title: Bulk RNA-seq analysis of gene expression changes at 8 hpf induced by miR-34a loss

city: Ottawa

first: Sergey

last: Prykhozhij

state: ON

address: Room 318/319 | CAREG, 20 Marie-Curie Private

institute: CHEO Research Institute/University of Ottawa

summary: Li-Fraumeni syndrome (LFS) is a disorder due to inherited mutations in the TP53 gene resulting in an increased risk of developing several types of cancer. MicroRNA miR-34a has been implicated downstream of p53 on the basis of being a direct transcriptional target and, when over-expressed, having pro-apoptotic phenotypes in cell lines. Moreover, miR-34a has been shown to be a modifier gene in the context of LFS, since its epigenetic silencing increases the likelihood of tumour development in patients with mutant TP53.  However, the in vivo consequences of miR-34 loss are still unclear. For example, mice lacking all three (a,b,c) miR-34 homologs show no detectable abnormalities in p53 responses. The relative expression of different miR-34 genes in zebrafish was studied using qRT-PCR with specific assays. The miR-34a, miR-34b and miR-34c display unique onset of developmental expression and expression levels, with miR-34a being the most abundant and constant in expression. All of the miR-34 genes also showed clear induction by p53 when DNA-damaging treatments are performed. Using CRISPR-Cas9 technology, we generated a zebrafish miR-34a deletion mutant to further investigate the roles of miR-34a on its own and its association with the p53 pathway. Predictably, a miR-34a deletion mutant demonstrated absence of miR-34a, though without miR-34b and miR-34c compensation beyond baseline expression levels. Mutants survive to adulthood, show no overt phenotypes and have normal apoptotic responses to DNA-damaging irradiation or camptothecin treatments. To further explore the effects of miR-34a, we performed gene expression profiling using RNA-seq of wild-type and miR-34a deletion mutant zebrafish embryos at 8 hpf. This experiment was motivated by a previous report knock-down of miR-34a in zebrafish leads to dramatic increases in expression of miR-34a target genes. We therefore expected that this experiment will help define the set of miR-34a target genes. The results of this RNA-seq experiment showed that the loss of miR-34a led to large transcriptomic effects at 8 hpf (1573 genes UP and 1679 genes DOWN at 1.5-fold change and FDR < 0.05). There was no significant enrichment of predicted miR-34a target genes among the differentially regulated genes but some interesting biological trends were found and will be described in the paper associated with this dataset.

overall_design: Zebrafish embryos at 8 hpf of wild-type (casper) and miR-34a-/- genotypes (4 groups of 30 embryos each) were dechorionated, flash-frozen followed by RNA extraction from all samples.

first: Jason

middle: N

last: Berman


title: Setting up a better infection: Overexpression of the early bacteriophage T4 gene motB during infection results in a more favorable tRNA pool for the phage

status: Public on Jun 14 2022

city: Bethesda

first: Deborah

last: Hinton

state: MD

address: 8 Center drive

country: USA

institute: NIDDK

type: Expression profiling by high throughput sequencing

summary:
Purpose: We investigated the effect of the T4 MotB protein on E. coli gene expression
Method: E. coli BL21 (DE3) containing either pNW129 or pNW129-MotB were grown to early log phase (OD600 ~ 0.3)  then induced with 0.2% arabinose for 20 minutes. T4 phage added to the culture at MOI10. Cells were then harvested at 0, 5, and 10 min time points and total RNA was isolated. The cDNA library was prepared using a modified RNATagSeq workflow as previously described (Shishkin, A.A. et al. 2015 Nat Methods). Optimum fragmentation of the total RNA samples in this library was determined to be 3 min at 94C in FastAP buffer (Thermo Fischer Scientific).  The cDNA Library was run on a Bioanalyzer using the Agilent High Sensitivity DNA Kit to evaluate the quality of the library.  The concentration of the cDNA library was determined by qPCR using the KAPA Library Quantification Kit (Kapa Biosystems, Wilmington, MA, USA) and CFX96 Real-Time PCR Detection System (Bio-Rad, Hercules, CA, USA). Sequencing was performed by the NIDDK Genomics Core facility using a MiSeq system with the single-end 50 bp Sequencing Kit (Illumina, San Diego, CA, USA). RNA-seq data was processed as previously described using E. coli str. K-12 substr. MG1655 (NC_000913.3) as the reference genome.  Differential expression between conditions was represented as a fold change, and genes with both a fold change ≥2 or ≤ 0.5 and adjusted p value ≤ 0.05 were considered significant.
Results: RNA-seq data revealed that the expression of 542 E. coli genes were significantly changed after motB expression. The expression of 704 and 706 E. coli genes were changed T4 5 min and 10 min post infection after MotB expression, respectively. After 5 min T4 infection, 34 T4 genes were significantly changed.
Conclusion: T4 MotB modifies the pool of host tRNAs, creating a better infection for T4

overall_design: To determine the effect of the T4 MotB protein on E. coli  gene expression, RNAseq was performed for E. coli BL21 (DE3) infected by T4 after producing T4 MotB for 20 minutes .

first: Bokyung

last: Son

first: Jennifer

last: Patterson-West

first: Christine

middle: O

last: Thompson

first: James

middle: R

last: Iben

middle: M


title: p53 regulates the conserved zebrafish microRNA-34 gene family, of which miR-34a has direct and p53-mediated effects including tumour suppression

country: Canada

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

overall_design: Refer to individual Series


title: Small molecule RIG-I agonist serves to adjuvant broad, multifaceted influenza virus vaccine immunity

city: Seattle

first: Richard

last: Green

state: WA

address: 750 Republican St, Seattle, WA 98109

department: Immunology

institute: UW

summary:
We identified a small molecule compound, KIN1148, that directly binds RIG-I to drive IRF3 and NF B activation and expression of innate immune genes, cytokines and chemokines. KIN1148 activates RIG-I in an RNA- and ATP-independent manner and does not induce a canonical antiviral interferon (IFN) gene program traditionally associated with RIG-I activation. When administered in combination with a vaccine against influenza A virus (IAV), KIN1148 induces both neutralizing antibody and broadly cross-protective IAV-specific T cell responses compared to vaccination alone, which induces poor responses.
In this study, we demonstrate that KIN1148 directly engages RIG-I to activate IRF3- and NFB-dependent innate immune responses, making it the first small molecule RIG-I agonist to be identified. Biochemical studies show that KIN1148 binds to RIG-I to drive RIG-I self-oligomerization and downstream signaling activation in an RNA- and ATP-independent manner. We further find that transcriptional programs induced by KIN1148 treatment exhibit shared and unique signatures to that induced by other methods of RIG-I activation, including Sendai virus (SeV) infection and PAMP RNA transfection. KIN1148 adjuvants a split virus (SV) vaccine at suboptimal dose to protect mice from lethal challenge with a recombinant highly pathogenic avian H5N1 influenza virus, A/Vietnam/1203/2004.

overall_design: Differentiated THP-1 cells were treated with small molecule compounds (KIN1000 and KIN1148 at 0.625, 2.5 or 10 micromoles/liter) or 25 IU/mL IFN Beta diluted in cRPMI supplemented with 0.5% (v/v) DMSO, or treated with cRPMI supplemented with 0.5% (v/v) DMSO alone (DMSO). Control cells infected with 25 HAU/mL SeV were maintained in cRPMI supplemented with 0.5% (v/v) DMSO after removal of the virus inoculum.

first: Emily

last: Hemann

first: Megan

middle: L

last: Knoll

first: Courtney

last: Wilkins

first: Renee

last: Ireton

first: Michael

last: Gale Jr


title: Cohesin-dependent chromosome loop extrusion is limited by transcription and stalled replication forks

status: Public on Jun 15 2022

city: Bunkyo-ku

first: Ryuichiro

last: Nakato

state: Tokyo

address: 1-1-1 Yayoi

department: Institute for Quantitative Biosciences

country: Japan

institute: The University of Tokyo

type: Genome binding/occupancy profiling by high throughput sequencing


title: Epigenetic profiling of mouse hematopoietic cells retrovirally transfected with MNX1 or empty vector (Ctr) from fetal liver or mouse bone marrow

city: Heidelberg

first: Dieter

last: Weichenhan

address: Im Neuenheimer Feld

department: Cancer Epigenomics

country: Germany

institute: Deutsches Krebsforschungszentrum


title: Transcriptomic analysis of hCMEC/D3 cells after SiNPs treatment

city: Kaifeng

first: Haigang

last: wu

state: Henan province

address: dongjing Road

department: School of Life Sciences

country: China

institute: Henan University

summary: hCMEC/D3 cells were treated with SiNPs for 24 h. It was discovered that SiNP_35 nm and SiNP_58 nm were both penetrated through the BBB via caveolin-mediated pathway, and both sizes induced the upregulation of the inflammation pathways.

overall_design: Examination of transcriptome level in hCMEC/D3 cells treated with SiNPs for 24 h.

first: Kaiting

last: Miao


title: Human midbrain dopaminergic neuronal differentiation markers predict cell therapy outcome in a Parkinson's disease model

city: Shanghai

first: Yuejun

last: Chen

state: Shanghai

address: 320 Yueyang Road

institute: Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences

summary: Human pluripotent stem cell (hPSC)-based replacement therapy holds great promise in treating Parkinson’s disease (PD). However, the heterogeneity of hPSC-derived donor cells and the low yield of midbrain dopaminergic (mDA) neurons after transplantation hinder its broad clinical application. Here, we depicted the single-cell molecular landscape during mDA neuron differentiation. We found that this process recapitulated the development of multiple but adjacent fetal brain regions including ventral midbrain, isthmus, and ventral hindbrain, resulting in heterogenous donor cell population. We reconstructed the differentiation trajectory of mDA lineage and identified CLSTN2 and PTPRO as specific surface markers of mDA progenitors, which were predictive of mDA neuron differentiation and could facilitate highly enriched mDA neurons (up to 80%) following progenitor sorting and transplantation. Marker sorted progenitors exhibited higher therapeutic potency in correcting motor deficits of PD mice. Different marker sorted grafts had a strikingly consistent cellular composition, in which mDA neurons were enriched, while off-target neuron types were mostly depleted, suggesting stable graft outcomes. Our study provides a better understanding of cellular heterogeneity during mDA neuron differentiation, and establishes a strategy to generate highly purified donor cells to achieve stable and predictable therapeutic outcomes, raising the prospect of hPSC-based PD cell replacement therapies.

