title: Transcriptional response in a sepsis mouse model reflects transcriptional response in sepsis patients

status: Public on Dec 31 2021

city: MARSEILLE CEDEX 09

first: Aurelie

last: Bergon

address: parc scientifique de Luminy, case 928

institute: TAGC INSERM U1090

summary: Mortality due to sepsis remains unacceptably high, especially for septic shock patients. Murine models have been used to better understand pathophysiology mechanisms. However, the mouse model is still under debate. Here we investigated the transcriptional response of mice injected with lipopolysaccharide (LPS) and compared it with that of human cells stimulated in vitro with LPS on the one hand, and with that of blood cells in septic patients on the other hand. We identified a molecular signature composed of 2331 genes with an FDR median of 0%. This molecular signature is highly enriched in regulated genes in peritoneal macrophages stimulated with LPS. There is a significant enrichment in several inflammatory signaling pathways, and in disease terms, such as pneumonia, sepsis, systemic inflammatory response syndrome, severe sepsis, an inflammatory disorder, immune suppression, and septic shock. A significant overlap between the genes up-regulated in mouse and human cells stimulated with LPS has been demonstrated. Finally, genes up-regulated in mouse cells stimulated with LPS are enriched in genes up-regulated in human cells stimulated in vitro and in septic patients, who are at high risk of death. Our results support the hypothesis of common molecular and cellular mechanisms between mouse and human sepsis.

overall_design: Twelve C57BL/6SJL mice were purchased from the Charles River Laboratory (L’Arbresle, France). C57BL/6SJL mice were housed under specific pathogen-free conditions and handled in accordance with French and European directives (EU Agreement N° A-13013 03). To model severe form of sepsis, mice were intraperitoneally injected with 200 μL (35 mg/kg) of lipopolysaccharide from Pseudomonas aeruginosa (L9143, Sigma-Aldrich®). The mice were sacrificed by cervical rupture before injection T0 (n=4) and 1 hour after LPS injection during the inflammatory peak T1 (n=4) or 40 hours after LPS injection, just before the death T2 (n=4). Peritoneal cavity cells were collected by peritoneal washes using PBS.

first: Florian

last: Rosier

first: Nicolas

last: Fernandez Nuñez

first: Magali

last: Torres

first: Béatrice

last: Loriod

first: Pascal

last: Rihet

first: Lydie

last: Pradel


title: Mechanical Compression Creates a Quiescent Muscle Stem Cell Niche

status: Public on Feb 06 2022

city: Washington

first: Jiaxiang

last: Tao

state: DC

address: 3970 Reservoir Road NW

department: Oncology

country: USA

institute: Georgetown University

type: Expression profiling by high throughput sequencing

summary: We discovered mechanical compression is able to bring activated MuSCs back to their quiescent stem cell state.

overall_design: Examination of RNA-seq results between uncompressed, compressed and freshly isolated MuSCs.

first: Chen-Ming

last: Fan


title: Comparison of gene expression profile of bronchoalveolar lavage fluid (BAL) from RAO- and IAD- affected horses with respect to control animals

status: Public on Dec 30 2021

city: Legnaro (PD)

first: Serena

last: Ferraresso

state: Padova

address: Viale dell'Università

department: Dept of Comparative Biomedicine and Food Science

country: Italy

institute: University of Padova

summary: An equine immuno-specific oligo microarray platform was designed to evidence differences in gene expression profiles in BAL fluid samples from eight (8) RAO-affected horses, ten (10) IAD-affected horses and seven (7) control animals. An unpaired t test was performed using the software Significant Analysis of Microarrays (SAM). 1763 and 379 genes were found differentially expressed between RAO and IAD horses respectively vs. controls.

overall_design: In this study, we analyzed twenty-five (25) equine BAL fluid samples: seven (7) from control animals, eight (8) from recurrent airway obstruction (RAO)-affected horses and ten (10) from inflammatory airway disease (IAD)-affected horses. Gene expression profiling was performed using an equine immuno-specific oligo microarray platform based on single-colour detection (Cyanine-3 only). Microarrays were scanned with Agilent scanner G2565BA (barcode on the left, DNA on the back surface, scanned through the glass) at a resolution of 5 microns; all slides were scanned twice at two different sensitivity settings (XDRHi 100% and XDRLo 10%); the scanner software created a unique ID for each pair of XDR scans and saved it to both scan image files. Feature Extraction (FE) 9.5 used XDR ID to link the pairs of scans together automatically when extracting data. The signal left after all the FE processing steps have been completed, is ProcessedSignal that contains the Multiplicatively Detrended, Background-Subtracted Signal.

first: Sara

last: Pegolo

first: Luca

last: Bargelloni


title: Microarray analysis of serum from esophageal squamous cell carcinoma

city: NAN JING

first: Kai

last: Xie

state: NAN JING

address: JIANG SU

country: China

institute: NAN JING MEDICAL UNIVERSITY

summary: Sample labeling and array hybridization were performed according to the Agilent One-Color Microarray-Based Gene Expression Analysis protocol (Agilent Technology) with minor modifications. Briefly, mRNA was purified from total RNA after removal of rRNA (mRNA-ONLY™ Eukaryotic mRNA Isolation Kit, Epicentre). Then, each sample was amplified and transcribed into fluorescent cRNA along the entire length of the transcripts without 3' bias utilizing a random priming method (Arraystar Flash RNA Labeling Kit, Arraystar). The labeled cRNAs were purified by RNeasy Mini Kit (Qiagen). The concentration and specific activity of the labeled cRNAs (pmol Cy3/μg cRNA) were measured by NanoDrop ND-1000. 1 μg of each labeled cRNA was fragmented by adding 5 μl 10 × Blocking Agent and 1 μl of 25 × Fragmentation Buffer, then heated the mixture at 60°C for 30 min, finally 25 μl 2 × GE Hybridization buffer was added to dilute the labeled cRNA. 50 μl of hybridization solution was dispensed into the gasket slide and assembled to the LncRNA expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven. The hybridized arrays were washed, fixed and scanned with using the Agilent DNA Microarray Scanner (part number G2505C).

overall_design: lncRNA microarray analysison four preoperative ESCC patients and matched normal healthy serum samples

first: K

first: C

last: Zheng

first: L

last: Hu

first: Y

last: Shen


title: Spatially resolved whole transcriptome profiling in human and mouse tissue using Digital Spatial Profiling [human cell pellet array]

status: Public on Mar 01 2022

city: Seattle

first: Stephanie

last: Zimmerman

state: WA

address: 530 Fairview Ave N

institute: NanoString Technologies

summary: Emerging spatial profiling technology has enabled high-plex molecular profiling in biological tissues, preserving the spatial and morphological context of gene or protein expression. Here we describe expanded chemistry for the Digital Spatial Profiling platform to quantify whole transcriptomes in human and mouse tissues using a wide range of spatial profiling strategies and sample types. We designed multiplexed in situ hybridization probe pools targeting the protein-coding genes in the human and mouse transcriptomes, hereafter referred to as the human or mouse Whole Transcriptome Atlas (WTA). We validated the human and mouse WTA assays using cell lines to demonstrate concordance with orthogonal gene expression profiling methods in profiled region sizes ranging from ∼10-500 cells. By benchmarking against bulk RNAseq and single-molecule fluorescence in situ hybridization, we demonstrate robust transcript detection possible down to ∼100 transcripts per region. To assess the performance of WTA across tissue and sample types, we applied WTA to biological questions in cancer, molecular pathology, and developmental biology. We show that spatial profiling with WTA can detect expected spatial gene expression differences between tumor and tumor microenvironment, identify spatial disease-specific heterogeneity in gene expression in histological structures of the human kidney, and comprehensively map transcriptional programs in anatomical substructures of nine organs in the developing mouse embryo. Digital Spatial Profiling technology with the WTA assays provides a flexible method for spatial whole transcriptome profiling applicable to diverse tissue types and biological contexts.

overall_design: This dataset contains RNA expression from 11 human cell lines from an FFPE cell pellet array using the human Whole Transcriptome Atlas for Digital Spatial Profiling


title: Transcriptom of Wild type and Lnc956 knockout mouse embryo

status: Public on Feb 16 2022

city: Kunming

first: Ping

address: Qingsong Road 21

institute: Kunming Institute of Zoology, Chinease Academy of Sciences

summary: Purpose: The goal of this study is to compare gene expression profiling of wild type and Lnc956 knockout mouse embryo.

overall_design: Wild type and lnc956 knockout mouse embryos were dissected and used for RNA extraction.

first: Weidao

last: Zhang

first: Min

last: Tang


title: Large scale transposon co-option in the Caenorhabditis germline regulatory network [RNA-Seq]

status: Public on Dec 26 2021

city: Cambridge

first: Francesco Nicola

last: Carelli

address: Henry Wellcome Building of Cancer and Developmental Biology,  Tennis Court Road

country: United Kingdom

institute: University of Cambridge

summary: The movement of repetitive elements in the germline creates widespread genomic alterations and pressure for resolution. Here we show that the Caenorhabditis clade took advantage of two transposon expansions by integrating hundreds of elements into its germline transcriptional network. We find that about one-third of C. elegans germline-specific promoters have been co-opted from CERP2 and CELE2 MITE elements and are regulated by HIM-17, a THAP domain-containing transcription factor related to a transposase. An ancestral CERP2 expansion took place in the common Caenorhabditis ancestor, concurrently with mutations in HIM-17 fixed by positive selection, whereas CELE2 expanded only in C. elegans. Through comparative analyses in C. briggsae, we find conservation as well as species-specific CERP2 co-option. Our work reveals the emergence of a novel transcriptional network driven by TE co-option and its impact on regulatory evolution.

overall_design: 4 single-end RNA-seq libraries were generated from polyA-selected RNA from C. elegans strains (wt and him-17 mutants).

first: Francesco

middle: N

first: Yan

last: Dong

first: Chiara

last: Cerrato

first: Alex

last: Appert

first: Julie

last: Ahringer


title: Suppressing circ_0008494 inhibits HSC activation by regulating the miR-185-3p/Col1a1 axis

status: Public on Dec 22 2021

city: shanghai

first: Xie

last: qin

address: Minchi first road, pujiang town, minhang district, Shanghai

institute: LC sciences

type: Non-coding RNA profiling by high throughput sequencing

summary: Background: Liver fibrosis is characterized by the extensive deposition of extracellular matrix components, especially collagens. However, effective antifibrotic therapies are still lacking. Recently, circular RNAs (circRNAs) have been a topic of high interest in the field of liver fibrosis.

overall_design: 6 liver fibrosis samples were sequencing

first: Bin-Bin

last: Li

first: Jia-Ming

last: Zhou

first: Yuan-Yuan

last: Luo

first: Ke-Gong

last: Tao

first: Li-Fen

first: Yong

last: Lin

first: Xin

last: Zeng

first: Hong-yu

last: Yu


title: Presurgical Phase IIB Trial of Oral CDB-4124 (TPA) vs. Placebo in Women with Stage 0-II Primary Breast Cancer: PACT-B (Progesterone Antagonists for Cancer Treatment for Breast) Study

status: Public on Feb 19 2022

city: Chicago

first: Seema

middle: Ahsan

last: Khan

state: IL

address: 303 E. Superior Street, 4-111

institute: Northwestern University

summary: Purposes: 1. to identify the genes which were significantly modulated by CDB-4124 (TPA) comapred to placebo treatment. 2. identify which the pathways were functionally impacted by CDB4124.

