spain).
All procedures were approved by the Ethical Committee for Animal Experimentation of the University of the Basque Country (EHU/UPV) in accordance with institutional,
national and international guidelines regarding the protection and care of animal use for scientific purposes.
Mice were kept in the animal facility of EHU/UPV and had access to standard chow and water ad libitum.
Colorectal cancer cells:
Murine CRC C26 cells (ATCC, Manassas, VA) syngeneic with Balb/c mice were grown under standard conditions in RPMI medium (Sigma-Aldrich, St. Louis, MO, USA)
supplemented with 10% fetal bovine serum (FBS), penicillin (100 U/ml), streptomycin (100 mg/ml) and amphotericin B (0.25 mg/ml), all purchased from Life technologies, Carlsbad, CA.
Control and tumor-activated hepatic cell isolation and culture:
Control and tumor-activated primary cultures of hepatic cells, LSECs, Ito cells and Kupffer cells were isolated from livers with CRC metastasis, or from healthy controls.
Balb/c mice were anesthetized with isofluorano and underwent surgical incisions on their left broadside.
Mice were inoculated into the spleen at the incision sites using 2x106 of C26 colon carcinoma cells.
Control mice were inoculated with PBS. Fourteen days later, all mice were sacrificed and the liver cells removed and purified by differential centrifugation.
In brief, the mice were perfused with Clostridium histolyticum collagenase P (Sigma-Aldrich, St. Louis, MO, USA)
through the cava vein and the obtained cell suspension twice centrifuged resulting in a parenchymal (PC)-enriched pellet and a non-PC-enriched supernatant.
The non-PC-enriched supernatant was layered on Percoll gradients (25% on top of 50% w/v) to obtain LSECs and on Percoll gradients (33% on top of 50% w/v) to obtain Ito cells.
After centrifugation the interphase between the two density cushions was collected and contained purified non-PC enriched LSECs in the first gradient and Ito cells in the second one.
Both solutions contained Kupffer cells which were further separated by adherence assays.
All cell fractions were washed and cultured with RPMI 1640 medium (Sigma-Aldrich, St. Louis, MO, USA)
supplemented with 10% FBS and used in different experiments a maximum of 24 hours after isolation (Life technologies, Carlsbad, CA).
miRNA microarray analysis :
The purified total RNA was analysed using Agilent mouse miRNA microarrays.
Release 18.0, 8x60K (v18) microarray slides (Agilent Technologies, USA), with 1,200 mouse miRNAs represented, were used.
Briefly, 100 ng of total RNA was labelled and hybridized following the standard Agilent Protocol for the miRNA Microarray System
with a miRNA Complete Labelling and Hybridization Kit, including Agilent miRNA Spike-ins,
and the results were scanned using an Agilent Microarray Scanner G2565CA (Agilent Technologies, USA).
The scanned TIFF image files were processed using Agilent Feature Extraction Software vs 10.7.3.1 to extract raw data and obtain QC reports.
The microarray raw data were normalized using the quantile method.

overall_design:
22 samples were analyzed
EC, Enthothelial control cells, 2 replicates
ET, Enthothelial tumour microenvironment (TME) cells, 3 replicates
IC, Ito control cells, 3 replicates
IT, Ito tumour microenvironment (TME) cells, 3 replicates
KC, Kupffer control cells, 3 replicates
KT, Kupffer tumour microenvironment (TME) cells, 2 replicates
TM, Tumor liver metastasis cells, 3 replicates
TP, Colorectal cancer (CRC) primary cells, 3 replicates

first: Daniela

last: Gerovska

first: Gorka

last: Larrinaga

first: Jon

middle: D

last: Solano-Iturri

first: Joana

last: Márquez

first: Patricia

last: Gallastegi

first: Abdel-Majid

last: Khatib

first: Gereon

last: Poschmann

first: Kai

last: Stühler

first: María

last: Armesto

first: Charles

last: Lawrie

first: Iker

last: Badiola


title: An Integrative Omics Approach Reveals Involvement of BRCA1 in Hepatic Metastatic Progression of Colorectal Cancer (mRNA)