overall_design: We collected cells for scRNA-seq at the end of five consecutive stages during the process of mDA neuron differentiation from hPSCs (day8-stage I; day 14-stage II; day 21-stage III; day 28-stage IV; day 35-stage V).  We  sought to define the cellular composition of these grafts by scRNA-seq. To remove the contamination of host (mouse) cells in the cell suspensions when dissecting graft tissue, we knocked a nuclear EGFP expressing cassette (EGFP- nucleoplasmin) into the AAVS1 locus of surface marker reporter hPSC lines (CLSTN2- tdT hPSCs or PTPTO-tdT hPSCs) to label all of the human cells . We transplanted surface marker-sorted or unsorted mDA progenitors derived from these hPSC lines into the striatum of PD mice. Four months after transplantation, we dissected grafts and prepared single-cell suspensions. Grafted human cells (EGFP+) were then isolated via FACS and subjected to scRNA-seq analysis.

first: Peibo

last: Xu

first: Hui

last: He

first: Qinqin

last: Gao

first: Ziyan

last: Wu

first: Yingying

last: Zhou


title: TET2-mutant clonal hematopoiesis and risk of gout

city: Boston

first: Mridul

last: Agrawal

address: 450 Brookline Ave.

institute: Dana-Farber Cancer Institute

summary: Gout is a common inflammatory arthritis caused by precipitation of monosodium urate (MSU) crystals in individuals with hyperuricemia. Acute flares are accompanied by secretion of pro-inflammatory cytokines, including interleukin-1 beta (IL-1B). Clonal hematopoiesis of indeterminate potential (CHIP) is an age-related condition predisposing to hematologic cancers and cardiovascular disease. CHIP is associated with elevated IL-1B, thus we investigated CHIP as a risk factor for gout. To test the clinical association between CHIP and gout, we analyzed whole exome sequencing data from 177,824 individuals in the MGB Biobank (MGBB) and UK Biobank (UKB). In both cohorts, the frequency of gout was higher among individuals with CHIP than without CHIP (MGBB, CHIP with variant allele fraction [VAF] ≥2%: OR, 1.69; 95% CI, 1.09-2.61; P=0.0189; UKB, CHIP with VAF ≥10%: OR, 1.25; 95% CI, 1.05-1.50; P=0.0133). Moreover, individuals with CHIP and a VAF ≥10% had an increased risk of incident gout (UKB: HR, 1.28; 95% CI, 1.06-1.55; P=0.0107). In murine models of gout pathogenesis, animals with Tet2 knockout hematopoietic cells had exaggerated IL-1B secretion and paw edema upon administration of MSU crystals. Tet2 knockout macrophages elaborated higher levels of IL-1B in response to MSU crystals in vitro, and this was ameliorated through genetic and pharmacologic Nlrp3 inflammasome inhibition. These studies show that TET2-mutant CHIP is associated with an increased risk of gout in humans and that MSU crystals lead to elevated IL-1B levels in Tet2 knockout murine models. We identify CHIP as an amplifier of NLRP3-dependent inflammatory responses to MSU crystals in gout patients.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for MSU-treated bone marrow derived macrophages from Tet2 knockout and wild-type mice


title: Epigenetic profiling of mouse hematopoietic cells retrovirally transfected with MNX1 or empty vector (Ctr) from fetal liver or mouse bone marrow [ATAC-seq]

summary: Acute myeloid leukemia (AML) results from aberrant hematopoietic processes and these changes are frequently initiated by chromosomal translocations. One particular subtype, AML with translocation t(7;12)(q36;p13), is found in children diagnosed before two years of age. The chromosomal breakage points of the t(7;12) have consistently been found to be located  after exon 1 in the Motor neuron and pancreas homeobox 1 (MNX1) gene in chromosome 7, and after exon 2 in the ETV6 gene in chromosome 12. The aim of this study is the investigation of the leukomogenic potenial of MNX1 overexpression and MNX1-ETV6 fusion using mouse models, in addition to the molecular pathway through which MNX1 is inducing leukemia.

overall_design: Fetal liver hematopietic cells from C57BL/6 mice were retrovirally transfected with MNX1 and transplanted into 10-12 weeks old NOD.Cg-KitW-41J Tyr + Prkdcscid Il2rgtm1Wjl/ThomJ (NBSGW) mice. After establishment of leukmeia in the NBSGW mice, cells were collected from the bone marrow for ATAC and H3K27me3 Act-Seq. Negative control of fetal liver hematopietic cells from C57BL/6 mice  retrovirally transfected with empty vector were used.

first: Christoph

last: Plass

first: Ahmed

last: Waraky

first: Lars

last: Palmqvist


title: Bulk RNA-seq analysis of gene expression changes at 72 hpf induced by miR-34a loss

first: Sergey

last: Prykhozhij

summary: Li-Fraumeni syndrome (LFS) is a disorder due to inherited mutations in the TP53 gene resulting in an increased risk of developing several types of cancer. MicroRNA miR-34a has been implicated downstream of p53 on the basis of being a direct transcriptional target and, when over-expressed, having pro-apoptotic phenotypes in cell lines. Moreover, miR-34a has been shown to be a modifier gene in the context of LFS, since its epigenetic silencing increases the likelihood of tumour development in patients with mutant TP53.  However, the in vivo consequences of miR-34 loss are still unclear. For example, mice lacking all three (a,b,c) miR-34 homologs show no detectable abnormalities in p53 responses. The relative expression of different miR-34 genes in zebrafish was studied using qRT-PCR with specific assays. The miR-34a, miR-34b and miR-34c display unique onset of developmental expression and expression levels, with miR-34a being the most abundant and constant in expression. All of the miR-34 genes also showed clear induction by p53 when DNA-damaging treatments are performed. Using CRISPR-Cas9 technology, we generated a zebrafish miR-34a deletion mutant to further investigate the roles of miR-34a on its own and its association with the p53 pathway. Predictably, a miR-34a deletion mutant demonstrated absence of miR-34a, though without miR-34b and miR-34c compensation beyond baseline expression levels. Mutants survive to adulthood, show no overt phenotypes and have normal apoptotic responses to DNA-damaging irradiation or camptothecin treatments. To further explore the effects of miR-34a, we performed gene expression profiling using RNA-seq of wild-type and miR-34a deletion mutant zebrafish embryos at 72 hpf. We wanted to explore how miR-34a loss affects differentiated organs in larval zebrafish. This resulted in 389 genes UP and 374 genes DOWN at 1.5-fold change and FDR < 0.05. There was no significant enrichment of predicted miR-34a target genes among the differentially regulated genes but some interesting biological trends were found and will be described in the paper associated with this dataset.

overall_design: Zebrafish embryos at 72 hpf of wild-type (casper) and miR-34a-/- genotypes (3 groups of 30 embryos each) were dechorionated, flash-frozen followed by RNA extraction from all samples.

middle: V

first: Jason

last: Berman


title: Epigenetic profiling of mouse hematopoietic cells retrovirally transfected with MNX1 or empty vector (Ctr) from fetal liver or mouse bone marrow [ACT-seq]


title: FOXA2 suppresses endometrial carcinogenesis and epithelial-mesenchymal transition by regulating enhancer activity

city: Dallas

first: Ram

last: Mani

state: Texas

address: 5323 Harry Hines Blvd NB6.444

department: Pathology

institute: UT Southwestern Medical Center

summary: FOXA2 encodes a transcription factor mutated in 10% of endometrial cancers (ECs), with a higher mutation rate in aggressive variants. FOXA2 has essential roles in embryonic and uterine development. However, FOXA2’s role in EC is incompletely understood. Functional investigations using human and mouse EC cell lines revealed that FOXA2 controls endometrial epithelial gene expression programs regulating cell proliferation, adhesion, and endometrial-epithelial transition. In live animals, conditional inactivation of Foxa2 or Pten alone in endometrial epithelium did not result in ECs, but simultaneous inactivation of both genes resulted in lethal ECs with complete penetrance, establishing potent synergism between Foxa2 and PI3K signaling. Studies in tumor-derived cell lines and organoids highlighted additional invasion and cell growth phenotypes associated with malignant transformation and identified key mediators, including Myc and Cdh1. Transcriptome and cistrome analyses revealed that FOXA2 broadly controls gene expression programs through modification of enhancer activity in addition to regulating specific target genes, rationalizing its tumor suppressor functions. By integrating results from our cell lines, organoids, animal models, and patient data, our findings demonstrated that FOXA2 is an endometrial tumor suppressor associated with aggressive disease and with shared commonalities among its roles in endometrial function and carcinogenesis.

overall_design: Examination of H3K27ac in Ishikawa cells transfected with FOXA2, using the empty vector (CMV) as control.

first: Subhransu

last: Sahoo

first: Susmita

last: Ramanand

first: Yunpeng

first: Chao

last: Xing

first: Diego

last: Castrillon


title: 4T1 mouse tumour single cell dataset for virtual reality exploration

city: Cambridge

first: Dario

last: Bressan

state: Cambridgeshire

address: Li Ka Shing Centre, Robinson Way

department: CRUK Cambridge Institute

country: United Kingdom

institute: University of Cambridge

summary: 50,000 cells were injected orthotopically into the inguinal fat pad of a Nod-Scid-Gamma (NSG) immuno-compromised mouse. Injected cells were 80% unlabelled 4T1 cells (parental population), and 20% ZsGreen-labelled 4T1-T cells (clone isolated in Wagenblast et Al, Nature, 2015). Tumour were allowed to develop for 20 days, and then collected during necropsy. Disaggegated cells were processed through the 10X genomics Single Cell 3' gene expression pipeline. This data is intended as an example dataset for a novel virtual reality viewer for single-cell data described in Bressan et Al, Nat. Cancer, 2021 (submitted)

overall_design: Two replicate animals, injected at the same time from the same pool of cells. One time point

first: Gregory

middle: J

last: Hannon


first: Yuejun

last: Chen

overall_design: Comparative gene expression profiling analysis of RNA-seq data for LMX1A+EN1+ cells and control cells (LMX1A+EN1- ,LMX1A-EN1+ ,LMX1A-EN1-).