overall_design:
A total of 60 patients were treated with either placebo control (n=29 ) or CDB-4124 (TPA)(n=31) for 2- 4 weeks prior to surgery. Two FFPE samples per subject (pre-Tx and post-Tx tumor specimen) were subjected to RNA extraction and RNA seq. Pre-Tx sample was a diagnostic core needle biopsy. Post-Tx sample was a surgical specimen. A total of 120 total RNA samples are submitted for RNA seq.
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Artificial Intelligene (AI, www.artificialintelligene.com)

first: Oukseub

last: Lee

first: Megan

last: Sullivan

first: Yanfei

last: Xu

first: Miguel

last: Muzzio

first: Irene

last: Helenowski

first: Ali

last: Shidfar

first: Zexian

first: Hari

last: Singhal

first: Borko

last: Jovanovic

first: Nora

last: Hansen

first: Kevin

last: Bethke

first: Peter

last: Gann

first: William

last: Gradishar

first: Susan

middle: E

last: Clare

middle: A


title: Microarray Analysis of GCNF mutant and wt E9.0 embryos in Mus musculus

status: Public on Feb 01 2022

city: Kansas City

first: Chris

middle: W

last: Seidel

state: MO

department: Genomics

institute: Stowers Institute

summary: Neural crest cells are migratory progenitor cells that contribute to nearly all tissues and organs throughout the body. Their formation, migration and differentiation are regulated by a multitude of signaling pathways, that when disrupted can lead to disorders termed neurocristopathies. While work in avian and amphibian species has revealed essential factors governing the specification and induction of neural crest cells during gastrulation and neurulation in non-mammalian species, their functions do not appear to be conserved in mice, leaving major gaps in our understanding of neural crest cell formation in mammals. Here we describe Germ Cell Nuclear Factor (GCNF/Nr6a1), an orphan nuclear receptor, as a critical regulator of neural crest cell formation in mice. Gcnf null mutant mice, exhibit a major disruption of neural crest cell formation. The purpose of this experiment is to examine gene expression changes in response to Gcnf mutation in E9.0 mouse embryos.

overall_design: Homozygous wt and mutant GCNF mouse embryos at developmental stage E9.0 were isolated in duplicate for RNA extraction and hybridization on Affymetrix microarrays.

first: Paul

last: Trainor


title: The Trifecta study: comparing plasma donor-derived cell-free DNA levels with the molecular phenotype of kidney transplant biopsies

status: Public on Dec 23 2021

city: Edmonton

first: Jessica

last: Chang

address: 250 Heritage Medical Research Centre

department: Medicine

country: Canada

institute: University of Alberta

summary: Background. The Trifecta study (ClinicalTrials.gov #NCT04239703) is a prospective investigator-initiated trial to evaluate the relationship between plasma %dd-cfDNA at the time of indication biopsy and the molecular phenotype of the biopsy.   Results. Median time of biopsy post-transplant was 455 days (5 days - 32 years), with case-mix similar to previous studies: 180 (60%) no rejection, 89 (30%) antibody-mediated rejection (ABMR), and 31 (10%) T cell-mediated rejection (TCMR) and mixed. In genome-wide mRNA measurements, all 20 top probesets correlating with %dd-cfDNA were previously annotated for association with ABMR and all rejection, either NK cell-expressed (e.g. GNLY, CCL4, TRDC, and S1PR5) or IFNG-inducible (e.g. PLA1A, IDO1, CXCL11, and WARS). Among gene set and classifier scores, %dd-cfDNA correlated very strongly with ABMR and all rejection, reasonably strongly with active TCMR, and weakly with inactive TCMR, kidney injury and atrophy-fibrosis. %dd-cfDNA was highest in active ABMR, mixed, and active TCMR but lower in late-stage ABMR and less active TCMR. By multivariate random forests and logistic regression, molecular rejection variables predicted %dd-cfDNA better than histologic variables.  Conclusions. %dd-cfDNA at time of indication biopsy strongly correlates with active molecular rejection and has potential to reduce unnecessary biopsies.

overall_design: Methods. We compared genome-wide gene expression (microarrays) in 300 biopsies to plasma %dd-cfDNA drawn at time of biopsy, from 19 centers in Europe and North America. We assessed rejection using the microarray-based Molecular Microscope® Diagnostic System (MMDx) using automatically assigned rejection archetypes (AA) and molecular report signouts, plus histology assessments by the center following current Banff guidelines.


title: Impairment of RNA polymerase I elongation rate induces defects in ribosomal RNA processing and ribosome biogenesis

status: Public on Feb 07 2022

city: Birmingham

first: Abigail

last: Huffines

address: 720 20th St S

institute: University of Alabama at Birmingham

summary: The objective of this study was to investigate the relationship between Pol I transcription elongation rate and ribosomal RNA processing in vivo by using a yeast strain containing a mutation in Pol I. This mutation, rpa190-F1205H, has been previously characterized to have a reduced elongation rate in vitro. Here, we used native elongating transcript sequencing (NET-seq) to determine the effect of this mutation on Pol I occupancy in vivo. Our findings demonstrate that this mutation induces increased Pol I pausing, especially in the first half of the 35S gene, and causes sequence-specific changes in Pol I occupancy.

overall_design: Triplicate NET-seq libraries of wild-type (WT) yeast and rpa190-F1205H yeast were prepared and analyzed.

middle: K

first: Krysta

middle: L

last: Engel

first: David

last: Schneider


title: Large scale transposon co-option in the Caenorhabditis germline regulatory network [longCapSeq]

first: Francesco Nicola

last: Carelli

overall_design: 4 paired-end RNA-seq libraries were generated from chromatin-associated long RNA from C. briggsae strains (wt and glp-1 mutants)

first: Francesco

first: Yan

last: Dong

first: Chiara

last: Cerrato

first: Alex

last: Appert

first: Julie

last: Ahringer


title: Large scale transposon co-option in the Caenorhabditis germline regulatory network [ChIP-Seq]

type: Genome binding/occupancy profiling by high throughput sequencing

overall_design: 2 single-end ChIP-seq libraries were generated from wild-type (N2), young adult C. elegans fixed nuclei (EGS+formaldehyde).


title: Activation of STAT3 through SRC and EGFR Drives Resistance to a Mitotic Kinesin Inhibitor in Glioblastoma

status: Public on Jan 09 2022

city: New York

first: Michael

middle: G

last: Argenziano

state: New York

address: 1130 St. Nicholas Ave

institute: Columbia University Vagelos College of Physicians and Surgeons

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

overall_design: Refer to individual Series


title: Many heart transplant biopsies currently diagnosed as no rejection have mild molecular antibody-mediated rejection-related changes

status: Public on Dec 28 2021

summary: Previous studies of rejection-associated transcript expression in heart transplant biopsies identified not only rejection but a group of early biopsies with injury but no rejection. The present analysis used an expanded population of biopsies to explore parenchymal injury in all biopsies, with or without rejection, and its relationship to function, and outcome.   Archetypal analysis defined five injury clusters: no-injury (N=376); mild (N=526); moderate (N=110); severe (N=87); and late (N=221). The late group, 62% of which had no rejection, had molecular characteristics associated with atrophy-fibrosis, depressed LVEF, and increased graft loss independent of rejection status. In random forest analysis, low LVEF was more strongly associated with injury scores than with rejection scores. Three-year graft failure was best predicted using a combination of injury and rejection scores.  In heart transplant biopsies, injury-related molecular scores correlate with dysfunction and risk of failure and identify an important new group of late heart transplants, many with no rejection, that have impaired function and a high risk of graft loss. (ClinicalTrials.gov #NCT02670408).

overall_design: We analyzed 1320 prospectively collected biopsies (645 patients) from 13 international centers in the INTERHEART study. Molecular injury was evaluated with archetypal analysis, using several injury-associated transcript set scores as inputs.


title: Outcomes of intrahepatic cholangiocarcinoma specific CRISPR screening in vivo [mouse exome]

status: Public on Dec 15 2021

city: Edinburgh

first: Luke

last: Boulter

address: IGC

institute: MRC Human Genetics Unit

summary: We report the genertion of a gRNA library that induces mutations often found in intrahepatic cholangiocarcinoma. We use this to screen in vivo mutations that when lost act synergistically with mutant Ras. Specifically KrasG12D and NrasG12V. We have included exome sequencing of these tumours in this data set.

overall_design: Whole exome from mouse intrahepatic cholangiocarcinoma containing a range of mutations


title: Large scale transposon co-option in the Caenorhabditis germline regulatory network

sample_organism:
Caenorhabditis briggsae
Caenorhabditis elegans


title: Activation of STAT3 through SRC and EGFR Drives Resistance to a Mitotic Kinesin Inhibitor in Glioblastoma [scRNA-seq]

summary: Inhibitors of the mitotic kinesin Kif11 are anti-mitotics that, unlike vinca alkaloids or taxanes, do not disrupt microtubules and are not neurotoxic. However, development of resistance has limited their clinical utility. While resistance to Kif11 inhibitors in other cell types is due to mechanisms that prevent these drugs from disrupting mitosis, we find that in glioblastoma (GBM), resistance to the Kif11 inhibitor ispinesib works instead through suppression of apoptosis driven by activation of STAT3. This form of resistance requires dual phosphorylation of STAT3 residues Y705 and S727, mediated by SRC and EGFR, respectively. Simultaneously inhibiting SRC and EGFR reverses this resistance, and combined targeting of these two kinases in vivo with clinically available inhibitors is synergistic and significantly prolongs survival in ispinesib-treated GBM-bearing mice. We thus identify a translationally actionable approach to overcoming Kif11 inhibitor resistance that may work to block STAT3-driven resistance against other anti-cancer therapies as well.

overall_design: scRNAseq on 2 samples: one naïve murine glioma cell line, one ispinesib resistant murine glioma cell line.

first: Steven

middle: S

last: Rosenfeld

first: Peter

last: Sims

last: Canoll

first: Athanassios

last: Dovas


title: Negative autoregulation mitigates collateral RNase activity of repeat-targeting CRISPR-Cas13d in mammalian cells

city: Gainesville

first: Eric

middle: T

last: Wang

state: FL

address: 2033 Mowry Road

department: Molecular Genetics and Microbiology

institute: University of Florida

summary: Cas13 is a unique family of CRISPR endonucleases exhibiting programmable binding and cleavage of RNAs and is a strong candidate for eukaryotic RNA knockdown in the laboratory and the clinic. However, sequence-specific binding of Cas13 to the target RNA unleashes non-specific bystander RNA cleavage, or collateral activity, which may confound knockdown experiments and raises concerns for therapeutic applications. Although conserved across orthologs and robust in cell-free and bacterial environments, the extent of collateral activity in mammalian cells remains disputed. Here, we investigate Cas13d collateral activity in the context of an RNA-targeting therapy for myotonic dystrophy type 1, a disease caused by a transcribed long CTG repeat expansion. We find that when targeting CUGn RNA in HeLa and other cell lines, Cas13d depletes endogenous and transgenic RNAs, interferes with critical cellular processes, and activates stress response and apoptosis pathways. We also observe collateral effects when targeting other repetitive and unique transgenic sequences, and we provide evidence for collateral activity when targeting highly expressed endogenous transcripts. To minimize collateral activity for repeat-targeting Cas13d therapeutics, we introduce gRNA excision for negative-autoregulatory optimization (GENO), a simple strategy that leverages crRNA processing to control Cas13d expression and is easily integrated into an AAV gene therapy. We argue that thorough assessment of collateral activity is necessary when applying Cas13d in mammalian cells and that implementation of GENO illustrates the advantages of compact and universally robust regulatory systems for Cas-based gene therapies.