summary:
The role of different cells in the tumor microenvironment (TME) is critical to the metastatic process.
Phenotypic transformation of the liver cells is one of the most important stages of hepatic metastasis progression of colorectal cancer (CRC).
Our aim was to identify the major molecules (i.e. genes, miRNAs and proteins) involved in this process.
We isolated and performed whole genome analysis of gene, miRNA and proteins expression in three types of liver cells
(Ito cells, Kupffer cells and liver sinusoidal endothelial cells) from the TME of a murine model of CRC liver metastasis.
We selected the statistically significant differentially expressed molecules using the Student's t-test with Benjamini-Hochberg correction.
and performed functional statistically-significant enrichment analysis of differentially expressed molecules with hypergeometric distribution using
the curated collection of molecular signatures, MSigDB.
To build a gene-miRNA-protein networks centered in Brca1, we developed software that collects miRNA targets from the union of the TargetScan, MicroCosm, mirTarBase and miRWalk databases.
This was used to select miRNAs targeting Brca1. We validated the most relevant miRNAs with real-time quantitative PCR. To investigate BRCA1 protein expression,
we built tissue microarrays (TMAs) from hepatic metastases of 34 CRC patients.
Using integrated omics analyses we observed that Brca1 gene is among the twenty transcripts simultaneously up-regulated in all three types of TME liver cells during metastasis.
Further analysis revealed that Brca1 is the last BRCA1-associated genome surveillance complex (BASC) gene activated in the TME.
We confirmed this finding in human reanalyzing transcriptomics datasets from 184 patients from non-tumor colorectal tissue,
primary colorectal tumor and colorectal liver metastasis of the GEO database. We found that the sequence of cell activation during metastasis is Endothelial->Ito->Kupffer.
Immunohistochemical analysis of human liver metastases showed BRCA1 protein was co-localized in Ito, Kupffer and endothelial cells in 81.8% of early or synchronous metastases.
However, in the greater part of the metachronous liver metastases this protein was not expressed in any of these TME cells.
These results suggest a possible role of the co-expression of BRCA1 in Ito, Kupffer and sinusoidal endothelial cells in the early occurrence of CRC liver metastases,
and point to BRCA1 as a potential TME biomarker.
Animals:
Balb/c mice (6- to 8-week-old males) were obtained from Charles River Laboratories Spain SA (Barcelona, Spain).
All procedures were approved by the Ethical Committee for Animal Experimentation of the University of the Basque Country (EHU/UPV) in accordance with institutional,
national and international guidelines regarding the protection and care of animal use for scientific purposes.
Mice were kept in the animal facility of EHU/UPV and had access to standard chow and water ad libitum.
Colorectal cancer cells:
Murine CRC C26 cells (ATCC, Manassas, VA) syngeneic with Balb/c mice were grown under standard conditions in RPMI medium (Sigma-Aldrich, St. Louis, MO, USA)
supplemented with 10% fetal bovine serum (FBS), penicillin (100 U/ml), streptomycin (100 mg/ml) and amphotericin B (0.25 mg/ml), all purchased from Life technologies, Carlsbad, CA.
Control and tumor-activated hepatic cell isolation and culture:
Control and tumor-activated primary cultures of hepatic cells, LSECs, Ito cells and Kupffer cells were isolated from livers with CRC metastasis, or from healthy controls.
Balb/c mice were anesthetized with isofluorano and underwent surgical incisions on their left broadside.
Mice were inoculated into the spleen at the incision sites using 2x106 of C26 colon carcinoma cells.
Control mice were inoculated with PBS. Fourteen days later, all mice were sacrificed and the liver cells removed and purified by differential centrifugation.
In brief, the mice were perfused with Clostridium histolyticum collagenase P (Sigma-Aldrich, St. Louis, MO, USA)
through the cava vein and the obtained cell suspension twice centrifuged resulting in a parenchymal (PC)-enriched pellet and a non-PC-enriched supernatant.
The non-PC-enriched supernatant was layered on Percoll gradients (25% on top of 50% w/v) to obtain LSECs and on Percoll gradients (33% on top of 50% w/v) to obtain Ito cells.
After centrifugation the interphase between the two density cushions was collected and contained purified non-PC enriched LSECs in the first gradient and Ito cells in the second one.
Both solutions contained Kupffer cells which were further separated by adherence assays.
All cell fractions were washed and cultured with RPMI 1640 medium (Sigma-Aldrich, St. Louis, MO, USA)
supplemented with 10% FBS and used in different experiments a maximum of 24 hours after isolation (Life technologies, Carlsbad, CA).
Omics analysis:
100ng of total RNA from each fraction was labeled and hybridized onto Agilent mouse RNA and miRNA microarrays, Release 18.0 (Agilent Technologies, USA)
following the standard Agilent Protocol with RNA and miRNA Complete Labelling and Hybridization Kits, including Agilent RNA Spike-ins.
Results were scanned using an Agilent Microarray Scanner G2565CA. Scanned TIFF image files were processed using Agilent Feature Extraction Software (v10.7.3.1) to extract the raw data.
All omics data were normalized with the quantile method.