first: Peibo

last: Xu

first: Hui

last: He

first: Qinqin


title: Bulk RNA-seq on mouse model of diabetic nephropathy and in vitro model of SRSF7 knockdown

city: St. Louis

first: Haojia

last: Wu

state: Missouri

address: 600 S Euclid Ave

department: Renal Division/Internal Medicine

institute: Washington University in St. Louis

summary: In this dataset, we utilized the db/db, uninephrectomy and renin-hypertension mouse model. We performed bulk RNA-seq and compared vehicle to ACE inhibitor, Rosiglitizone, SGLT2 inhibitor, ACEi + Rosiglitizone and ACEi + SGLT2i at two time points (2 days and 2 weeks). To study the mechanism, we also performed bulk RNA-seq on human primary tubular epithelial cells with or without SRSF7 siRNA knockdown.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: We performed bulk RNA-seq to a model of murine DKD and in vitro model of SRSF7 siRNA knockdown.

first: Benjamin

last: Humphreys


title: Next Generation Sequencing of mouse hematopoietic cells retrovirally transfected with MNX1 or empty vector (Ctr) from fetal liver or mouse bone marrow and treated with or without pan-inhibitor against SAM-dependent methyltransferases Sinefungin

city: Gothenburg

first: Lars

address: Sahlgrenska University Hospital

department: Institute of Biomedicine

country: Sweden

institute: University of Gothenburg

overall_design:
Fetal liver hematopietic cells from C57BL/6 mice were retrovirally transfected with MNX1 or empty vector and transplanted into 10-12 weeks old NOD.Cg-KitW-41J Tyr + Prkdcscid Il2rgtm1Wjl/ThomJ (NBSGW) mice. After establishment of leukmeia in the NBSGW mice, cells were collected from the bone marrow for mRNA expression profile. Additional controls with fetal liver hematopietic cells from C57BL/6 mice  retrovirally transfected with empty vector were used.
For InVitro experiments, fetal liver hematopietic cells were retrovirally transfected with empty vector or MNX1 overexpression and treated with or without 5μM Sinefungin

first: Ahmed


title: Hsa_circ_0005358 suppresses cervical cancer metastasis via interacting with PTBP1 protein to destabilize CDCP1 mRNA [vector vs has_circ005358 overexpression]

city: Hangzhou

first: Yixuan

last: Cen

state: Zhejiang

address: No.1 Xue Shi Road, Hangzhou City, Zhejiang Province, China

institute: Women's Hospital School Of Medicine Zhejiang University

summary: Functional exploration of hsa_circ_0005358 in cervical cancer cells ;Mechanism of how hsa_circ_0005358 works via its protein partner PTBP1.

overall_design: To investigate downstream target mRNA of hsa_circ_0005358 we conducted RNA-seq in SiHa cells.Seq: vector vs has_circ005358 overexpression


title: Transplanted human organoids empower PK/PD assessment of drug candidate for the clinic

first: Eva

last: Fast

state: MA

address: 1 Portland st

institute: Pfizer

summary: Pharmacokinetic/pharmacodynamic (PK/PD) studies are an essential component of pre-clinical drug discovery. Current best approaches for PK/PD studies, including the analysis of novel kidney disease targeting therapeutic agents, are limited to animal models with unclear translatability to the human condition. To address this challenge, we developed a novel approach for PK/PD studies using transplanted human kidney organoids. We performed PK studies with GFB-887, an investigational new drug now in Phase 2 trials. Orally dosed GFB-887 to athymic rats that had undergone organoid transplantation resulted in measurable drug exposure in human transplanted organoids. Importantly, we established the efficacy of orally dosed GFB-887 in PD studies, where quantitative analysis showed significant protection of kidney filter cells in human organoids and endogenous rat host kidneys. This widely applicable approach demonstrates, for the first time, feasibility of using transplanted human organoids in PK/PD studies, empowering organoids to revolutionize drug discovery.

sample_organism:
Homo sapiens
Rattus norvegicus

platform_organism:
Homo sapiens
Rattus norvegicus

overall_design: scRNAseq profiles of in vitro and in vivo matured IPSC derived organoids

first: Amy

middle: D

last: Westerling-Bui

first: Peter

last: Mundel


title: Transcriptome profiling of  control cells or the A549 cells in the presence or absence of acetate following TGF-β1 stimulation

city: St. Petersburg

first: Jianhua

last: Xiong

state: Florida

address: 600 5th Street S

department: Medicine

institute: Johns Hopkins University

summary: Purpose: To compare the transcriptome profiling of control cells or the A549 cells in the presence or absence of acetate following TGF-β1 stimulation.  Methods:  A549 cells were treated with control, TGF-β1 or TGF-β1 plus acetate for 2 days. And then, we performed RNA sequencing for transcription profiling of control cells or the A549 cells in the presence or absence of acetate following TGF-β1 stimulation. The raw data were processed with bcl2fastq software and HISAT2.  Results: From the transcriptome profiling and analysis, we got gene expression data of control group and in the presence or absence of acetate following TGF-β1 stimulation using A549 cells.  Conclusion: The gene expression data revealed the transcriptome variation by TGF-β1 treatment and acetate can reverse TGF-β1 effect.

overall_design: A549 cells were treated with control, TGF-β1 or TGF-β1 plus acetate. And then, we performed RNA sequencing for transcription profiling of control cells or the A549 cells in the presence or absence of acetate following TGF-β1 stimulation. The raw data were processed with bcl2fastq software and HISAT2.

first: Junfang

last: Lyu


title: A NEW PRECLINICAL MODEL OF WESTERN DIET-INDUCED PROGRESSION OF NON-ALCOHOLIC STEATOHEPATITIS TO HEPATOCELLULAR CARCINOMA

city: Richmond

first: Mikhail

last: Dozmorov

state: VA

address: 830 E Main St

department: Biostatistics

institute: Virginia Commonwealth University

summary: Non-alcoholic steatohepatitis (NASH) results from accumulation of excessive liver lipids leading to hepatocellular injury, inflammation, and fibrosis that greatly increase the risk for hepatocellular carcinoma (HCC). Despite the well-characterized clinical and histological pathology for NASH-driven HCC in humans, its etiology remains unclear and there is a deficiency in pre-clinical models that recapitulate the progression of the human disease. Therefore, we developed a new mouse model amenable to genetic manipulations and gene targeting that mimics the gradual NASH to HCC progression observed in humans. C57BL/6NJ mice were fed a Western high fat diet and sugar water (HFD/SW) and monitored for effects on metabolism, liver histology, tumor development, and liver transcriptome for up to 54 weeks. Chronic HFD/SW feeding led to significantly increased weight gain, serum and liver lipid levels, liver injury, and glucose intolerance. Hepatic pathology progressed and mice developed hepatocellular ballooning, inflammation, and worse fibrosis was apparent at 16 weeks, greatly increased through 32 weeks, and remained elevated at 54 weeks. Importantly, hepatocellular cancer spontaneously developed in 75% of mice on HFD/SW, half of which were HCC, whereas none of the mice on chow diet developed HCC. Chronic HFD/SW induced molecular markers of de novo lipogenesis, endoplasmic reticulum stress, inflammation, and accumulation of p62, all of which also participate in the human pathology. Moreover, transcriptome analysis revealed activation of HCC-related genes and signatures associated with poor prognosis of human HCC. Overall, we have identified a new preclinical model that recapitulates known hallmarks of NASH-driven HCC that can be utilized for future molecular mechanistic studies of this disease.

overall_design: Liver mRNA profiles from mice treated with chow or high fat diet plus sugar water for 54 weeks.

first: Christopher

last: Green

first: Cynthia

last: Weigel

first: Ryan

last: Brown

first: Pierre

last: Bedossa

first: Arun

last: Sanyal

first: Sarah

last: Spiegel


title: Hsa_circ_0005358 suppresses cervical cancer metastasis via interacting with PTBP1 protein to destabilize CDCP1 mRNA


title: Hsa_circ_0005358 suppresses cervical cancer metastasis via interacting with PTBP1 protein to destabilize CDCP1 mRNA [si-oligo vs si-PTBP1]

summary: Function exploration of hsa_circ_0005358 in cervical cancer cells ;Mechanism of how hsa_circ_0005358 works via its protein partner PTBP1.

overall_design: To investigate the downstream target mRNA of  PTBP1, we conducted RNA-seq in SiHa cells.Seq:si-oligo vs si-PTBP1;


title: Mapping the Single Cell Transcriptomic Response of Murine Diabetic Kidney Disease to Therapies

first: Haojia

summary: In this study, we utilized the db/db, uninephrectomy and renin-hypertension mouse model. We compared vehicle to ACE inhibitor, Rosiglitizone, SGLT2 inhibitor, ACEi + Rosiglitizone and ACEi + SGLT2i at two time points (2 days and 2 weeks). We generated snRNA-seq datasets from all groups comprising nearly 1 million cells. Analysis of the resulting atlas revealed that the different medications affected strikingly different cell types. Combination therapy had the largest effects, but these effects were also largely non-overlapping. This study highlights the cellular complexity of diabetic nephropathy and the effects of different classes of therapy. The results support the development of combination therapies since different drug classes target different cell types in kidney.

overall_design: We employed single nucleus RNA-sequencing (snRNA-seq) to a model of murine DKD and five treatment regimens. We profiled a total of 70 mice from 14 groups, with N=4-6 mice per group.