overall_design: RNA-seq was performed to screen for transcriptomic off-targets of repeat-targeting approaches (Cas13d, dCas9, and shRNA). CUG-targeted systems and non-targeting controls (no gRNA or non-targeting shRNA) were transfected in triplicate into HeLa cells. CUG480 target plasmid was omitted to enrich for off-target events. After 68 hr incubation, RNA was extracted with 300 uL TRIzol and purified using the Direct-zol RNA Miniprep Kit. Ribosomal RNA was depleted from 300 ng total RNA using the NEBNext rRNA Depletion Kit (NEB, #E6310L), and libraries were generated for next-generation sequencing using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (NEB, #E7760L) according to the manufacturer protocol. Libraries were multiplexed with Illumina i5 and i7 barcoding primers, pooled at 4 nM, and sequenced in a 2x76 bp paired-end format on an Illumina NextSeq 500. >14 million paired end reads were sequenced per library and recorded in FASTQ format.

first: Chase

middle: P

last: Kelley

first: Maja

middle: C

last: Haerle


title: Expression data from MDA-MB-231 breast cancer cell lines with DAXX Knockdown

status: Public on Dec 11 2021

first: Daiqing

last: Liao

address: 2033 Mowry Rd

department: Anatomy and Cell Biology

summary: DAXX, originally discovered as a context-dependent regulator of cell death or survival. Emerging evidence suggests that DAXX has an oncogenic role in diverse cancer types. However, the molecular mechanisms underlying DAXX's oncogenic function remain to be defined. We used Affymetrix Human Transcriptome Array 2.0 (HTA-2_0) to analyze genes that were up or downregulated upon DAXX knockdown.

overall_design: The shRNA that target DAXX gene was transfected in human triple negative brest cancer MDA-MB-231 cell line by lentiviral particles and selected stable DAXX knockdown MDA-MB-231 cells. Scrambled control shRNA-transfected MDA-MB-231 cells were applying as control. Both stable DAXX knockdown and control MDA-MB-231 cells were seeded with 2 x 10^5 cells at 10 cm culture plate. Stable expression of shRNA was established through lentiviral transduction of MDA-MB-231 cell line and puromycin (2 µg/mL) selection. Three biological replicates were applied.

first: Iqbal

last: Mahmud


title: To explore the changes in the genes expression profile  between brain and skull after ischemic injury

status: Public on Dec 20 2021

city: Philadelphia

first: Chunjie

last: Liu

state: PENNSYLVANIA

address: Colket Translational Research Building, 3501 Civic Center BLVD

department: Center for Computational and Genomic Medicine

institute: Children's Hospital of Philadelphia

summary: We longitudinally monitor the cortical/calvarial vascular injury and regeneration processes after ischemic injury in awake mice. Results show that calvarial blood vessels recover earlier than the cortex. Further, by transcriptome analysis, we finds gene expression patterns and immune cells abundances exist great differences between brain and skull after ischemic injury, which may be one of the causes for the time lag between their vascular recovery. These findings bring great enlightenment to the vascular regeneration and reconstruction.

overall_design: Transcriptome changes in cortex and skull at representative time points after ischemic injury were analyzed. The mouse brain and skull tissue contains ischemic and peri-ischemic regions were harvested for RNA sequencing.

first: Chao

last: Zhang

first: Chun-Jie

first: Xue

last: Li

first: Wei

last: Feng


title: Single cell RNA sequencing analysis of the center region of healing skin at wound day 11  in wild type mouse

status: Public on Dec 06 2021

city: Baltimore

first: Sam

last: Lee

state: MD

address: 1550 Orleans Street

institute: Johns Hopkins University School of Medicine

summary: We have applied single_x0002_cell RNA sequencing analysis to investigate keratinocyte populations and their specific gene expression on the day of would closure (post wounding day 11-12) at the center of re-epithelialized mouse wound site (1 cm2 of whole skin was removed, multiple pooled mice, n = 10).  Viable cells from single cell suspension were selected by sorting and single cell library was prepared by 10x Genomics and sequenced by Illumina NovaSeq 6000. Downstream bioinformatic analysis of single cell RNAseq was performed by using Seurat package in R. Clusters of cells were denoted according to their identity. Keratinocytes were identified from the entire sample of cells using known keratinocyte markers. Specific gene expression over the keratinocytes were visulaized by heatmap. Overlap expression of Cd14, Tlr3, Krt15, Krt19 and Rarα at the day of wound closure supports morphogenesis.

overall_design: Single sample was analyzed to investigation of keratinocyte gene expression on the day of would closure (post wounding day 11-12) at the center of re-epithelialized mouse wound site

first: Gaofeng


title: Large scale transposon co-option in the Caenorhabditis germline regulatory network [ATAC-seq]

first: Francesco Nicola

last: Carelli

overall_design: 4 single-end ATAC-seq libraries from C. briggsae strains (wt and glp-1 mutant)

first: Francesco

first: Yan

last: Dong

first: Chiara

last: Cerrato

first: Alex

last: Appert

first: Julie

last: Ahringer


title: Urine single cell RNA-sequencing in focal segmental glomerulosclerosis reveals inflammatory signatures in immune cells and podocytes

status: Public on Dec 07 2021

city: Bethesda

first: Jeffrey

middle: B

last: Kopp

state: Maryland

address: 10 Center Drive, 3N116

department: NIDDK - KDB

institute: National Institutes of Health

summary: The diagnosis of focal segmental glomerulosclerosis (FSGS) requires a renal biopsy  which is invasive and can be problematic in children and in some adults. We used single cell RNA-sequencing to explore disease-related cellular signatures in 17 urine samples from 12 FSGS subjects. We identified immune cells in urine  predominantly monocytes  and renal epithelial cells  including podocytes. Analysis revealed M1 and M2 monocyte subsets  and podocytes showing increased expression of genes involved in epithelial-to-mesenchymal transition (EMT). We confirmed M1 and M2 gene signatures using published monocyte/macrophage data from lupus nephritis and cancer. Using renal transcriptomic data from the Nephrotic Syndrome Study Network (NEPTUNE)  we found that urine immune and EMT signature genes also showed higher expression in FSGS biopsies compared to minimal change disease biopsies. These results suggest that urine cell profiling may serve as a diagnostic and prognostic tool in nephrotic syndrome and may assist in identifying novel biomarkers and developing personalized therapeutic strategies.

overall_design:
single cell RNAseq of urine cells from 12 subjects with focal segmental glomerulosclerosis
Contributor: The Nephrotic Syndrome Study Network (NEPTUNE)
Contributor: The Accelerating Medicines Partnership in Rheumatoid Arthritis and Systemic Lupus Erythematosus (AMP RA/SLE) consortium

first: Khun

middle: Z

last: Latt

first: Jurgen

last: Heymann

first: Joseph

middle: H

last: Jessee

first: Avi

last: Rosenberg

first: Celine

last: Berthier

first: Sean

last: Eddy

first: Teruhiko

last: Yoshida

first: Yongmei

last: Zhao

first: Vicky

last: Chen

first: George

last: Nelson

first: Margaret

last: Cam

first: Parimal

last: Kumar

first: Monika

last: Mehta

first: Michael

last: Kelly

first: Matthias

last: Kretzler

first: Patricio

last: Ray

first: Marva

last: Moxey-Mims

first: Gregory

last: Gorman

first: Brent

last: Lechner

first: Renu

last: Regunathan-Shenk

first: Dominic

last: Raj

first: Katalin

last: Susztak

first: Cheryl

last: Winkler

first: Arnon

last: Arazi


title: TETs during directed differentiation of human embryonic stem cells to cardiomyocyte

city: Beijing

first: Heng

last: Pan

address: 49 North Garden Rd., Haidian District

institute: Peking University Third Hospital

type: Methylation profiling by high throughput sequencing


title: DAXX genome binding/occupancy profiling by high throughput ChIP sequencing

first: Daiqing

last: Liao

summary:
Purpose: The goal of this study is to compare DAXX genome binding/occupancy by high throughput ChIP sequencing in control, wild type DAXX overexpressd cells (WT DAXX OE), and wild type DAXX SUMO interacting motif mutant overexpressd cells (DSM OE).
Methods: The panel of MDA_MB-231 derived cell lines (control, WT DAXX OE, DSM OE) were cultured in complete DMEM. At about 90% confluency, the cells were crosslinked by adding 37 % formaldehyde to the final concentration of 1% for 10 min at room temperature. Crosslinking was stopped by adding glycine to the final concentration of 125 mM. Cells were lifted, washed with cold PBS, and pelleted by centrifugation. The cells were resuspended in a swelling buffer in the presence of the protease inhibitor cocktail (Sigma) and then pelleted and resuspended in the SDS lysis buffer. The lysates were transferred to a Covaris microTUBE and sonicated with an E220 Covaris Ultrasonicator. Chromatin fragmentation (~500 bps) was verified through agarose gel electrophoresis. The fragmented chromatins were diluted and incubated with a control IgG and the DAXX mAb (5G11) along with protein A/G magnetic beads. The beads were washed sequentially with a low salt buffer, high salt buffer, LiCl buffer, and TE buffer (twice). The immunoprecipitated chromatins were eluted at 65 °C for 15 min, and the eluted chromatins were subjected to proteinase K digestion at 65 °C for 3 h. The DNAs were recovered through a Qiagen mini-prep column. The immunoprecipitated DNAs were used for qPCR and library construction and high throughput sequencing using an Illumina Hi-Seq 2500 sequencer.
Results: We surveyed genome-wide occupancy of DAXX using ChIP-seq technology. Overexpression of WT DAXX  increased DAXX?s chromatin association specifically with lipogenic pathway genes where DSM OE reverse.
Conclusions: Our study represents the first detailed analysis of DAXX occupancy in lipoegenic gene transciption.

overall_design: Genome sequencing of MDA_MB-231 derived cell lines (control, WT DAXX OE, and DSM OE)

first: Iqbal

last: Mahmud


title: Outcomes of intrahepatic cholangiocarcinoma specific CRISPR screening in vivo

first: Luke

last: Boulter


title: Integration of a scalable parallel reporter genes for qRT-PCR based enhancer screening with CRISPR dCas9 activation and supershift EMSA identifies impaired BACH1 repression of ST8SIA1 to contribute to the increased risk for periodontitis in smokers

status: Public on Jan 01 2022

city: Berlin

first: January

last: Weiner

address: Charitéplatz 1

department: CUBI

country: Germany

institute: Berlin Institute of Health, Charité Medical University of Berlin

summary: Genome-wide association studies identified many loci associated with human diseases, but assigning causal alleles remains difficult. Likewise, generating biological meaning underlying a statistical association is challenging. To address these problems, we developed a scalable reporter gene system for the simultaneous transfection and parallel quantification of enhancer activity. Additionally, we characterized a genetic association at the gene ST8SIA1 that increases the risk for periodontitis in smokers. We cloned unique DNA barcode sequences to a reporter gene. Transcript levels of the barcodes and firefly luminescence of the reporter was compared in HeLa cells for several known enhancers. We mapped the associated SNPs to DNA elements with predictive features of regulatory function on gene expression from ENCODE. We tested whether associated alleles changed predicted transcription factor binding sites  and proved transcription factor binding using antibody electrophoretic mobility shift assays. Using CRISPR dCas9 activation, we pinpointed the target gene of the association. Using RNA-Sequencing, we identified gene sets that are associated with ST8SIA1. Two associated repressor elements bind the transcription factor BACH1 with the putative causative variant rs2012722 decreasing BACH1 binding by 40%. ST8SIA1 is the likely target gene of the association, regulating the cell cycle arrest and integrin cell surface signaling. We found the periodontitis risk gene ABCA1 as the most significantly up-regulated gene. In conclusion, we developed a low cost, easy to use parallel reporter gene system that facilitates enhancer screening, and described an experimental pipeline for straightforward identification and characterization of causal variants and of their target genes.