overall_design:
Two-channel microarrays were 32 samples were analyzed as single channels in 16 Agilent microarrays.
EC, Enthothelial control cells, 4 replicates
ET, Enthothelial tumour microenvironment (TME) cells, 4 replicates
IC, Ito control cells, 4 replicates
IT, Ito tumour microenvironment (TME) cells, 4 replicates
KC, Kupffer control cells, 4 replicates
KT, Kupffer tumour microenvironment (TME) cells, 4 replicates
TM, Tumor liver metastasis cells, 4 replicates
TP, Colorectal cancer (CRC) primary cells, 4 replicates

middle: H


title: An Integrative Omics Approach Reveals Involvement of BRCA1 in Hepatic Metastatic Progression of Colorectal Cancer

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: Interepithelial inflammatory crosstalk mediated by stromal fibroblast communication through hyaluronan [dataset 2]

status: Public on Oct 19 2020

city: La Jolla

first: Jason

middle: Samuel

last: Seidman

address: 9500 Gilman Drive, Mail code 0651

department: Cellular and Molecular Medicine

institute: University of California San Diego

summary: Inflammation of epithelial structures frequently leads to disease at distal organs, but the mechanism responsible for this is unknown. We report evidence that digestion of hyaluronan in the extracellular matrix signals distal stromal cells to become primed for increased inflammation. By applying transgenic mouse models and single cell RNA sequencing of mouse and human tissues we demonstrate that skin injury or infection promotes an adipogenic response in distinct populations of distant submucosal fibroblasts of the colon. This response can be recapitulated without skin inflammation by expression of hyaluronidase in skin that promotes a highly amplified and frequently fatal host response to intestinal injury. Our results uncover an innate system of communication between epithelial stroma that is not initiated by immunocyte trafficking or cytokine signaling.

overall_design: RNA-seq of mouse colon lamina propria

first: Tatsuya

last: Dokoshi

first: Richard

last: Gallo


title: Transcriptome analysis of gene expression of strains JNB5-1 and SK8-37

status: Public on Oct 01 2020

city: Wuxi

first: Xuewei

last: Pan

address: 1800, Lihu Avenue

institute: Jiangnan University

summary: RNAseq analysis was performed to evaluate gene expression differences between strains JNB5-1 and SK8-37. S. marcescens JNB5-1 and SK8-37 cells were grown in LB medium for 10 h before harvesting. The collected cells were then treated with RNAprep pure Kit (TIANGEN) to extract total bacterial RNA and delivered to GENEWIZ in dry ice for transcriptome resequencing analysis. For annotation, the genome of S. marcescens WW4 (NC_020211.1) was used as reference. A total of 31206068 reads matched to the referenced genome in the sample of JNB5-1, and 29862970 reads in the sample of SK8-37. The significantly differentially expressed genes (DEGs) were determined between strains JNB5-1 and SK8-37 with the standards of p-value≤ 0.05, fold change |log2Ratio|≥1. Transcriptome data showed that expression of 60 genes were significantly up-regulated while 558 genes were significantly down-regulated by at least 2-fold when comparing the psrA mutant strain SK8-37 to its parent strain JNB5-1. Based on the annotation of KEGG_B_class, the up-regulated and down-regulated genes were classified into 10 and 19 major cellular processes, respectively. The transcriptome data indicated that PsrA may controls a variety of cellular processes in S. marcescens, including prodigiosin synthesis, serrawttin W1 biosynthesis, extracellular polysaccharide production, biofilm formation, cell motility and T6SS-mediated antibacterial activity.

overall_design: Gene expression of strains JNB5-1 and SK8-37 were generated by deep sequencing, in triplicate, using Illumina HiSeq X10

first: Zhiming

last: Rao


title: Large chromosomal rearrangements associated with recombination between homologous prophage regions are a common feature in E. coli O157 strains and promote phenotypic plasticity in the population

status: Public on Oct 02 2020

city: Edinburgh

first: Stephen

last: Fitzgerald

state: United Kingdom

address: Roslin Institute, University of Edinburgh, Easter Bush

summary: This study characterises large chromosomal rearrangements (LCRs) and their associated phenotypes in E. coli O157 strains. RNA-seq was performed on wildtype strain 9000 and three derivative strains in which LCRs had occurred following experimental bovine colonisation. Gene expression differences between these strains was analysed when each was grown in either LB or M9 media.

overall_design: mRNA and sRNA profiles of strain 9000 and derivative strains Z1615, Z1723 and Z1767 grown in LB medium

middle: F

first: Nadejda

last: Lupolova

first: Sharif

last: Shaaban

first: Timothy

last: Dallman

first: David

last: Greig

first: Lesley

last: Allison

first: Tom

middle: N

last: McNeilly

first: James

middle: L

last: Bono

last: Gally


title: Interepithelial inflammatory crosstalk mediated by stromal fibroblast communication through hyaluronan