title: Effect of IL1β on A498 ccRCC  cell lines

status: Public on Jun 13 2022

city: Milano

first: Michela

last: Riba

address: via Olgettina 60

department: COSR, Center for Omics Sciences

country: Italy

institute: IRCCS, Ospdale San Raffaele

summary: Renal cell carcinoma is highly inflamed and tumor cells are embedded into a microenvironment enriched of IL1. While inflammatory pathways are well characterized in the immune system, less is known about these same pathways in epithelial cells; it is unclear if and how innate immune signals directly impact on cancer cells, and if we could we manipulate these for therapeutic purpose.  To address these questions, we first focused on the inflammatory receptors belonging to the IL1- and Toll-like receptors family including negative regulators in a small cohort of 12 clear cell RCC (ccRCC) patients’ samples as compared to their coupled adjacent normal tissues. Our data demonstrated that renal epithelial cancer cells showed a specific and distinctive pattern of inflammatory receptors expression marked by a consistent downregulation of the inhibitory receptor SIGIRR mRNA. This repression was confirmed at the protein level in both cancer cell lines and primary tissues. When we analyzed in silico data of different kidney cancer histotypes, we identified the clear cell subtype as the one where SIGIRR was mostly downregulated; nonetheless, papillary and chromophobe tumor types also showed low levels as compared to their normal counterpart.  RNA-sequencing analysis demonstrated that IL1 stimulation of the ccRCC cell line A498 triggered an intrinsic signature of inflammatory pathways activation characterized by the induction of distinct “pro-tumor” genes including several chemokines, the autocrine growth factor IL6, the atypical co-transcription factor NFKBIZ and the check-point inhibitor PD-L1. When we looked for the macroareas most represented among the differentially expressed genes, additional clusters emerged including pathways involved in cell differentiation, angiogenesis and wound healing. To note, SIGIRR over-expression in A498 cells dampened IL1 signaling as assessed by a reduced induction of NFKBIZ, IL6 and PD-L1. Our results suggest that SIGIRR downregulation unleashes IL1 signaling intrinsic to tumor cells and that manipulating this pathway may be beneficial in ccRCC.

overall_design: 4x A498 ccRCC  cell lines untreated or exposed for 4 hrs to IL1β

first: Maria

middle: E

last: Mantione

first: Ilenia

last: Sana

middle: G

last: Vilia

first: Claudio

last: Doglioni

first: Alessandro

last: Larcher

first: Umberto

last: Capitanio

first: Marta

last: Muzio


title: e-CLIP

city: Cologne

first: Leo

last: Kurian

address: Robert-Kochstr. 21

institute: University of Cologne

summary: To RBPMS RNA targts in hESCs, eCLIP-seq (Van Nostrand et al., 2016) was performed in 4 biological replicates

overall_design: Cell culture dishes containing around 20M hESCs were UV-crosslinked with (400 mJ/cm2 constant energy) to stabilize protein-RNA interaction. IP was performed with a RBPMS-specific antibody.

first: Deniz

last: Bartsch


title: Gene expression profile in mock or virus-infected SOD1-G85R transgenic mice, a mouse model of amyotrophic lateral sclerosis

city: Vancouver

first: Honglin

last: Luo

state: British Columbia

address: Room 166, 1081 Burrard Street

institute: University of British Columbia

summary: Profilling of inflammatory-related genes signatures that contributes to the progression of ALS in SOD1-G85R C57BL/6 mice model upon mock or viral infection.

overall_design: In the study presented here, spinal cord from week-20, or whole brain from week-20 and week-60 of Mock or CVB3 infected WT or SOD1-G85R transgenic mice was used to acquire expression profiles of a total of ≈600 unique genes


title: Expression profile of oral tumors from KR versus KHR mice treated with tamoxifen

city: Chicago

first: John

last: Bechill

state: IL

department: Radiation Oncology

institute: The University of Chicago

summary: kRas (KR) tumors grow faster when expressing kRas and HPV E6E7 (KHR). The hypothesis was to test for differences in gene expression between KR and KHR oral tumors.

overall_design: Total RNA was extracted from mouse KR + tamoxifen  (n=6) and KHR + tamoxifen (n=6) tumors. Expression profiles were examined by Affymetrix Mouse Gene ST 2.0 exon array (GPL18233). Studies were conducted from oral tumor formed in C57BL/6 × FVB F1 mice.

first: Michael

last: Spiotto


title: Transcriptomic points of departure calculated from rainbow trout gill, liver, and gut cell lines exposed to methylmercury and fluoxetine

city: Sainte-Anne-de-Bellevue

first: Krittika

last: Mittal

state: Quebec

address: 21,1111 Lakeshore Blvd, Apt #2

institute: McGill University

summary: The primary objective of this study was to determine if quantitative tPOD values could be derived from chemical exposure studies that followed OECD Test No. 249 (rainbow trout gill cell line) and expanding the assay to rainbow trout liver and intestinal epithelial cell lines. The secondary objective was to determine if tPOD values could be derived from similar studies on rainbow trout liver and gut cell lines.  Methylmercury and fluoxetine were the test chemicals. From the resulting data, we compared tPOD data with literature-derived apical BMDs, characterized mechanisms of action through analysis of the transcriptomic data, and compared responses (cytotoxicity and transcriptomic) across the three cell types. By bringing together transcriptomics measurements and tPOD calculations with the OECD Test, and expanding the number of cell lines, we believe that this work can help establish a cost effective in vitro test method that can yield quantitative tPOD values that are protective of in vivo concentrations associated with adverse outcomes while also advancing knowledge of a test chemical’s  mechanism of action.

overall_design: RNA Sequencing analysis of rainbow trout gill, liver, and gut cell lines exposed to methylmercury (in duplicate) and fluoxetine (in single replicates)

first: Jessica

last: Ewald

first: Niladri

last: Basu


title: Fibroblast-specific reduction of heparan sulfate suppresses colon tumor growth in mice

city: Gifu City

first: Ayumi

last: Niwa

address: 1-1 Yanagido

institute: Gifu University

summary: we examined how HS-reduced fibroblasts are involved in the proliferation of colon cancer by using mice in which Ext-1 expression is specifically suppressed in fibroblasts.

overall_design: We prepared mouse colon cancer (MC38) cells. We transplanted the cells to wild type (WT) and Ext1 flox/flox; S100a4-Cre mice (N=4 each). We removed the developing tumors from mouse subcutaneous area  25 to 30 days later and extracted RNA from whole tumors. Finally, we performed RNA microarray analysis.

first: Hiroyuki

last: Tomita


title: Fibroblast-specific reduction of heparan sulfate suppresses colon tumor growth and pancreatic cancer growth in mice

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

overall_design: Refer to individual Series


title: Inhibition of PGE2-EP2/EP4 signaling elicits anti-tumor immunity through reprograming of inflammatory myeloid cells and   suppression of mRegDC-Treg axis

city: Kyoto

first: Shuh

last: Narumiya

state: Kyoto

address: Yoshidahonmachi

department: Department of Drug Discovery Medicine

institute: Kyoto University

summary: The discovery of immune checkpoint inhibitor (ICI) has highlighted the clinical importance of immune evasion in cancer. However, only a fraction of cancer patients show response to ICI, raising a question on immune suppression mechanisms other than immune checkpoint. In this study, we examined the role of the lipid inflammatory mediator PGE2 in immune evasion of the ICI-insensitive Lewis Lung Carcinoma line 1 (LLC1) mouse model. Inhibition of PGE receptors, EP2 and EP4, significantly suppressed tumor growth through the modulation of host immune cells. Single cell RNA-sequencing analysis revealed that EP2/4 inhibition elicited anti-tumor immunity through the reprogramming of inflammatory myeloid cells by downregulating expression of genes in NFB signaling and actions and suppression of the mregDC-regulatory T cell axis by downregulating genes associated with regulatory T cell recruitment and activation. Taken together, our work suggests that PGE2-EP2/EP4 signaling induces proinflammatory myeloid and tolerogenic lymphoid environments in ICI-insensitive tumors, which is amenable to EP2 and EP4 inhibitors..

overall_design: Using 10x genomics to measure single-cell RNA sequence (scRNA-seq) to comprehensively characterize the tumor immune microenvironment in mouse LLC1 tumor syngeneic transplantation model from EP2 and EP4 antagonists treatment and Treg depletion.

first: Siwakorn

last: Punyawatthananukool

first: Dean

last: Thumkeo


title: Fibroblast-specific reduction of heparan sulfate suppresses pancreatic cancer growth in mice

summary: we examined how HS-reduced fibroblasts are involved in the proliferation of pancreatic cancer by using mice in which Ext-1 expression is specifically suppressed in fibroblasts.

overall_design: We prepared mouse pancreatic cancer (Pan02) cells. We transplanted the cells to wild type (Control) and Ext1 flox/flox; S100a4-Cre mice  (Mutant) (N=4 each). We removed the developing tumors from mouse subcutaneous area  36 to 40 days later and extracted RNA from whole tumors. Finally, we performed RNA microarray analysis.


title: TS-seq

summary: To investigate the effect of RBPMS KO on mRNA ribosomes occupancy, polysome profiling was performed on lysates from WT and RBPMS KO hESCs.

overall_design: Ribosomes were stalled on mRNAs by using cycloheximide and ribosomal fractions were isolated using polysome profiling. RNA was isolated using Trizol and equal amounts were used for library generation for stranded, paired-end RNA sequencing.


title: UBP12 and UBP13 deubiquitinases destabilize the CRY2 blue light receptor to regulate Arabidopsis growth

city: Cold Spring Harbor

first: Ullas

middle: V.

last: Pedmale

state: NY

address: 1 Bungtown RD

department: Pedmale

institute: Cold Spring Harbor Laboratory

summary: Light is a crucial exogenous signal sensed by cryptochrome (CRY) blue light receptors to modulate growth and the circadian clock in plants and animals. However, how CRYs interpret light quantity to regulate growth in plants remains poorly understood. Furthermore, CRY2 protein levels and activity are tightly regulated in light to fine-tune hypocotyl growth; however, details of the mechanisms that explain precise control of CRY2 levels are not fully understood. We show that in Arabidopsis, UBP12 and UBP13 deubiquitinases physically interact with CRY2 in light. UBP12/13 negatively regulates CRY2 by promoting its ubiquitination and turnover to modulate hypocotyl growth. Growth and development were explicitly affected in blue light when UBP12/13 were disrupted or overexpressed, indicating their role alongside CRY2. UBP12/13 also interacted with and stabilized COP1, which is partially required for CRY2 turnover. Our combined genetic and molecular data support a mechanistic model in which UBP12/13 interact with CRY2 and COP1, leading to the stabilization of COP1. Stabilized COP1 then promotes the ubiquitination and degradation of CRY2 under blue light. Despite decades of studies on deubiquitinases, the knowledge of how their activity is regulated is limited. Our study provides insight into how exogenous signals and ligands, along with their receptors, regulate deubiquitinase activity by protein-protein interaction. Collectively, our results provide a framework of cryptochromes and deubiquitinases to detect and interpret light signals to control plant growth at the most appropriate time.