overall_design: 500-1000 ng of total RNA of the 6 transfected independent HeLa cell cultures was sequenced with 16 million reads (75bp single end) on a NextSeq 500 using the NextSeq 500/550 High Output Kit v2.5 (75 Cycles). RNA-Seq was performed at the Berlin Institute of Health Core Facility Genomics.

first: Avneesh

last: Chopra

first: Ricarda

last: Mueller

first: Jennifer

last: Rosowski

first: Henrik

last: Dommisch

first: Elisabeth

last: Grohmann

first: Arne

last: Schaefer


title: Intratumor genomic heterogeneity in stage II colon cancer

status: Public on Dec 01 2021

city: Barcelona

first: Sara

last: Lahoz

address: Carrer del Rosselló, 149

country: Spain

institute: Institut D'Investigacions Sanitàries August Pi i Sunyer

type:
Genome variation profiling by SNP array
Other


title: Outcomes of intrahepatic cholangiocarcinoma specific CRISPR screening in vivo [RNA-seq]

summary: We report the genertion of a gRNA library that induces mutations often found in intrahepatic cholangiocarcinoma. We use this to screen in vivo mutations that when lost act synergistically with mutant Ras. Specifically KrasG12D and NrasG12V. We have included RNA sequencing of these tumours in this data set.

overall_design: RNAseq from mouse intrahepatic cholangiocarcinoma containing a range of mutations


title: The astroglial and stem cell functions of adult rat folliculostellate cells

status: Public on Dec 12 2021

first: Patrick

middle: Allen

last: Fletcher

address: 9000 Rockville Pike

institute: NIDDK

summary: The mammalian pituitary gland is a complex organ consisting of hormone-producing cells, anterior lobe folliculostellate cells (FSCs), posterior lobe pituicytes, vascular pericytes and endothelial cells, and Sox2-expressing stem cells. We present single-cell RNA sequencing and immunohistofluorescence analyses of pituitary cells of adult female rats with a focus on the transcriptomic profiles of nonhormonal cell types. Samples obtained from whole pituitaries and separated anterior and posterior lobe cells contained all expected pituitary resident cell types and lobe-specific vascular cell subpopulations. FSCs and pituicytes expressed S100B, ALDOC, EAAT1, ALDH1A1, and VIM genes and proteins, as well as other astroglial marker genes, some common and some cell type-specific. We also found that the SOX2 gene and protein were expressed in ~15% of pituitary cells, including FSCs, pituicytes, and a fraction of hormone-producing cells, arguing against its stem cell specificity. FSCs comprised two Sox2-expressing subclusters; FS1 contained more cells but lower genetic diversity, while FS2 contained proliferative cells, shared genes with hormone-producing cells, and expressed genes consistent with stem cell niche formation, regulation of cell proliferation and stem cell pluripotency, including the Hippo and Wnt pathways. FS1 cells were randomly distributed in the anterior and intermediate lobes, while FS2 cells were localized exclusively in the marginal zone between the anterior and intermediate lobes. These data indicate the identity of the FSCs as anterior pituitary-specific astroglia, with FS1 cells representing differentiated cells equipped for classical FSC roles and FS2 cells exhibiting additional stem cell-like features.

overall_design: Single-cell transcriptome analysis of anterior and posterior pituitary cells from adult female rats


title: MiR-484 Knockdown Ameliorates Liver Fibrosis by Blocking Wnt/β-catenin Pathway via Tar-geting HIPK1

status: Public on Dec 09 2021

first: Xie

last: qin

summary: Liver fibrosis is a common pathological process of various chronic liver diseases that can develop into liver cancer. MicroRNAs (miRNAs) are a king of non-coding RNA which are closely related to liver diseases. Thus, this research hope to explore the effect of miR-484 on liver fibrosis and reveal its mechanism. The miRNAs profiles were screened by microRNA sequencing and the location of miR-484 was identified by fluorescence in situ hybridization (FISH) in human liver fibrotic tissues. MiR-484 expression was detected by qRT-PCR in rat primary hepatic stellate cells (HSCs). Bioinformatics analysis and dual-luciferase reporter assay were performed to determine the target gene of miR-484. Liver fibrosis specific signatures were analyzed by qRT-PCR and western blot after miR-484 mimic/inhibitor transfection. The cell apoptosis was detected by Annexin V-FITC/PI double staining. The effect of miR-484 silencing on fibrosis in vivo was investigated in thioacetamide (TAA) induced mice model using the adeno-associated virus carrying miR-484 inhibitor. Enrichment of miR-484 was observed in human liver fibrosis tissues and activated rat primary HSCs.  FISH showed that miR-484 was prominently located at fibrotic region and the cytoplasm of HSCs in human liver tissues. Dual-luciferase reporter assay verified that the homeodomain-interacting protein kinases 1 (HIPK1) was the direct target of miR-484. After transfecting miR-484 inhibitor into HSC-T6, HIPK1 were significantly up-regulated, and α-SMA, col1a1, Wnt-3a, Wnt-5a, β-catenin and p-β-catenin were down-regulated, suggesting the restrain effect of miR-484 knockdown on HSCs activation. Conversely, the results were opposite with miR-484 mimic transfection. In addition, the apoptosis of HSC-T6 altered significantly after miR-484 modulation. Moreover, adeno-associated virus carrying miR-484 inhibitor alleviated mice liver fibrosis induced by TAA. In conclusion, miR-484 knockdown ameliorates liver fibrosis by promoting the apoptosis and suppressing HSCs activation via blocking Wnt/β-catenin signaling pathway. MiR-484 and its downstream gene HIPK1 might be selected as novel therapeutic targets of liver fibrosis.

overall_design: six human liver fibrotic tissues (provided by the Second Affiliated Hospital of Naval Medical Universi-ty, Shanghai, China) were gathered and sent for miRNA sequencing (Lc-bio technologies, Hangzhou, China).

first: Bin-Bin

last: LI

first: Yuan-Yuan

last: LUO

first: Ke-Gong

last: TAO

first: Li-Fen

last: ZHANG

first: Hong-Yu

last: YU

first: Yong

last: LIN

first: Xin

last: ZENG


title: Dual RNA-seq on pneumococcal infection to zebrafish embryo model

status: Public on Dec 14 2021

city: Groningen

first: Rieza

last: Aprianto

state: Groningen

address: Nijenborg 7

department: Molecular Genetics Department

country: Netherlands

institute: University of Groningen

summary: Zebrafish embryo has been emerging as an interesting model of infection due to their fecundity, transparency and availability of genetic tools. Streptococcus pneumoniae, the pneumococcus, is the main etiological agent of pneumonia, sepsis and meningitis. The bacteria expresses a very important virulence factor, pneumolysin able to form pores on cholesterol-based membranes and to activate innate immune system. Here, we exploited the recently described dual RNA-seq to simultaneously measure genome-wide expression of host and pathogen eight hours into infection. Functional enrichment analysis showed certain pathways such as autophagy and apoptosis being activated in the host while stress responses including pneumococcal competence. The study is the first to describe dual RNA-seq application in whole organism sequencing in infection model.

sample_organism:
Streptococcus pneumoniae
Danio rerio

platform_organism:
Streptococcus pneumoniae
Danio rerio

overall_design: Six samples are analysed, categorized into two conditions: infection with ply mutant and complemented strain

middle: R

first: Jan-Willem

middle: J

last: Veening

first: Kinki

last: Jim

first: Astrid

last: van der Sar

first: Diederik

middle: D

last: van de Beek

first: Christina

last: Vandenbroucke-Grauls

first: Wilbert

last: Bitter


title: Probiotic Lacticaseibacillus rhamnosus HA-114 suppresses age-dependent neurodegeneration via mitochondrial beta-oxidation

status: Public on Dec 05 2021

city: Montreal

first: J Alex

last: Parker

state: Quebec

address: 900 rue St-Denis

institute: CRCHUM, Université de Montréal

summary: The human microbiota is believed to influence health. Microbiome dysbiosis may be linked to neurological conditions like Alzheimer’s disease (AD), amyotrophic lateral sclerosis (ALS) and Huntington’s disease (HD). We report the ability of a probiotic bacterial strain in halting neurodegeneration phenotypes. We show that Lacticaseibacillus rhamnosus HA-114 is neuroprotective in C. elegans models of ALS and HD. Our results show that neuroprotection from L. rhamnosus HA-114 is unique from other L. rhamnosus strains, and resides in its fatty acid content. Neuroprotection by L. rhamnosus HA-114 requires acdh-1/ACADSB, kat-1/ACAT1 and elo-6/ELOVL3/6, which are key fatty acid metabolism and mitochondrial b-oxidation genes. Our data suggest that disrupted lipid metabolism contributes to neurodegeneration and that dietary intervention with L. rhamnosus HA-114 restores lipid homeostasis and energy balance through mitochondrial b-oxidation. L. rhamnosus HA-114 is suitable for human consumption opening the possibility of modifying disease progression by dietary intervention.

overall_design: N2 worms (WT Caenorhabditis elegans) were grown on  OP50, Bifidobacterium animalis subsp. lactis B94 or Lacticaseibacillus rhamnosus HA-114. Two or three samples per condition were used for this experiment. Sample were biological and technical replicates.

first: Audrey

last: Labarre

first: J


title: Next Generation Sequencing reveals transcriptome profiling and genome-wide bindings of KLF4 and P53  in Wild Type and Lnc956 loss mouse embryonic stem cells

status: Public on Nov 19 2021

city: Kunming

first: Ping

last: Zheng

address: Qingsong Road 21

institute: Kunming Institute of Zoology, Chinease Academy of Sciences

summary: Purpose: The goals of this study are to compare the transcriptome profiling between wild type (WT) and Lnc956 loss mouse embryonic stem cells (mESCs) and to discover binding sites of KLF4 and P53 in  WT and Lnc956 loss mESCs.

overall_design: mRNA profiles of WT mESCs and Lnc956 loss mESCs; Examination of KLF4 and P53 binding sites in WT and Lnc956 loss mouse embryonic stem cells.

first: Huaixiao

last: Ma

first: Yuqi

last: Ning

first: Lin

last: Wang


title: Notch1-CD22-Dependent Immune Dysregulation in the SARS-CoV2-Associated Multisystem Inflammatory Syndrome in Children

city: Boston

first: KLaus

last: Schmitz-Abe

state: MA

address: 300 Longwood Avenue

department: Asthma, Allergy and Immunology

institute: Children's Hospital Boston

summary: Multisystem inflammatory syndrome in children (MIS-C) evolves in some pediatric patients following acute infection with SARS-CoV-2 by hitherto unknown mechanisms. Whereas acute-COVID-19 severity and outcome were previously correlated with Notch4 expression on regulatory T (Treg) cells, here we show that the Treg cells in MIS-C additionally upregulated Notch1 expression. Genetic analysis revealed that MIS-C patients were enriched in rare deleterious variant impacting inflammation and autoimmunity pathways. Several variants mapped to Notch signaling-related genes including a loss of function mutation in the negative Notch1 regulator NUMB . Notch1 signaling in Treg cells induced CD22, leading to their destabilization in an mTORC1 dependent manner and to the promotion of systemic inflammation in proxy mouse models. These results establish the Notch1-CD22 axis as an immune dysregulatory mechanism critically involved in MIS-C and point to distinct immune checkpoints controlled by individual Treg cell Notch receptors that shape the inflammatory outcome in SARS-CoV-2 infection.