overall_design: compartive gene expression profiling analysis of RNA-seq data on Arabidopsis thaliana Col-0 (WT), cry2, CRY2oe, UBP13oe, and ubp12ubp13 genotypes

first: Louise

last: Lindbäck


title: UVB-induced skin autoinflammation due to NLRP1 mutation and its inhibition by anti-IL-1b antibody

city: Nagoya

first: yuya

last: murase

address: 65 Tsurumai-cho, Showa-ku

institute: Nagoya University Graduate School of Medicine

summary: To analyze the pathogenic mechanism of the autoinflammatory skin syndromes caused by an NLRP1 mutation, we sought to generate Nlrp1b mutation knock-in mice mice as a mouse model of cutaneous autoinflammatory lesions due to NLRP1 mutants, and examine the expression of proteins, such as cytokines, and the mRNA expression profile, including inflammasome-related genes in the model lesions.

overall_design: RNA sequencing was performed using extracted RNA from the skin of 5-week-age Nlrp1b knock in (KI) mice (homo and hetero) and wild type (WT) mice before and after UVB irradiation. We performed a gene set enrichment analysis using the hallmark gene set database.

first: Yuya

last: Murase

first: Takuya

last: Takeichi

first: Jun

last: Koseki

first: Yuki

last: Miyasaka

first: Tamio

last: Ohno

first: Teppei

last: Shimamura

first: Masashi

last: Akiyama


title: BRD2 interconnects with BRD3 to facilitate Pol II transcription initiation and elongation to prime promoters for cell differentiation [ChIP-seq]

city: Beijing

first: Xiong

last: Ji

address: Haidian District, Beijing Summer Palace Road No. 5

institute: Peking University

summary: Understanding the precise functions and relationship of BRD2 with other bromodomain and extraterminal motif (BET) proteins is central for the application of BET-specific and pan inhibitors. Here, we used acute protein degradation and quantitative genomic and proteomic approaches to investigate the primary functions of BRD2 in transcription. We report that BRD2 is required for TAF3-mediated Pol II initiation at low levels of H3K4me3-modified promoters and Pol II elongation by suppressing R-loops. Single and double depletion revealed that BRD2 and BRD3, but not BRD4, redundantly and independently function in Pol II transcription at different promoters and cooperatively occupy enhancers. Interestingly, we found that depletion of BRD2 affects the expression of different genes during differentiation processes, priming with promoter regulation in ES cells. Therefore, our results suggest complex interconnections between BRD2 and BRD3 at promoters to fine-tune Pol II initiation and elongation for control of cell state.

overall_design: We have finished the ChIA-PET, ChIP-seq, RNA-seq and 4C-seq to investigate the roles of BRD2 in transcription initiation. The V6.5 mouse ES (mES) cell line was used to perform all the high-throughput analysis. BRD2-degron mES cells were pre-treated with 1 μg/ml doxycycline for 18 hours and treated with or without 500 μM indole-3-acetica cid for 24 hours.

first: Chenlu

last: Wang

first: Qiqin

last: Xu

first: Jie

last: Huang


title: BRD2 interconnects with BRD3 to facilitate Pol II transcription initiation and elongation to prime promoters for cell differentiation [ChIA-PET]


title: BRD2 interconnects with BRD3 to facilitate Pol II transcription initiation and elongation to prime promoters for cell differentiation [4C-seq]

overall_design: We have finished the ChIA-PET, ChIP-seq, RNA-seq and 4C-seq to investigate the roles of BRD2 in transcription initiation. The V6.5 mouse ES (mES) cell line was used to perform all the high-throughput analysis. BRD2-degron mES cells were pre-treated with 1 μg/ml doxycycline for 18 hours and treated with or without 500 μM indole-3-acetic acid for 24 hours.


title: Pu-miR172d negatively regulates stomatal differentiation response to drought stress in poplar

city: Harbin City

first: Chenghao

last: Li

state: HeiLongJiang Province

address: No. 51 HeXing Road,Xiangfang District

department: College of Forestry

institute: Northeast Forestry University

summary: We characterized Pu-miR172d, a miRNA that negatively regulates stomatal differentiation in Populus ussuriensis. Overexpression of Pu-miR172d significantly decreased stomatal density in P. ussuriensis. Molecular analysis indicated that PuGTL1 as a major target of Pu-miR172d by cleavage. Moreover, chimeric dominant repressor version of PuGTL1 (PuGTL1-SRDX) lines resembled the Pu-miR172d overexpression (Pu-miR172d-OE) lines, resulting in reduced stomatal density, net photosynthetic rate, stomatal conductance, transpiration and enhanced instantaneous water use efficiency, and thus improving tolerance to drought stress. QRT-PCR analysis showed that PuSDD1 transcript abundance in young leaves of PuGTL1-SRDX and Pu-miR172d-OE plants compared to WT plants, suggesting that Pu-miR172d positively regulates PuSDD1 expression through repression of PuGTL1. RNA-seq analysis showed Pu-miR172d overexpression significantly reduced the expression of many genes related to photosynthesis under drought stress. Overall, the present results demonstrate that Pu-miR172d/PuGTL1/PuSDD1 module plays an important role in stomatal differentiation and regulate WUE, illustrating its capacity for engineering drought tolerance improvement in poplar under future drier environments.

overall_design: The 7th and 8th leaves from shoot tips were harvested from 70-day-old Pu-miR172d-OE and WT plants transplanted in soil growing under well-watered and drought treatment (4 days) conditions were used for transcriptome analysis. Total RNA isolation, library construction and sequencing were performed by Annoroad Gene Technology Corporation (Beijing, China). A total amount of 2 μg RNA per sample was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (#E7530L, NEB, USA) following the manufacturer’s recommendations and index codes were added to attribute sequences to each sample. The library preparations were sequenced on an Illumina Hiseq 2500 platform and paired-end reads were generated. About 4 Gb of high-quality 150-bp paired-end reads were generated from each library. The DEGs were defined according to the following thresholds: fold change ≥ 1, false discovery rate (FDR) < 0.05.

first: Quangang

last: Liu

first: Zhanchao


title: Expression profiling of retinoic acid targets in lateral mesoderm from Xenopus laevis embryos

city: PARIS

first: Jean-Francois

last: RIOU

address: 9, quai Saint-Bernard, case 24

department: Institut de Biologie Paris-Seine

country: France

institute: CNRS/Sorbonne Universite

summary:
Renal precusors of the Xenopus pronephros arise from dorso-lateral mesoderm at the early neurula stage. This process is under the control of retinoic acid (RA).
We have used microarrays to identify RA targets in dorso-latearl mesoderm by performing differential expression analysis between control and RA-depleted situations

overall_design: mRNA encoding GFP (control) or a mix of GFP and RA-catabolizing enzyme cyp26 (RA-depleted) were microinjected in 4-cell stage embryos. Lateral marginal zone (LMZ) explants were dissected at the early gastrula stage and cultured for 1h (early gastrula), 3 h (mid-gastrula) or overnight (early neurula)

last: Riou

first: Ronan

last: LeBouffant

first: Muriel

last: Umbhauer


title: BRD2 interconnects with BRD3 to facilitate Pol II transcription initiation and elongation to prime promoters for cell differentiation


title: BRD2 interconnects with BRD3 to facilitate Pol II transcription initiation and elongation to prime promoters for cell differentiation [RNA-seq]


title: Effects of Magnesium Nutrition on Soybean Growth and Transcriptomic Response to Arbuscular Mycorrhizal Fungal Inoculation

city: Guangzhou

first: Jinzhuan

last: Qin

address: Wushan Road

institute: South China Agricultural University

summary:
In the study, two soybean genotypes were selected to conduct high and low P, high and low Mg, and AM fungal inoculation treatments, combined with RNA-seq sequencing technique in order to investigate the physiological and molecular mechanisms of the symbiosis between soybean and AM fungi affected by P and Mg treatments.  The results showed that both Mg application and AM fungal inoculation were beneficial to promote soybean growth under low P condition. And there was a synergistic effect between the Mg concentration and the P concentration in the root of HN112 under the inoculation condition.
RNA-seq sequencing was carried out using the roots of P-efficient soybean HN89 under different Mg and inoculation treatments with low P condition, and the difference of gene expression profiles was analyzed between high and low Mg treatments, and different inoculation treatments. According to the analysis of GO function classification and KEGG enrichment, under high Mg condition, the metabolic pathway was mainly enriched in lipid metabolism and glucose metabolism pathway under the inoculation treatment compared with the non-inoculation treatment, which regulated carbon metabolism pathway. Under the low Mg condition, the metabolic pathway was mainly enriched in the photosynthesis- antenna protein pathway to regulate the photosynthesis pathway under the inoculation treatment compared with the non-inoculation treatment. At the same time, the inoculation treatment significantly increased soybean root starch concentration under low Mg condition, compared with the non-inoculation treatment, suggesting that the significant up-regulation of a large number of photosynthesis related genes might be related to the significant increase of starch concentration at this treatment.

overall_design: we employed pot experiment and RNA-seq sequencing technology to investigate the effects of Mg nutrition on plant growth, P and Mg contents, and root transcriptomes in soybean plants inoculated with AM fungi, every treatment had three replicates.

first: Xiurong


title: Differential transcriptomic changes in the central nervous system and urinary bladders of mice infected with a coronavirus

status: Public on Jun 11 2022

city: Aurora

first: Anna

last: Malykhina

state: CO

institute: University of Colorado

summary: Gene expression of 770 genes from the Nanostring Neuroinflammation panel in the coronavirus-induced encephalomyelitus (CIE) murine model of multiple sclerosis.

overall_design:
In the study presented here, spinal cords and bladders were collected from 4 control mice, 4 CIE mice at week 1, and 4 CIE mice at week 4.
Two spinal cord samples (SC 406 Control and SC 455 CIE 4 wk) were excluded due to QC flags.

first: Taylor

last: Clarkson


title: Gene expression analysis of PJA1 knock down HepG2 cells or HepG2 cells treated with TGF-beta1.

city: Houston

first: Jian

last: Chen

state: TX

address: 1515 Holcombe Boulevard - Unit 1466

institute: The University of Texas M. D. Anderson Cancer Center

summary: Gene expression profiling was carried out in four (HepG2-shCtrl,HepG2-shPJA1, HepG2/Ctrl and HepG2/TGFb1 cells) different liver cancer cells.  PJA1 is an E3 ligase that regulates TGF-beta signaling pathway. In this study, we report a major role of the PJA1-mediated desregulation of TGF-beta/Smad3 in liver cancers . To determine the mechanism for the oncogenic function of PJA1 in TGF-β signaling transduction, we analyzed gene profiles the cells of knockdown PJA1 in HepG2 cells and the cells treated with TGFb1 in HepG2.

first: Lopa

last: Mishra


title: Oviductal Glycoprotein 1 Promotes Hypertension by Inducing Vascular Remodeling through an Interaction with MYH9

status: Public on Jun 12 2022

city: Peking

first: Zhengqin

last: Rong

address: Beiqing Road

institute: Basepedia

summary: Hypertension is a common cardiovascular disease that is related to genetic and environmental factors, but its mechanisms remain unclear. DNA methylation, a classical epigenetic modification, not only regulates gene expression but is also susceptible to environmental factors, linking environmental factors to genetic modification. Therefore, globally screening differential genomic DNA methylation in hypertensive patients is important for investigating novel hypertension mechanisms.Differential genomic DNA methylation in hypertensive, prehypertensive, and healthy control individuals was screened using the Illumina 450K BeadChip and verified by pyrosequencing. Plasma oviduct glycoprotein 1 (OVGP1) levels were determined using an enzyme-linked immunosorbent assay. Ovgp1 transgenic and knockout mice were generated to analyze the function of OVGP1. The blood pressure levels of the mouse models were measured using the tail-cuff system and radiotelemetry methods. The role of OVGP1 in vascular remodeling was determined by vascular relaxation studies. Protein–protein interactions were investigated using a pull-down/mass spectrometry assay and verified with coimmunoprecipitation and pull-down assays. We found a hypomethylated site at cg20823859 in the promoter region of OVGP1, and the plasma OVGP1 levels were significantly increased in hypertensive patients. This finding indicates that OVGP1 is associated with hypertension. In Ovgp1 transgenic mice, OVGP1 overexpression caused an increase in blood pressure, dysfunctional vasoconstriction and vasodilatation, remodeling of arterial walls, and increased vascular superoxide stress and inflammation, and these phenomena were exacerbated by angiotensin II infusion. In contrast, Ovgp1 deficiency attenuated angiotensin II-induced vascular oxidase stress, inflammation, and collagen deposition. These findings indicate that OVGP1 is a prohypertensive factor that directly promotes vascular remodeling. Pull-down and coimmunoprecipitation assays showed that myosin heavy chain II-A (MYH9) interacted with OVGP1, whereas inhibition of MYH9 attenuated OVGP1-induced hypertension and vascular remodeling.  CONCLUSIONS: Hypomethylation at cg20823859 in the promoter region of OVGP1 is associated with hypertension and induces upregulation of OVGP1. The interaction between OVGP1 and MYH9 contributes to vascular remodeling and dysfunction. Therefore, OVGP1 is a prohypertensive factor that promotes vascular remodeling by binding with MYH9.

overall_design: Artery mRNA profiles of 12 weeks old Ovgp1 knock out (KO) and wild type (WT) mice

first: Congxia

last: Bai

first: Ming

last: Su

first: Jingzhou

first: Hao


title: Genome-wide maps of rH2A.X in wildtype and DNMT3b-KO HCT116 lines

city: Taipei

first: Wei-Yi

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

department: Institute of Biochemistry and Molecular Biology

country: Taiwan

institute: National Yang Ming Chiao Tung University

summary: Dysfunction of DNA methyltransferase 3b (DNMT3b) causes centromere instability but the underlying mechanism is unclear. We found that enforced expression of RNase H1 that removes R-loops, nucleic structures comprising an DNA-RNA hybrid, was sufficient to abolish DNA double-strand breaks (DSBs) at (peri-)centromeric sites in immunodeficiency-centromeric instability-facial anomalies (ICF) patient cells carrying DNMT3b mutation. However, ICF cells had lower steady-state level of centromeric R-loops than normal cells. Simultaneous knockdown of two DNA endonucleases, XPG and XPF, restored centromeric R-loops in ICF cells while reducing DSBs and chromosome segregation error. This suggests that (peri-)centromeric R-loops are more vulnerable to XPG or XPF in ICF cells, thus increasing centromeric breaks. This mechanism is recapitulated in DNMT3b-knockout HCT116 cells. Moreover, we present evidence for the choice of alternative end-joining (alt-EJ) repair of (peri-)centromeric breaks in ICF cells. Thus, DNA cleavages of (peri-)centromeric R-loops and mutagenic alt-EJ repair undermine centromere stability in DNMT3b defective cells.

overall_design: Genome-wide binding/occupancy profiling of rH2A.X by ChIP-seq in wildtype and DNMB3b-KO HCT116 cells


title: SS18/BAFs regulate chromatin accessibility dynamics during pluripotent-somatic transition

city: hangzhou

first: kuang

last: junqi

address: No.18 Shilongshan Road

institute: westlake University

summary: The exit from pluripotency or pluripotent-somatic transition (PST) landmarks an event of mammalian development, and is also a representative cell-fate transition model, but remains largely unresolved. Recently, we reported construction of robust JUN-induced PST completed in one cell cycle and whose dominant regulator SS18/BAFs (Brg1/Brahma-associated factors). However, the transition process in the chromatin architecture and the roles played by BAF are still unknown. Here we report the dynamic changes of chromatin accessibility during JUN-induced PST. Meanwhile, SS18/BAFs mediates PST process by relocating from pluripotent loci to AP-1 associated ones and once compromised, JUN fails to open chromatin and PST will be delayed. Furthermore, we show that the relocation of SS18/BAF partially relays on histone modification H3K27ac, instead of JUN-centric protein-protein interaction. These results reveal the orchestration of master transcription factor, epigenetic machine, and histone modification in the cell fate transition.

overall_design: We analyzed the transcriptome expression profiles (RNA-seq) in WT and bromodomain inhibitors treatment mESCs during the process of JUN triggered differentiation

first: Junqi

last: Kuang

first: Duanqing

last: Pei

first: Jing


title: Single Cell sequencing of mononuclear cells from regenerating skeletal muscle

city: Boulder

first: Jesse

last: Kurland

address: 1945 Colorado Ave

department: MCDB

institute: University of Colorado Boulder

summary: Skeletal muscle stem cells, or satellite cells (SCs), are essential to regenerate and maintain muscle. Quiescent SCs reside in an asymmetric niche between the basal lamina and myofiber membrane. To repair muscle, SCs activate, proliferate, and differentiate, fusing to repair myofibers or reacquiring quiescence to replenish the SC niche. Little is known about when SCs reacquire quiescence during regeneration or the cellular processes that direct SC fate decisions and progression through myogenesis. Single cell sequencing of myogenic cells in regenerating muscle identifies SCs reacquiring quiescence and reveals that non-cell autonomous signaling networks influence SC fate decisions during regeneration. Single cell RNA-sequencing of regenerating skeletal muscle reveals that RBP expression, including numerous neuromuscular disease-associated RBPs, is temporally regulated in skeletal muscle stem cells and correlates to stages of myogenic differentiation. By combining machine learning with RBP engagement scoring, we discover that the neuromuscular disease associated RBP Hnrnpa2b1 is a differentiation-specifying regulator of myogenesis controlling myogenic cell fate transitions during terminal differentiation.

overall_design: Mononuclear cells were isolated from mouse tibialis anterior muscle at 4 and 7 days following injury with Barium Chloride. The muscle was mechanically dissociated and digested with collagenase prior to affinity based removal of debris and dead cells. The isolated cells were barcoded for seqeuncing using the Chromium Next GEM Single Cell 3' Reagent kit v2.

first: Bradley

middle: B

last: Olwin

first: Alicia

middle: A

last: Cutler

last: Pawlikowski

first: Joshua

last: Wheeler


title: Cabozantinib enhances anti-PD1 efficacy and elicits a neutrophil-based immune response in murine models: implications for human HCC

city: Barcelona

first: Roger

last: Esteban-Fabró

state: Barcelona

address: Rosselló, 149-153

country: Spain

institute: IDIBAPS

summary: Background & Aims: Immune checkpoint inhibitors combined with anti-angiogenic agents produces benefits in the treatment of advanced hepatocellular carcinoma (HCC). We investigated the efficacy and immunomodulatory activity of cabozantinib alone and combined with anti-PD1 in experimental models of HCC, and explored the potential target population that might benefit from this combination. Approach & Results: C57BL/6J mice bearing subcutaneous Hepa1-6 or Hep53.4 tumours received cabozantinib, anti-PD1, their combination or placebo. Tumour and blood samples were analysed by flow cytometry, immunohistochemistry, transcriptome and cytokine profiling. Cabozantinib-related effects were validated in a colorectal cancer PDX model. Transcriptomic data from three human HCC cohorts (Cohort 1: n=167, Cohort 2: n=57, TCGA: n=319) were used to cluster patients according to neutrophil features, and assess their impact on survival. The combination of cabozantinib and anti-PD1 showed increased anti-tumour efficacy compared to monotherapy and placebo (P<0.05). Cabozantinib alone significantly increased neutrophil infiltration and reduced CD8+PD1+ T cell proportions in the tumour, while the combination with anti-PD1 further stimulated both effects and significantly decreased T regulatory cell infiltration (all P<0.05). In blood, cabozantinib and especially combination increased the proportions of overall T cells (P<0.01) and memory/effector T cells (P<0.05), while lowering the neutrophil-to-lymphocyte ratio (P<0.001 for combination). Unsupervised clustering of human HCCs revealed that high tumour enrichment in neutrophil features observed with the treatment combination was linked to less proliferative phenotypes, and well-differentiated HCC with better prognosis. Neutrophil recruitment in both humans and mice was linked to CXCR2 ligands and CXCL12 (P<0.05). Conclusions: Cabozantinib in combination with anti-PD1 enhanced anti-tumour immunity by bringing together innate neutrophil-driven and adaptive immune responses, a mechanism of action which favours this approach for the treatment of HCC.

overall_design: Upon patients' consent, we obtained human HCC and non-tumour adjacent tissue samples from 57 resected patients with preserved liver function (Child-Pugh A). Transcriptomic data obtained from Human Genome U219 Array (Affymetrix) was used to cluster samples according to enrichment in active neutrophil profiles in those tissues and to identify key molecular features associated with the tumour biology among the groups that were generated.