overall_design: WT Tregs or Tregs CD22positive from Foxp3EGFPCre  or  Foxp3EGFPCre R26 N1c  mice were cell sorted with a Sony sorter and Snap Freeze before RNA preparation

first: Mehdi

last: Benamar

first: Klaus

first: Talal

last: Chatila


title: Expression data of microRNAs from human lung cancer at different clinical types

status: Public on Nov 12 2021

city: Tangshan

first: Guogui

last: Sun

state: China

address: 73 Jian-She Road, Tangshan, Hebei, P.R. China

institute: North China University of Science and Technology Affiliated Hospital

summary: Brain metastasis is a major complication of lung cancer. An investigation of the pathogenic mechanisms of brain metastasis, as well as the identification of appropriate molecular markers, is necessary. We used microarrays to determine the expression patterns of microRNAs in lung cancer tissue with or without brain metastasis and to investigate the biological role of these miRNAs during tumorigenesis.

overall_design: The different types human lung cancer tissue were selected for RNA extraction and hybridization on Affymetrix microarrays, including non first brain metastasis (11 cases),  first brain metastasis (5 cases), non metastasis (8 cases) and only extracranial metastasis (21 cases).


title: Differential neoadjuvant chemotherapy response in urothelial carcinoma molecular subtypes

city: Lund

first: Gottfrid

last: Sjodahl

address: Medicon Village

department: Department of Translational Medicine

country: Sweden

institute: Lund Univeristy

summary: For patients with muscle-invasive bladder cancer, there are no biomarkers in clinical use that can identify patients that are sensitive or resistant to neoadjuvant chemotherapy. This study investigates how molecular subtypes impact pathological response and survival in 149 patients receiving preoperative cis-platin based chemotherapy by tumor classification using transcriptomic profiling and a 13-marker immunostaining panel. Furthermore, we explored to what extent gene expression signatures can predict chemotherapy response beyond molecular subtypes. We observed improved pathological response rates and survival outcomes for patients presenting with genomically unstable (GU) and urothelial-like (Uro) subtypes compared to the basal-squamous (BASQ) subtype following neoadjuvant chemotherapy and radical cystectomy. Also, SPP1, coding for osteopontin, displayed a clear subtype-dependent effect on chemotherapy response, confirmed at the protein level. Based on our findings, we hypothesize that urothelial cancer of the luminal-like GU- and Uro-subtypes are more responsive to cisplatin-based chemotherapy which may influence patient selection pending further research.

overall_design:
Retrospective cohort study aiming to determine the association between molecular subtypes and neoadjuvant chemotherapy response in muscle-invasive bladder cancer.

Please note that the 'normalized_by_gene.txt' was updated on Feb 28, 2022 to include the corresponding sample titles as the data column headers.

last: Sjödahl

first: Johan

last: Abrahamsson

first: Karin

last: Holmsten

first: Carina

last: Bernardo

first: Gunilla

last: Chebil

first: Pontus

last: Eriksson

first: Iva

last: Johansson

first: Petter

last: Kollberg

first: Claes

last: Lindh

first: Kristina

last: Lövgren

first: Nour-al-Dain

last: Marzouka

first: Hans

last: Olsson

first: Mattias

last: Höglund

first: Anders

last: Ullén

first: Fredrik

last: Liedberg


title: Transcriptional alterations in Type II Rothmund-Thompson Syndrome

city: Houston

first: An

last: Xu

state: TX

institute: UThealth Houston

summary: We generated induced pluripotent stem cell lines (iPSCs) from paired RTS patient/ parental control fibroblasts. These iPSCs were differentiated to mesenchymal stem cells (MSCs), and then to osteoblasts. We next compared the transcriptional profiles of MSCs, pre-osteoblasts, and osteoblasts.

overall_design: Comparison of Type II RTS (RECQL4 mut/mut) iPCS derived mesenchymal stem celks, early osteoblasts, and late osteoblasts to Family controls (RECQL4wt/mut).

first: Brittany

middle: E

last: Jewell

first: Dung-Fang

last: Lee


title: A longitudinal RNA sequencing dataset in primary human fibroblasts with mitochondrial genetic, metabolic, and endocrine perturbations

first: Gabriel

last: Sturm

state: NY

address: 40 Haven Avenue

department: Psychiatry

institute: Columbia University

summary: The current RNA sequencing dataset (n=345) provides a transcriptional map of cultured primary human fibroblasts (in vitro) aged through the cellular lifespan. The dataset was generated from paired-end (150bp) Illumina HiSeq sequencing, with an average coverage of 38.3million reads per sample. These data contain gene expression trajectories collected at high temporal frequency (every 11 days, ~7timepoints per donor) across the replicative lifespan of primary human fibroblast cell lines from n=4 healthy male and female donors, and n=3 donors with lifespan-altering mitochondrial disease caused by mutations in the SURF1 gene. The dataset can be integrated with other measures collected in parallel along the lifespan, including cytological (cell size, morphology), bioenergetic (energy expenditure, derived ATP synthesis rates from Seahorse), epigenetic (DNA methylation, GSE179847), secreted proteins, telomere length, and whole-genome sequencing (WGS) data. This dataset also includes experimental manipulations with treatments targeting oxidative phosphorylation (OXPHOS) and glycolysis, and glucocorticoid signaling, providing an opportunity to examine the influence of stress and bioenergetics on human aging biology. All processed data can be accessed and browsed at our webtool: https://columbia-picard.shinyapps.io/shinyapp-Lifespan_Study/

overall_design: Primary human fibroblasts are cultured and passaged over ~250 days of replicative lifespan in various treatment conditions.

first: Balaji

last: Santhanam

first: Martin

last: Picard


title: RNA promotes the formation of spatial compartments in the nucleus

status: Public on Nov 04 2021

city: Princeton

first: Sofia

middle: Agustina

last: Quinodoz

state: NJ

address: 25 William St

department: CBE

institute: Princeton University

summary: The nucleus is a highly organized arrangement of RNA, DNA, and protein molecules that are compartmentalized within three-dimensional (3D) structures involved in shared functional and regulatory processes. Although RNA has long been proposed to play a global role in organizing nuclear structure, exploring the role of RNA in shaping nuclear structure has remained a challenge because no existing methods can simultaneously measure RNA-RNA, RNA-DNA, and DNA-DNA contacts within 3D structures. To address this, we developed RNA & DNA SPRITE (RD-SPRITE) to comprehensively map the location of all RNAs relative to DNA and other RNAs. Using this approach, we identify many RNAs that are localized near their transcriptional loci (RNA-DNA) together with other diffusible ncRNAs (RNA-RNA) within higher-order DNA structures (DNA-DNA). These RNA-chromatin compartments span three major classes of nuclear functions: RNA processing (including ribosome biogenesis, mRNA splicing, snRNA biogenesis, and histone mRNA processing), heterochromatin assembly, and gene regulation. More generally, we identify hundreds of ncRNAs that form stable nuclear compartments in spatial proximity to their transcriptional loci. We find that dozens of nuclear compartments require RNA to guide protein regulators into these 3D structures, and focusing on several ncRNAs, we show that these ncRNAs specifically regulate heterochromatin assembly and the expression of genes contained within these compartments. Together, our results demonstrate a unique mechanism by which RNA acts to shape nuclear structure by forming high concentration territories immediately upon transcription, binding to diffusible regulators, and guiding them into spatial compartments to regulate a wide range of essential nuclear functions.

overall_design: RD-SPRITE was used to simultaneously map the higher-order organization of RNA and DNA in mouse embryonic stem cells.

middle: A

first: Peter

last: Chovanec

first: Mitchell

last: Guttman


title: Affymetrix OncoScan SNP arrays for profiling copy-number alterations (CNAs) and loss-of-heterozygosity (LOH) in colon cancer

first: Sara

last: Lahoz

summary: Intratumor heterogeneity fosters the evolution of the genome leading to metastatic progress and therapy resistance . Here, we investigate the relative contribution of tumor aneuploidy and genomic heterogeneity involving CNAs and mutational events as prognostic and predictive determinants for disease recurrence in early-stage colon cancer patients. We combined SNP arrays, targeted next-generation sequencing, fluorescence in situ hybridization and inmunohistochemistry on a retrospective cohort of 84 untreated stage II colon cancer patients. We assessed the subclonality of copy-number alterations (CNAs) and mutations, CD8+ lymphocyte infiltration levels and their association with time to recurrence  (TTR).

overall_design: Two hundred nanograms of tumor DNA were processed for hybridization on arrays following  the  commercial  protocol  OncoScan  FFPE  assay  kit  (Affymetrix,  Santa  Clara,  CA,  USA). After  hybridization, microarrays were processed and scanned using Affymetrix scanners. Raw array florescence intensity data generated on the Affymetrix scanners in the form of CEL files were loaded into the OncoScan Console software v.1.1.0 (Affymetrix, Santa Clara, California). Quality control statistics as well as integrated OSCHP files were generated by OncoScan Console. To then  generate  whole-genome  copy-number  and  allele-frequency  calls,  raw SNP-array fluorescence intensity data were processed using SNP-FASST2 segmentation algorithm from Nexus Biodiscovery software.

first: Jordi

last: Camps


title: Morc3 silences endogenous retroviruses in mouse embryonic stem cells

status: Public on Oct 28 2021

city: Leiden

first: Lucia

last: Daxinger

address: Albinusdreef 2

institute: Leiden Univeristy Medica Center

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

overall_design: Refer to individual Series


title: High calcium inducible genes in triple negative breast cancer cells

status: Public on Nov 26 2021

city: Nashville

first: LaMonica

middle: V

last: Stewart

state: TN

address: 1005 DB Todd JR BLVD

department: School of Graduate Studies and Research (SOGSR)

institute: Meharry Medical College

summary: Malignant breast cancer is often associated with skeletal metastasis and co-morbidities such as cancer-induced hypercalcemia. Although circulating calcium has been linked to aggressive and larger breast tumors, the mechanisms underlying the potential oncogenic effects of high calcium remain poorly understood. Here, we assessed the effects of high extracellular calcium on the modulation of gene expression in TNBC cells. This study showed that chronic high calcium attenuates the activity of the calcium sensing receptor but promotes the growth and migration of triple negative breast cancer cells. We also show that in TNBC cells, high extracellular calcium triggered the expression of early response genes such as FOS/FOSB and subsequently, mailgnancy assoicated genes including the cancer testis antigen MAGEC2. This study provides novel mechanistic insights into the hitherto unappreciated oncogenic effects of high extracellular calcium especially in malignant or late-stage breast cancers.

overall_design: MDA-MB-231 cells were cultured in complete medium or complete medium supplemented with 3.0 mM or 5.0 mM calcium for 48 h (n=4) and differential gene expression was assessed by gene expression profiling on Human Gene 2.0ST Affimetrix Genechip. After hybridization, the cartridge arrays were washed, and stained per standard Affymetrix protocols using an Affymetrix Fluidics Station 450. The arrays were then scanned in an Affymetrix 7G plus scanner and the resulting data were analyzed by Affymetrix Expression Console v.1.2 using an Robust Multi-array Aver-age (RMA) normalization algorithm producing log base 2 results. Differential gene expression was assessed between replicate groups (n=4) using a moderated t-test and Benjamini-Hochberg multiple testing correction, with significance determined by adjusted p-value <0.05 and absolute value fold change >2.0. Hierarchical clustering was performed in GeneSpring on both averaged and non-averaged normalized expression values.