first: Josep

last: Llovet

first: Catherine

last: Willoughby


title: miRNA expression in EBCs

city: Bloomington

first: Patricia

last: Silveyra

email: psilveyr@iu.edu, ptrcslvr@gmail.com

state: Indiana

department: Environmental and Occupational Health

institute: Indiana University Bloomington

summary: We analyzed expression of miRNAs in Exhaled Breath Condensates from pediatric patients with sickle cell disease, asthma, sickle cell disease and asthma, and controls

overall_design:
We extracted RNA  from 0.5 mL of EBCs using the Plasma/Serum RNA Purification Kit. The expression of 1,066 human miRNAs was quantified using the Human miRNome miScript® miRNA PCR Array (QIAGEN, MIHS-3216Z). Results (Ct values) were normalized to global means, and fold changes were calculated using the 2-ΔΔCT method.
SCD: 1,2,3,4,5,6,7,8,9  SCD+ASTHMA: 10,11,12,13  CONTROL: 14,15,16,17,18,19,20,21,22  ASTHMA: 23,24,25,26

first: Pritish

last: Mondal

first: Roopa

last: Siddaiah


title: Mechanisms of drugs-resistance in small cell lung cancer II

city: Chiyoda-ku

first: Shogo

last: Okazaki

state: Tokyo

address: 1-8-13 Kanda-Surugadai

department: School of Dentistry

institute: Nihon University

summary: Malignant transformation of SCLC often occurs simultaneously with the acquisition of chemotherapy resistance, suggesting that phenotypic malignant transformation is associated with adaptation to chemotherapy-induced stress. Recently, it has been reported that autophagy deficiency is involved in chemotherapy resistance in SCLC. To analyze gene expression changes associated with SCLC malignant transformation, we established ATG7 knockout and ATG7/SQSTM1 double knockout SCLC cell lines and performed RNA sequencing.

overall_design: ATG7 Knockout and ATG7/SQSTM1 double knockout DMS273 cells were generated by CRISPRv2 vector. RNAs were isolated from each generated DMS273 cells and subjected RNA-sequencing analysis.

first: Kentaro

last: Suina

first: Juntaro

last: Yamasaki


title: c-Rel-dependent monocytes are potent immune suppressor cells in cancer

city: Philadelphia

first: Ting

last: Li

address: 422 Curie Blvd.

institute: University of Pennsylvania

summary: Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of leukocytes that are important for tumorigenesis and tumor immunotherapy. They comprise up to 10% of leukocytes in the blood of tumor patients and their depletion may be required for successful tumor immunotherapy. However, the identity of MDSCs remains obscure, primarily due to their heterogeneity and lack of a known lineage-specific transcription factor specifying their differentiation. Using single-cell transcriptomics and gene knockout approaches, we now describe a subset of murine and human myeloid suppressor cells, named rel-dependent monocytes (rMos), which are programmed by the transcription factor c-Rel of the NF-B family. Unlike MDSCs described previously, the c-Rel-dependent monocytes expressed a high amount of the proinflammatory cytokine IL-1β together with a low level of suppressive molecule arginase 1. Both in vitro and in tumor-bearing mice, these c-Rel+IL-1βhiArg1 monocytes promoted tumor growth by potently suppressing T cell function and showed a strong migratory phenotype, all of which were impaired by c-Rel deficiency or inhibition. Mechanistic studies revealed that c-Rel controlled the expression of monocyte signature genes through a unique transcriptional complex called the c-Rel enhanceosome, and IL-1CCL2 crosstalk between tumor cells and the rel-dependent monocytes maintained the suppressive tumor microenvironment. Thus, c-Rel specifies the development of a suppressive monocyte population and could be selectively targeted for treating cancer.

overall_design: To understand the effect of c-Rel in MDSC development in the TME, we injected wild-type (WT) (LysM-cre x Rel+/+) and myeloid conditional Rel knockout mice (LysM-cre x RelloxP/loxP) with B16F10 melanoma tumor cells. At 2 weeks post tumor cells injection, viable CD45+ cells from similarly-sized tumors were collected for single-cell RNA sequencing.

first: Mayassa

last: Bou-Dargham

first: Norman

last: Fultang

first: Xinyuan

first: Warren

middle: S

last: Pear

first: Honghong

last: Sun

first: Youhai

middle: H

last: Chen


title: Exercise promotes bone marrow microenvironment by inhibiting Adipsin in diet-induced obese mice

city: Shanghai

first: Jinwen

last: Luan

address: No. 200, Hengren Road

institute: Shanghai University of Sport

summary: To study how exercise induces bone remodeling in diet-induced obesity in a comprehensive way.8-week male C57/B6 mice were fed with HFD to establish an obese mice model. Then mice were randomly divided into 2 groups: the exercise and the control groups. The exercise group of mice was put on a moderate-intensity treadmill running for 12 weeks.We employed unbiased RNA-Seq to comprehensively investigate the exercise effect on the bone marrow of diet-induced obesity mice in vivo.

overall_design: To investigate how exercise induces bone remodeling in diet-induced obesity in a comprehensive way, we isolated the bone marrow from the the control and the exercise group tibia after 12-weeks exercise intervention and analyzed them by RNA-seq.

first: Zunhan

last: Shi

first: Lihui

last: Wang

first: Liqin

last: Yin

first: Xiaohui

last: Ji

first: Wenqian

last: Zhang

first: Bingxiang

last: Xu

first: Wei

last: Cheng

first: Ru

first: Longhua

last: Liu


title: Next Generation Sequencing Facilitates Quantitative Analysis of Reverb fl/fl and  Reverb fl/fl /MHC+ cardiac Transcriptomes

city: Hefei

first: Yanfeng

last: Xue

state: Anhui

address: Changjiang Western Road 130

institute: Anhui Agricultural University

summary: We use reverb heart specific ko mouse model to exclude the function of reverb in heart

overall_design: We harvest mice at 6 ZTs and snap frozen the hear samples for transcrptomic analysis.

first: Shiyang

last: Song

first: Zheng


title: Effect of pelleted or non-pelleted high-grain diet on ruminal epithelium in Hu-lambs

summary: Fifiteen male Hu-lambs were randomly assigned to three groups (n = 5 for each group). Lambs in the control (CON), HG, and HP groups received low-grain nonpelleted diet (30% concentrate), HG diet (70% concentrate), and HP diet containing the same ingredients and nutritions with HG group, respectivley. After 60-day treatment, all the lambs were slaughtered to collect ruminal epihelium samples for transcriptome analysis.

overall_design: Fifiteen ruminal epithelium samples were analyzed by transcriptome sequencing (5 from each group).

first: Shengyong

last: Mao

first: Yuyang

first: Ehab

last: Trabi


title: miRNA sequencing in synovium of human temporomandibular joint osteoarthritis

status: Public on Jun 10 2022

city: shanghai

first: Chuyao

address: The Ninth People's Hospital, No. 639, Zhizaoju Road, Huangpu District, Shanghai

institute: shanghai ninth people hospital

type: Non-coding RNA profiling by high throughput sequencing

summary: To investigate differentially expressed miRNAs in synovium of human temporomandibular joint osteoarthritis (TMJOA), we performed miRNA high-throughput sequencing in synovium of human TMJOA.

overall_design: Patients with reducible anterior disc displacement were  used as control groups and patients with TMJOA were used as experimental group.

first: Dongmei

last: He

first: Yihui

last: Hu

first: Huimin

last: Zhu

first: Luxiang

last: Zou


title: Transcriptomic analysis of mice heart following transverse aortic constriction (TAC) surgery

summary: We did TAC surgery for mice and daily treated the mice with Vehicle or SR9009

overall_design: We harvest mice at around ZT10 and snap frozen the heart samples for transcrptomic analysis.

last: Sun

first: Lilei


title: P. canariensis Resprouting

city: Göttingen

first: Victor

last: Chano

address: Buesgenweg 2

department: Forest Genetics and Forest Tree Breeding

institute: University of Göttingen

summary:
Expression profiling by microarray hybridization
Newly formed resprouts from Pinus canariensis at three different stages (R0, R1 and R2), and associated phloematic and meristematic tissue, was used for gene expression analysis against control tissues, sampled at the same dates from remote parts of the plant, in order to check transcriptomical variations of an unusual trait in conifers.

overall_design: Two-color experiment microarray using three biological replicates for sampling date

first: Alvaro

last: Soto

first: Carmen

last: Collada


title: An in vitro vascularized micro-tumor model of human colorectal cancer recapitulates in vivo responses to standard-of-care therapy

city: Irvine

first: Stephanie

middle: Jeanelle

last: Hachey

state: CA

address: 2400 Biological Sciences III

institute: UC Irvine

summary: Around 95% of anti-cancer drugs that show promise during preclinical study fail to gain FDA-approval for clinical use. This failure of the preclinical pipeline highlights the need for improved, physiologically-relevant in vitro models that can better serve as reliable drug-screening and disease modeling tools. The vascularized micro-tumor (VMT) is a novel three-dimensional model system (tumor-on-a-chip) that recapitulates the complex human tumor microenvironment, including perfused vasculature, within a transparent microfluidic device, allowing real-time study of drug responses and tumor–stromal interactions. Here we have validated this microphysiological system (MPS) platform for the study of colorectal cancer (CRC), the second leading cause of cancer-related deaths, by showing that gene expression, tumor heterogeneity, and treatment responses in the VMT more closely model CRC tumor clinicopathology than current standard drug screening modalities, including 2-dimensional monolayer culture and 3-dimensional spheroids.

overall_design: Here we used single-cell mRNA sequencing (scRNAseq) to profile the transcriptomes of cells isolated from HCT116-containing VMT and matched HCT116 CRC cells, EC and fibroblasts growing in 2D monocultures. We further sought to validate the robustness of our system by performing additional scRNAseq on cells isolated from SW480-derived VMT, matched monolayer cultures, and tumor spheroids.