first: Amos

middle: M

last: Sakwe

first: Siddharth

last: Pratap


title: Spatiotemporal transcriptional atlas of lung adenocarcinoma from adenocarcinoma in situ to invasive carcinoma [scRNA-seq]

status: Public on Nov 30 2021

city: Xi'an

first: Yue

last: Fan

address: Yanta West Road No.76

institute: Xi'an Jiaotong University

summary: To illuminate the evolutionary trajectory of LUAD from AIS to IAC, high-throughput scRNA-seq and ST data were generated and integrated to create a large-scale, single-cell spatiotemporal atlas of LUAD. We compiled a multi-omics atlas of the early-stage LUAD invasion process that reflects the heterogeneity of cancer cells, the competitive polyclonal origin of LUAD, signalling interactions between cancer cells and the TME, and the pseudo-chronological nature of LUAD invasion. The spatial distribution characteristics of LUAD cells revealed the spatial heterogeneity of LUAD and the mechanism of spatial immune escape in LUAD, which provides strong evidence supporting clinical diagnosis and surgical intervention at the single-cell level.

overall_design: Nine resected samples (TD1-9, three each from AIS, MIA and IAC cases) from nine treatment-naïve patients were collected for droplet-based scRNA-seq. In parallel, six paired samples were subjected to ST

first: Tao

last: Jiang

first: Shiquan

last: Sun

first: Jianfei

last: Zhu


title: Morc3 silences endogenous retroviruses in mouse embryonic stem cells [RNA-seq CRISPR]

summary: Background:   MORC proteins are involved in epigenetic gene silencing in a wide variety of eukaryotic organisms. Deletion of MORCs result in several developmental abnormalities and their dysregulation has been implicated in developmental disease and multiple cancers. Specifically, mutations of mammalian MORC3 have been associated with immune system defects, Down syndrome and human cancers such as bladder, uterine, stomach, and lung cancers, and diffuse large B cell lymphomas. While previous studies have shown that MORC3 binds to H3K4me3 in vitro and overlaps with H3K4me3 ChIP-seq peaks in mouse embryonic stem cells, the mechanism by which MORC3 regulates gene expression is unknown.   Results:  In this study, we find that MORC3 functions as an epigenetic silencer of endogenous retroviruses (ERVs) in mouse embryonic stem cells (mESCs). Loss of MORC3 results in upregulation of ERVs, specifically those belonging to the LTR class of retrotransposons. Using ChIP-seq, we measure the genome-wide localization of MORC3 in wild-type cells and find that MORC3 binds to ERVs suggesting its direct role in regulating ERV expression. Previous studies have shown that these ERVs are marked by repressive histone mark H3K9me3 which plays a key role in their silencing. Interestingly, we find that the levels of H3K9me3 do not change substantially upon the loss of MORC3 indicating that MORC3 possibly acts downstream of the TRIM28/SETDB1 complex that deposits H3K9me3 at these loci. Instead, we discover that loss of MORC3 results in increased chromatin accessibility at the ERVs suggesting that MORC3 silences ERVs by compacting DNA in mESCs.    Conclusions:  Our results reveal MORC3 as a novel regulator of ERV silencing in mouse embryonic stem cells. As early mammalian development is characterized by dynamic changes in ERV expression, the role of MORC3 in silencing ERVs is exciting and could potentially explain the abnormalities observed due to its misregulation during mammalian development.

overall_design: Expression profiling by high throughput sequencing   The RNA-seq analysis includes 3 biologicate replicates of the WT mESC and 3 biologicate replicates of the Morc3 mutant for each RNA-seq experiment

first: Varsha

last: Desai

first: Jihed

last: Chouaref

first: Haoyu

last: Wu

first: Steven

last: Jacobsen


title: Global alternative splicing analysis of PRP8 and BRR2/SNRP200 cryptic splicing suppressors in C. elegans

status: Public on Nov 23 2021

city: Santa Cruz

first: Alan

last: Zahler

state: CA

address: Sinsheimer Labs

department: MCD Biology

institute: UC Santa Cruz

summary: The spliceosome undergoes extensive rearrangements as it assembles in multiple steps onto the precursor messenger RNA. In the earliest assembly step, U1snRNA identifies the 5' splice site through base-pairing interactions. However, U1snRNA leaves the spliceosome relatively early in the assembly process. The 5' splice site identity is subsequently maintained through interactions with U6snRNA, protein factor PRP8, and other components of the spliceosome during the complex assembly and rearrangements that build the catalytic site. Using a forward genetic screen in C. elegans, we have identified splicing suppressors of a locomotion defect caused by a 5'ss mutation. Here we report three new extragenic suppressor alleles from this screen, two in PRP8 and one in SNRNP200/Brr2. mRNASeq studies of these suppressor strains indicates that there are specific native targets with alternative 5' and alternative 3' splicing events affected by these suppressors, especially for the suppressor PRP8 D  1549N (position D1556 in humans). The strong suppressor at the unstructured N-terminus of SNRP200, N18K, indicates a potential regulatory role for this region. By examining distinct changes in the splicing of native genes, and by mapping these conserved suppressor residues onto cryoEM structural models of assembling human spliceosomes, we conclude that there are multiple interactions in the spliceosome that are required to ensure that the initial 5'ss identified by U1snRNA early in spliceosome assembly is the one that gets loaded into the catalytic core.The spliceosome undergoes extensive rearrangements as it assembles in multiple steps onto the precursor messenger RNA. In the earliest assembly step, U1snRNA identifies the 5' splice site through base-pairing interactions. However, U1snRNA leaves the spliceosome relatively early in the assembly process. The 5' splice site identity is subsequently maintained through interactions with U6snRNA, protein factor PRP8, and other components of the spliceosome during the complex assembly and rearrangements that build the catalytic site. Using a forward genetic screen in C. elegans, we have identified splicing suppressors of a locomotion defect caused by a 5'ss mutation. Here we report three new extragenic suppressor alleles from this screen, two in PRP8 and one in SNRNP200/Brr2. mRNASeq studies of these suppressor strains indicates that there are specific native targets with alternative 5' and alternative 3' splicing events affected by these suppressors, especially for the suppressor PRP8 D  1549N (position D1556 in humans). The strong suppressor at the unstructured N-terminus of SNRP200, N18K, indicates a potential regulatory role for this region. By examining distinct changes in the splicing of native genes, and by mapping these conserved suppressor residues onto cryoEM structural models of assembling human spliceosomes, we conclude that there are multiple interactions in the spliceosome that are required to ensure that the initial 5'ss identified by U1snRNA early in spliceosome assembly is the one that gets loaded into the catalytic core.

overall_design:
Strains with cryptic splicing suppressor mutations in PRP-8 (R540K and D1549N) and Brr2/SNRP-200 (N18K) were compared to a control strain to identify changes in native gene alternative splicing
Samples from strain SZ299 are from animals homozygous for the PRP-8(R540K) mutation
Samples from strain SZ304 are from animals homozygous for the PRP-8(D1549N) mutation

first: Sol

last: Katzman


title: Morc3 silences endogenous retroviruses in mouse embryonic stem cells [Morc3 ChIP-seq]

overall_design: Endogenous Morc3 (Rockaland 100-401-N97) ChIP-seq experiment in mESc


title: Morc3 silences endogenous retroviruses in mouse embryonic stem cells [H3K9me3 ChIP-seq]

overall_design: H3K9me3 ChIP-seq experiment performed in 2 biological replicates for the WT and 2 biological replicaties for the Morc3D41/D41


title: Targeted Next-Generation Sequencing for profiling frequently mutated genes in early-stage colon cancer

summary: Intratumor heterogeneity fosters the evolution of the genome leading to metastatic progress and therapy resistance. Here, we investigate the relative contribution of genomic heterogeneity involving mutations and CNAs as prognostic and predictive determinants for disease recurrence in early-stage colon cancer. We combined targeted next-generation sequencing (NGS) and SNP arrays on a retrospective cohort of untreated stage II colon cancer patients to assess the association of genomic subclonality with time to recurrence (TTR).

overall_design: Enriched libraries were prepared from 50 ng of DNA per tumor sample using the SMARTer ThruPLEX DNA-seq Library Preparation kit (TaKaRa, Japan) and a predesigned gene capture pool of IDT xGen Lockdown probes (IDT, Coralville, Iowa, USA). Paired-end sequencing runs were performed on a MiSeq platform (Illumina, San Diego, CA, USA) following a protocol with 2x130 bp reads.


title: 3D ECM-rich environment sustains the identity of naïve human iPSCs [Bulk RNA-seq in triplicate after 1, 4, and 7 days of culture]

status: Public on Nov 11 2021

city: Santander

first: Lucio

last: Di Filippo

state: Cantabria

address: PCTCAN - Cl. Albert Einstein, 22

institute: IBBTEC

summary: The establishment of in vitro naïve human pluripotent stem cell cultures opened new perspectives for the study of early events in human development. In the last 5 years the signalling environment promoting human naïve pluripotency has been extensively investigated. However, little is known about the role exerted by the extracellular matrix and three-dimensional organization in naïve maintenance. Here, by using an unbiased and integrated approach we found that naïve, but not primed, hiPSC colonies are characterized by an extracellular matrix-rich microenvironment, and by a specific extracellular matrix fingerprint at transcriptional, proteomic and secretome levels. Based on this, we developed a 3D culture system which supports robust long-term feeder-free expansion of naïve hiPSCs. In this same 3D ECM-rich environment, we can recapitulate timely developmental morphogenesis simply by modulating the signalling environment. Our study opens new perspectives for future applications of naïve hiPSCs to study critical stages of human development in three-dimensions starting from a single cell.

overall_design: Bulk RNA-seq in triplicate after 1, 4, and 7 days of culture. Microfluidic and standard plate culture of human fibroblasts (HFF-1), naïve iPSCs (isogenic, HFF-1-derived), and induced primed (isogenic, naïve iPSCs-derived) .

first: Elisa

last: Cesare

first: Anna

last: Urciuolo

first: Hannah

last: Stuart

first: Erika

last: Torchio

first: Cecilia

last: Laterza

first: Onelia

last: Gagliano

first: Sebastian

last: Martewicz

first: Meihua

last: Cui

last: Manfredi

first: Patrizia

last: Sabatelli

first: Stefano

last: Squarzoni

first: Irene

last: Zorzan

first: Graziano

last: Martello

first: Davide

last: Cacchiarelli

first: Camilla

last: Luni

first: Nicola

last: Elvassore


title: Sequencing of Mus musculus brain (hippocampus) after enriched environment 3

status: Public on Oct 19 2021

city: Jena

first: JenAge

last: Project

address: Beutenbergstr. 11

institute: Leibniz Institute for Age Research - Fritz Lipmann Institute

summary: Comparison of gene expression profiles from Mus musculus brain (hippocampus) of animals kept in standard environment and enriched environment. The RNA-seq data comprise 4 groups: 2 age groups, each w/ and w/o enriched environment.    Jena Centre for Systems Biology of Ageing - JenAge (www.jenage.de)

overall_design: 39 samples in 4 groups: 5 months control  (10 samples), 5 months enriched environment (9 samples), 24 months control (15 samples), 24 months enriched environment (5 samples)

first: Christiane

last: Frahm

first: Otto

middle: W

last: Witte

first: Madlen

last: Günther

first: Rolf

last: Hühne

first: Marco

last: Groth

first: Matthias

last: Platzer


title: Morc3 silences endogenous retroviruses in mouse embryonic stem cells [ATAC-seq]

first: Lucia

last: Daxinger

overall_design: Chromatin accessibility profile  The ATAC-seq analysis includes 2 biological replicates for the WT cell lines and 2 biological replicates for the mutant cell lines

first: Varsha

last: Desai

first: Jihed

last: Chouaref

first: Haoyu

last: Wu

first: Steven

last: Jacobsen


title: Morc3 silences endogenous retroviruses in mouse embryonic stem cells [Morc3_mESCs_RNAseq]


title: Differentiating mouse ESC towards gonadal progenitors cells

status: Public on Oct 15 2021

city: Ramat Gan

first: Nitzan

last: Gonen

address: Bar-Ilan University

department: Life Sciences

country: Israel

institute: Bar-Ilan University

summary: We developed a gradual differentiation protocol to differentiate mESC towards the EpiSC lineage, followed by early mesoderm cells. These were later differentiated to intermediate mesoderm and we found a condition (IM3) that best generates early gonadal progenitor cells which are normally present at the bipotential gonad at E11.5. Forced expression of 2 transcription factors: Nr5A1 (SF1) and Dmrt1 induced a specific-Sertoli marker (TESCO-CFP) and the cells started to self-aggregate and form tubule-like structures, highly resembling the testis cords. There was proper induction of relevant markers at the correct time-points as shown by both qPCR, immuno-staining and RNA-Seq that includes comparison to in-vivo gonads.

overall_design: Bulk RNA-Seq of cells at different stages of the differentiation: ESC, EpiSC, Early Mesoderm, Intermediate mesoderm, induced embryonic Sertoli like cells.

first: Robin

last: Lovell-Badge


title: Longitudinal DNA methylation and RNA-seq datasets in primary human fibroblasts with mitochondrial genetic, metabolic, and endocrine perturbations

city: New York

first: Gabriel

last: Sturm

summary: The current RNA sequencing and DNA methylation datasets provide an epigenomic map across the entire replicative lifespan of primary human fibroblast cell lines from n=6 healthy male and female donors and n=3 donors with lifespan-altering mitochondrial disease caused by mutations in the SURF1 gene. These datasets can be integrated with other measures collected in parallel along the lifespan, including cytological (cell size, morphology), bioenergetic (energy expenditure, derived ATP synthesis rates from Seahorse), secreted proteins, telomere length, and whole-genome sequencing (WGS) data. This dataset also includes experimental manipulations with treatments targeting oxidative phosphorylation (OXPHOS) and glycolysis, and glucocorticoid signaling, providing an opportunity to examine the influence of stress and bioenergetics on human aging biology. All processed data can be accessed and browsed at our webtool: https://columbia-picard.shinyapps.io/shinyapp-Lifespan_Study/


title: Single-cell RNA-sequencing of thymic myeloid cells from Csf1rgfp/gfp (MaFIA) and Cd11cyfp/yfp mice

status: Public on Nov 03 2021

city: Taipei

first: Tyng-An

last: Zhou

state: Taiwan

address: No.155, Sec.2, Linong Street

department: Institute of Microbiology and Immunology

country: Taiwan

institute: National Yang Ming Chiao Tung University

summary:
Purpose: The goals of this study are to identify different sub-populations of mouse thymic myeloid cells and to compare their differentially expressed genes.
Methods: Single-cell mRNA libraries of EGFP-positive and YFP-positive cells from 5-weeks-old Csf1rgfp/gfp and 9-weeks-old Cd11cyfp/yfp mice were generated with Chromium X (10X Genomics), in two biological replicates, each, followed by sequencing using Illumina NovaSeq. The sequence reads that passed quality filters were aligned against Ensembl GRCm39 mouse genome assembly with GRCm39.104 annotation by STARsolo, downsampled by DropletUtils, and analyzed by SingleCellExperiment.
Results: we obtained about 1 billion sequence reads per sample, except for 1 sample (MaFIA rep1), which contained reads mostly derived from mitochondria and was excluded from downstream analysis. In the other three samples, over 90% of reads had valid barcodes and reached sequencing saturation about 70%. Over 80% of reads could be mapped to mouse genome uniquely (build GRCm39). Each sample had estimated number of cells over 6,000. Downstream analysis identified different subsets of thymic macrophages and dendritic cells, along with small amount of plasmacytoid dendritic cells, T cells and B cells, all of these populations could be clearly clustered and presented on tSNE or UMAP.
Conclusions: We could identify 4 populations of dendritic cells, 2 populations of macrophages and one monocyte population in the thymus. This dataset is a rich resource for exploring the diversity of thymic myeloid cells at single cell resolution.

overall_design: Transcriptional profiling of thymic macrophages at single-cell level, generated from single-cell preparation of the MaFIA and CD11c-YFP mouse thymus EGFP or YFP positive cells, sequenced in several batches on illumina NovaSeq platform.

first: Ivan

middle: L

last: Dzhagalov


title: High quality mapping of chromatin at or near the nuclear lamina using cTSA-seq

status: Public on Oct 27 2021

city: Baltimore

first: Joseph

last: Tran

address: 3520 San Martin Drive

department: Embryology

institute: Carnegie Institution for Science

summary: We presented an adaptation of the TSA-seq antibody based method to map chromatin at the nuclear lamina from lower numbers of cells, which we term chromatin-based TSA-seq or cTSA-seq. We provide evidence that the method maps closed heterochromatin at or near the nuclear lamina that is in the B-compartment and show that it is useful down to 50,000 cells. We applied this emthod to the early G1 cell population to verify its utility and show that telomeric ends are indeed NL-proximal during this stage. We further provide evidence that the early G1 LADs profile is reminiscent of the profiles observed for oncogene-induced senescence.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: Development of cTSA-seq and application to low numbers of cells

first: Yixian

last: Zheng

first: Xiaobin

first: Stephen

last: Adam

first: Robert

last: Goldman


title: 3D ECM-rich environment sustains the identity of naïve human iPSCs

first: Lucio

last: Di Filippo

overall_design: Bulk RNA-seq. Microfluidic 3-day culture of human naïve iPSCs on MEF feeder layer (3 replicates) and on Vitronectin coating (3 replicates).

first: Elisa

last: Cesare

first: Anna

last: Urciuolo

first: Hannah

last: Stuart

first: Erika

last: Torchio

first: Cecilia

last: Laterza

first: Onelia

last: Gagliano

first: Sebastian

last: Martewicz

first: Meihua

last: Cui

last: Manfredi

first: Patrizia

last: Sabatelli

first: Stefano

last: Squarzoni

first: Irene

last: Zorzan

first: Graziano

last: Martello

first: Davide

last: Cacchiarelli

first: Camilla

last: Luni

first: Nicola

last: Elvassore


title: Glioblastoma Genetic Drivers Dictate the Function of Tumor-Associated Macrophages/Microglia and Responses to CSF1R Inhibition

status: Public on Oct 08 2021

city: Cincinnati

first: Qing

middle: Richard

last: Lu

email: raorr@mail.uc.edu, richard.lu@cchmc.org

state: OH

address: 3333 Burnet Ave

institute: Cincinnati Children's Hospital Medical Center

summary: Tumor-associated macrophages/microglia (TAMs) are prominent microenvironment components in human glioblastoma (GBM) that are potential targets for anti-tumor therapy. However, TAM depletion by CSF1R inhibition showed mixed results in clinical trials. We hypothesized that GBM subtype-specific tumor microenvironment convey distinct sensitivities to TAM targeting.We generated syngeneic PDGFB-driven and RAS-driven GBM models that resemble proneural-like and mesenchymal-like gliomas, and determined the effect of TAM targeting by CSF1R inhibitor PLX3397 on glioma growth. We also investigated the co-targeting of TAMs and angiogenesis on PLX3397-resistant RAS-driven GBM. Using single-cell transcriptomic profiling, we further explored differences in tumor microenvironment cellular compositions and functions in PDGFB- and RAS-driven gliomas. We found that growth of PDGFB-driven tumors was markedly inhibited by PLX3397. In contrast, depletion of TAMs at the early phase accelerated RAS-driven tumor growth and had no effects on other proneural and mesenchymal GBM models. In addition, PLX3397-resistant RAS-driven tumors did not respond to PI3K signaling inhibition. Single-cell transcriptomic profiling revealed that PDGFB-driven gliomas induced expansion and activation of pro-tumor microglia, whereas TAMs in mesenchymal RAS-driven GBM were enriched in pro-inflammatory and angiogenic signaling. Co-targeting of TAMs and angiogenesis decreased cell proliferation and changed the morphology of RAS-driven gliomas.Our work identify functionally distinct TAM subpopulations in the growth of different glioma subtypes. Notably, we uncover a potential responsiveness of resistant mesenchymal-like gliomas to combined anti-angiogenic therapy and CSF1R inhibition. These data highlight the importance of characterization of the microenvironment landscape in order to optimally stratify patients for TAM-targeted therapy.

overall_design: Single Cell RNA-seq (10X and Drop-Seq) for glioma driven by HRasV12 and dominant negative p53. Bulk RNA-seq of Ras-driven glioma tumor tissues from mouse treated with vehicle and PLX3397. Bulk RNA-seq of cell lines derived from PDGFB and Ras-driven GBM. Bulk RNA-seq of Cd11b+ positive cells from PDGFB-driven GBM after three days of vehicle or PLX3397 treatment

first: Rohit

last: Rao

first: Rong

last: Han

first: Sean

last: Ogurek

first: Chengbin

last: Xue

first: Lai

last: Wu

first: Liguo

last: Zhang

first: Li

first: Jian

last: Hu

first: Timothy

middle: N

last: Phoenix

first: Stephen

last: Waggoner

first: Q


title: A longitudinal DNA methylation dataset in primary human fibroblasts with mitochondrial genetic, metabolic, and endocrine perturbations

first: Gabriel

last: Sturm

summary: The current DNA methylation dataset (n=479) provides an epigenetic map of cultured primary human fibroblasts (in vitro) aged through the cellular lifespan. The dataset was generated using the Illumina EPIC bead array, which includes DNA methylation levels (0-1.0) at 866,091 CpG sites. This provides DNA methylation trajectories collected at high temporal frequency (every 8 days, ~7timepoints per a donor) across the entire replicative lifespan of primary human fibroblast cell lines from n=6 healthy male and female donors and n=3 donors with lifespan-altering mitochondrial disease caused by mutations in the SURF1 gene. The dataset can be integrated with other measures collected in parallel along the lifespan, including cytological (cell size, morphology), bioenergetic (energy expenditure, derived ATP synthesis rates from Seahorse), gene expression (RNA sequencing, GSE179848), secreted proteins, telomere length, and whole-genome sequencing (WGS) data. This dataset also includes experimental manipulations with treatments targeting oxidative phosphorylation (OXPHOS) and glycolysis, and glucocorticoid signaling, providing an opportunity to examine the influence of stress and bioenergetics on human aging biology. All processed data can be accessed and browsed at our webtool: https://columbia-picard.shinyapps.io/shinyapp-Lifespan_Study/

overall_design: Primary human fibroblasts are cultured and passaged over ~250 days of replicative lifespan in various treatment conditions.

first: Martin

last: Picard


title: Sequencing of Mus musculus brain (hippocampus) after enriched environment

first: JenAge

last: Project

overall_design: 20 samples in 4 groups: 5 months control (5 samples), 5 months enriched environment (5 samples), 24 months control (5 samples), 24 months enriched environment (5 samples)

first: Christiane

last: Frahm

first: Otto

last: Witte

first: Madlen

last: Günther

first: Rolf

last: Hühne

first: Marco

last: Groth

first: Matthias

last: Platzer


title: Elimination of radiation-induced senescence in the brain tumor microenvironment attenuates glioblastoma recurrence

status: Public on Sep 22 2021

city: Dallas

first: Ralf

last: Kittler

state: Texas

address: 5323 Harry Hines Blvd

institute: University of Texas Southwestern Medical Center

summary: We report gene expression changes in the brains of C57BL6/J mice 30 days after cranial irradiation with 10 Gy of X-rays

overall_design: Examination of RNA profiles of brains of C57BL6/J mice at 30 days post irradiation compared to mock-irradiated mice (3 mice per group)

first: Sandeep

last: Burma

first: Eliot

last: Fletcher-Sananikone

first: Rahul

last: Kollipara


title: Droplet-based Single-cell Total RNA-seq Reveals Differential Non-Coding Expression and Splicing Patterns during Mouse Development

status: Public on Sep 16 2021

first: Anna

last: Alemany

email: annalemany@gmail.com, a.alemany@lumc.nl

state: South Holland

address: ALBINUSDREEF 2

department: Anatomy and Embryology

institute: LUMC

summary: Single-cell RNA-seq is one of the most important and widely used approaches to characterize cell types and to understand major parts of biological systems. As of today, most methods are only able to capture parts of the whole transcriptome, mainly the protein-coding genes, and therefore lack information about non-coding biotypes or full-length transcripts. Here, we present “Vast transcriptome Analysis of Single-cells by dA-tailing (VASA-seq)”, a method for highly-sensitive, full-length and total RNA-seq in single-cells. VASA-seq is compatible with plate-formats for sorting of rare cell populations, and with droplet microfluidics for high-throughput applications and atlasing. We applied VASA-seq to >30,000 single cells in the developing mouse embryo during gastrulation and early organogenesis. Our in-depth analysis revealed expression of novel cell-type specific non-coding RNA markers, enhanced cell-cycle characterization and alternative splicing patterns. We believe the method and mouse dataset will serve as a useful tool to further investigate the expression of different RNA biotypes and splicing patterns in large datasets.

overall_design: total RNA sequencing data (VASA-seq) from Hek293T cells, mES cells and E6.5, E7.5, E8.5 and E9.5 mouse embryo

first: Fredrik

last: Salmen

first: Joachim

last: De Jonghe

first: Tomasz

last: Kaminski

last: Alemanny

first: Guillermo

last: Parada

first: Joe

last: Verity-Legg

first: Timo

last: Kohler

first: Nicholas

last: Battich

first: Floris

last: van den Brekel

last: Ellermann

first: Alfonso

last: Arias

first: Jennifer

last: Nichols

last: Hemberg

first: Florian

last: Hollfelder

first: Alexander

last: van Oudenaarden


title: Mitogen induced-nucleosomal response pathway and transcriptional programming in mammalian fibroblasts

status: Public on Sep 13 2021

city: Winnipeg

first: James

middle: Ronald

last: Davie

state: Manitoba

address: 745 Bannatyne Avenue, RM 333A

department: Biochemistry and Medical Genetics

institute: University of Manitoba

summary: Stimulation of the RAS-MAPK pathway activates mitogen- and stress-activated kinases (MSK) 1 and 2, which phosphorylate histone H3 at S10 (H3S10ph) or S28 (H3S28ph) (nucleosomal response) located at the regulatory regions of immediate-early genes, setting in motion a series of chromatin remodeling events that result in transcription initiation. To investigate immediate-early genes regulated by the MSK, we have completed transcriptome analyses (RNA Sequencing) of human normal fibroblast cells (CCD-1070Sk) stimulated with EGF or TPA ± H89, a potent MSK/PKA inhibitor.

overall_design: Treatment of quiescent human cells with fetal bovine serum (FBS), epidermal growth factor (EGF) or the phorbol ester (12-O-tetradecanoylphorbol-13-acetate, TPA) ± H89 following RNA Sequencing.

first: Kiran

last: Sharma

first: Shuo

last: Jia

first: Tasnim

last: Beacon

first: Ifeoluwa

last: Adewumi

first: Camila

last: López

first: Pingzhao

last: Hu

first: Wayne

last: Xu


title: The effect of PTP1B inhibitor (MSI-1436) on neutrophils.

status: Public on Sep 18 2021

city: Cold Spring Harbor

middle: K

last: Tonks

address: 1 Bungtown Rd

institute: Cold Spring Harbor Laboratory

summary: Acute Lung Injury (ALI) can cause Acute Respiratory Distress Syndrome (ARDS), a lethal condition with limited treatment options and currently a common global cause of death due to COVID-19-induced ALI. ARDS secondary to Transfusion-Related Acute Lung Injury (TRALI) has been recapitulated pre-clinically by anti-MHC-I antibody administration to LPS-primed mice. In this model, we demonstrated that inhibitors of PTP1B, a protein tyrosine phosphatase that regulates signaling pathways of fundamental importance to homeostasis and inflammation, prevented lung injury and increased survival. Treatment with PTP1B inhibitors attenuated the aberrant neutrophil function that drives ALI, and was associated with release of myeloperoxidase, suppression of Neutrophil Extracellular Trap (NET) formation, and inhibition of neutrophil migration. Mechanistically, reduced signaling through the CXCR4 chemokine receptor, particularly to the activation of mTOR, was essential for these effects, linking PTP1B in  hibition to promoting an aged neutrophil phenotype. Considering dysregulated activation of neutrophils is implicated in sepsis and can cause collateral tissue damage, we demonstrated also that PTP1B inhibitors improved survival and ameliorated lung injury in the LPS-induced sepsis model. Our data highlight PTP1B inhibition for prevention of TRALI and ARDS from multiple etiologies.

overall_design: RNA-seq: Neutrophils were isolated from mouse bone marrow, and were treated with MSI-1436 for 2 hours.

first: Dongyan

last: Song

first: Jose

last: Adrover

first: Xiaoli

last: Wu


title: Transcriptomic profiling of oncogenic KRAS and TP53 in lung adenocarcinoma cells

status: Public on Oct 01 2021

city: Seattle

first: Alice

last: Berger

state: WA

address: 1100 Fairview Ave N

institute: Fred Hutchinson Cancer Center

summary: Based on the previous eVIP study (Cancer Cell, 2016), we profiled the transcriptomes of lung adenocarcinoma cell lines expressing oncogenic KRAS and TP53 to compare methods and investigate the effects of these oncogenes on the cells' whole transcriptomes.

overall_design: mRNA profiles

first: Lo

last: April

first: Berger

last: Alice


title: Single-cell transcriptomics identifies Keap1-Nrf2 regulated collective invasion in a Drosophila tumor model

status: Public on Sep 19 2021

city: New Orleans

first: Wu-Min

last: Deng

state: LA

address: 1700 Tulane Avenue

department: School of Medicine, Department of Biochemistry

institute: Tulane University

summary: Apicobasal cell polarity loss is a founding event in epithelial-mesenchymal transition and epithelial tumorigenesis, yet how pathological polarity loss links to plasticity remains largely unknown. To understand the mechanisms and mediators regulating plasticity upon polarity loss, we performed single-cell RNA sequencing of Drosophila ovaries, where inducing polarity-gene l(2)gl-knockdown (Lgl-KD) causes invasive multilayering of the follicular epithelia. Analyzing the integrated Lgl-KD and wildtype transcriptomes, we discovered the cells specific to the various discernible phenotypes and characterized the underlying gene expression. A genetic requirement of Keap1-Nrf2 signaling in promoting multilayer formation of Lgl-KD cells was further identified. Ectopic expression of Keap1 increased the volume of delaminated follicle cells that showed enhanced invasive behavior with significant changes to the cytoskeleton. Overall, our findings describe the comprehensive transcriptome of cells within the follicle cell tumor model at the single-cell resolution and identify a previously unappreciated link between Keap1-Nrf2 signaling and cell plasticity at early tumorigenesis.

overall_design: Whole-tissue RNA-Seq (2 replicates per genotype; 2 time-points - 24h and 96h) of tjTS>GFP (Control) and tjTS>lglRNAi (Sample) and single-cell RNA-Seq (1 replicate; 1 time-point - 72h) of w1118 cells (Control) of the Drosophila ovary and tjTS>lglRNAi cells (Sample) of the Drosophila ovary. Single-cell RNA-Seq: 10x Genomics CHROMIUM Single Cell 3’ Solution V2 Chemistry and Sequencing platform: Illumina NovaSeq 6000.

first: Deeptiman

last: Chatterjee


title: Non-coding genetic variation in GATA3 increases acute lymphoblastic leukemia risk through local and global changes in chromatin conformation

city: CHICAGO

first: Yu

last: Luan

state: Illinois

address: 303 E. Superior Simpson Querrey 7-518

department: Department of Biochemistry and Molecular Genetics

institute: Northwestern University Feinberg School of Medicine

summary: Inherited non-coding genetic variants confer significant disease susceptibility in many cancers. However, the molecular processes by which germline variants contribute to somatic lesions are poorly understood. We performed targeted sequencing in 5,008 patients and identified a key regulatory germline variant in GATA3 strongly associated with Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL). By creating an isogenic cellular model with a CRISPR-Cas9 system, we showed that this variant activated a strong enhancer that significantly upregulated GATA3 transcription, which in turn reshaped the global chromatin accessibility and 3D genome organization. Remarkably, this genotype switch induced a chromatin loop between the CRLF2 oncogene and a distal enhancer, similar to the somatically acquired super-enhancer hijacking event in patients. GATA3 genotype-related alterations in transcriptional control and 3D chromatin organization were further validated in Ph-like ALL patients. Finally, we showed that GATA3 directly regulates CRLF2 and potentiates the oncogenic effects of JAK-STAT signaling in leukemogenesis. Altogether, our results provide evidence for a novel mechanism by which a germline non-coding variant contributes to oncogene activation epigenetic regulation and 3D genome reprogramming.

overall_design: GM12878 cells were engineered to knock in the A allele at rs3824462. Consequences of the epigenetics and chromatin topology change between GM12878 cells/patients(CC) and engineered cells/patients(AA)  were investigated using ChIP-seq, RNA-seq, and Hi-C.

first: Hongbo

last: Yang

first: Hui

last: Zhang

first: Feng

last: Yue

first: Jun


title: RNA sequencing of preB-ALL NALM-6 cells with or without XIAP-knockdown, using prime-seq

status: Public on Sep 15 2021

city: Munich

first: Irmela

last: Jeremias

address: Feodor-Lynenstr. 21

department: Research Unit Apoptosis in Hematopoietic Stem Cells

institute: Helmholtz Zentrum München, German Center for Environmental Health (HMGU), Munich, Germany

summary: In this study we performed molecular manipulation of XIAP in vivo using microRNA-30 flanked shRNA expression in cell lines models of r/r ALL. Interestingly, upon strong XIAP knockdown,  r/r ALL cells were outcompeted in vivo, while intermediate knockdown sensitized cells to chemotherapy. Using prime-seq we wanted to characterized NALM-6 cells expressing different XIAP levels.

overall_design:
XIAP KD and control NALM-6 cells were isolated out of the murine bone marrow at different time points. the mRNA expression pattern was analyzed.
NOTE FROM SUBMITTER: Due to legal regulations in Germany, we are not allowed to publish raw sequencing data. We already asked for permission to only publish the raw counts.

first: Jeremias

last: Irmela

first: Carlet

last: Michela

first: Fabian

last: Klein

first: Johannes

last: Bagnoli