first: Christopher

middle: W

last: Hughes


title: Effect of long non-coding RNA LL35/Falcor depletion on gene expression in murine hepatocytes in vivo and in vitro

city: Moscow

first: Evgeniya

last: Shcherbinina

address: Bolshoy Boulevard 30, bld. 1

country: Russia

institute: Skolkovo Institute of Science and Technology

summary: To determine and compare a functional role of long non-coding RNA LL35/Falcor in murine hepatocytes in vitro and in vivo we performed a transcriptome analysis of AML12 hepatocytes cell line and murine liver after LL35 depletion.

overall_design: Differential gene expression profiling analysis of RNA-seq data for AML12 cells with depleted LL35 (AML12 LL35 ASO) in comparison with control (AML12 Luc ASO), and for murine liver (Liver LL35 ASO) in comparison with control (Liver Luc ASO).

first: E

middle: Y

first: O

last: Sergeeva


title: RNA sequencing analysis of gene expression changes during the development of radiation proctitis in a mouse model

city: Guangzhou

first: Yanmei

last: Cui

address: Yuancun erheng Road

institute: Sun Yat-sen University

summary: Background: Radiation proctitis (RP) is the most common complication of radiotherapy for pelvic tumor. Currently there is a lack of effective clinical treatment and its underlying mechanism is poorly understood. In this study, we aimed to dynamically reveal the mechanism of RP progression from the perspective of RNomics using a mouse model, so as to help develop reasonable therapeutic strategies for RP.   Results: Mice were delivered a single dose of 25Gy rectal irradiation, and the rectal tissues were removed at 4 hours, 1 day, 3 days, 2 weeks and 8 weeks post-irradiation (PI) for both histopathological assessment and RNA-seq analysis. According to the histopathological characteristics, we divided the development process of our RP animal model into three stages: acute (4 hours, 1 day and 3 days PI), subacute (2 weeks PI) and chronic (8 weeks PI), which could recapitulate the features of different stages of human RP. Bioinformatics analysis of the RNA-seq data showed that in the acute injury period after radiation, the altered genes were mainly enriched in DNA damage response, p53 signaling pathway and metabolic changes; while in the subacute and chronic stages of tissue reconstruction, genes involved in the biological processes of vessel development, extracellular matrix organization, inflammatory and immune responses were dysregulated. We further identified the hub genes in the most significant biological process at each time point using protein-protein interaction (PPI) analysis and verified the differential expression of these genes by quantitative real-time-PCR (qRT-PCR) analysis.  Conclusions: Our study revealed the molecular events sequentially occurred during the course of RP development and might provide molecular basis for designing drugs targeting different stages of RP development.

overall_design: A total of 27 RNA samples  from the rectal tissues of mice in both irradiated and control group at 4 hours, 1 day, 3 days, 2 weeks and 8 weeks post-irradiation were collected and subjected to high-throughput RNA sequencing. Three biological replicates of the RNA sample were prepared for each time point in both groups. The same control samples were shared for 4 hours and 1 day.

first: Qingzhi

last: Zeng

first: Jingyang

last: Cheng

first: Haiyong

last: Wu


title: Single cell RNA Sequencing Facilitates understanding of cell-type specific function of miR-21 during Atherosclerosis

city: New Haven

first: Rolando

last: Garcia-Milian

state: CT

address: 333 Cedar St

department: CWML

institute: Yale University

summary:
Purpose: using single cell transcriptome profiling to find cell-type specific genes regulated by microRNA-21 following western diet feeding in atherosclerotic mouse models.
Methods: WT/mTmG/ Myh11ERT2 and miR-21fl/flmTmG/Myh11ERT2 littermates were injected Tamoxifen at 5weeks old for CRE recombination. Later received AAV8-PCSK9 injection then fed on western diet (1.25% cholesterol; D12108; Research Diets, Inc) for 20 weeks to induce atherosclerosis. Aortas were harvested from WT/mTmG/ Myh11ERT2 and miR-21fl/flmTmG/Myh11ERT2 mice and digested in 1.5mg/ml collagenase A (10103578001, Roche) + 0.5mg/ml elastase (6365, Worthington) buffer supplemented with 0.1% actinomycin at 37oC for 1h with gentle shaking. Digested aortas were passed through a 70 μm Cell Strainer and stained with a live/dead viability dye eFluor 450 (Thermo Fisher Scientific). Viable eGFP+ (SMCs) and eGFP-/tdTomato+ (non-SMCs aortic cells) were sorted by FACS Aria III (BD Biosciences). The sorted eGFP+ (SMCs) and eGFP-/tdTomato+ (non-SMCs aortic cells) cells were mixed to represent complete profile of aortic cells and then encapsulated into droplets and processed following manufacturer’s specifications using 10X Genomics GemCode Technology. Equal numbers of cells per sample were loaded on a 10x Genomics Chromium controller instrument to generate single-cell Gel Beads in emulsion (GEMs) at Yale Center for Genome Analysis. Lysis and barcoded reverse transcription of polyadenylated mRNA from single cells were performed inside each GEM followed by cDNA generation using the Single Cell 3’ Reagent Kits v2 (10X Genomics). Libraries were sequenced on an Illumina HiSeq 4000 as 2 × 100 paired-end reads.
Results: 5404 cells from WT/mTmG/ Myh11ERT2 sample and 6911 cells from miR-21fl/flmTmG/Myh11ERT2 sample for expression analysis and distinguished phenotypes were identified for SMCs lacking miR-21. Following pathway analysis on the DEGs revealed pathways regulated by miR-21 deficiency in SMCs that contributes to advancing atherosclerotic plaque.
Conclusions: Our study represents the first detailed transcriptomic analysis of atherosclerotic aortas from SMC specific knockout of miR-21. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that when lacking miR-21, a subset of SMCs are prone to apoptosis that supported the overall pro-atherogenic phenotype by miR-21 deficiency.

overall_design: Single cell transcriptomic profiling of 30-week old wild type (WT) and miR-21fl/flmTmG/Myh11ERT2 mice

first: Wen

last: Ding

first: Yajaira

last: Suarez


title: Ethylene-induced flowering in Aechmea fasciata

city: Danzhou

first: Jiabin

last: Wang

state: Hainan

address: Baodaoxincun

institute: Chinese Academy of Tropical Agricultural Sciences

summary: This study aims to understand ethylene-induced flowering in Aechmea fasciata. During ethylene-induced flowering, AfFTL2 expression is induced and targets the EIN3 binding site ‘ATGTAC’ by AfEIL1-like.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for AfEIL1-like.

first: Zhiying

last: Li

first: Li

last: Xu


title: H2A monoubiquitination targeting by cohesin subunits in Arabidopsis

first: Yu

last: Zhang

address: DongChuan Road No. 800

institute: Shanghai Jiaotong University

summary: Histone H2A monoubiquitination (H2Aub1) function as a conserved post-translational modification in eukaryotes to maintain gene repression and ensure cellular identity. Arabidopsis H2Aub1 is catalyzed by polycomb repressive complex 1 (PRC1) contributed by AtRING1s and AtBMI1s. Because PRC1 components lack known DNA binding domains, how H2Aub1 is established at the specific genomic sites remains a topic of debate. Here we show that the cohesin subunits AtSYN4 and AtSCC3 interact with each other and AtSCC3 binds to AtBMI1s. We find that H2Aub1 levels are significantly reduced in the atsyn4 mutant or AtSCC3 RNAi knockdown plants. ChIP-seq analysis indicated that AtSYN4 and AtSCC3 tend to associate with H2Aub1 in the genome regions of transcription activation independent on H3K27me3. Finally, we show that AtSYN4 binds directly to the G-box element and targets H2Aub1to these sites. Our study thus revealed a mechanism for cohesin-mediated recruitment of AtBMI1s to specific loci to mediate H2Aub1.

overall_design: Detecting the genome-wide occupancy of AtSYN4 and AtSCC3.

first: Yuda

last: Fang

first: Min

last: Ma

first: Meng

last: Liu


title: Single-cell RNA sequencing reveals tissue architecture during human high-grade serous ovarian cancer progression

city: Hangzhou

first: Junfen

state: Other (Non U.S.)

address: No.1 Xueshi Road

department: Department of Gynecologic Oncology

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

summary: As the most fatal type of gynecological cancers, high-grade serous ovarian cancer (HGSOC) exhibits heterogeneity that obstacles therapeutics. Up to date, the pathogenesis of HGSOC remains poorly understood. Here, we  performed the deep single-cell RNA sequencing (scRNA-seq) using 59,324 cells isolated from seven treatment-naïve HGSOC patients at early or late tumor stages and five age-matched non-malignant ovarian samples. Our sequencing data showed a highly complex ecosystem of tumor, immune and stromal cells based on their marker genes and functional properties. We showed that normal ovary-specific fibroblasts were carried with the properties of mesenchymal stem cells (MSCs), which could be reprogrammed to cancer-associated fibroblasts. During tumor progression, matrix CAFs (mCAFs) could induce the EMT properties of the tumor cells. Furthermore, the analysis of the early stage tumors illuminated enrichments of IDO-expressing macrophages, CD8+ TRM cells, and the activated CD8+ TEX cells (TRM-CXCL13), whereas those tumors at the late stages loss these features and expressed signatures related to epithelial-to-mesenchymal transition (EMT) program. Also, we characterized a specific chemokine-receptor communication in HGSOC tumor microenvironment, which may contribute to shape the HGSOC features. Our compendia of scRNA-seq results provide valuable insights for understanding the landscape of HGSOC ecosystem, which will be helpful in advancing HGSOC cancer diagnosis and therapy and enabling personalized approaches to the treatment.

overall_design: landscape of HGSOC ecosystem


title: Paternal imprinting of dosage-effect defective1 contributes to seed weight xenia in maize

city: Gainesville

first: Dawei

last: Dai

state: Florida

address: 2550 Hull Rd

department: Horticultural Sciences

institute: University of Florida

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

overall_design: Refer to individual Series


title: Analysis of Wild Type and miR-21-/- Smooth Muscle Cell Transcriptomes after atherosclerosis

summary:
Purpose: The goals of this study are to investigate the effect of miR-21 knockout in aortic smooth muscle cell transcriptome profiling (RNA-seq) to understand how miR-21 regulates SMC plasticity during atherosclerosis.
Methods: Aortic smooth muscle cell (SMC) mRNA profiles of 30-week-old wild-type (WT) and SMC specific miR-21 knockout (miR-21fl/flmTmG/Myh11ERT2) mice fed on western diet for 20 weeks were generated by deep sequencing, in triplicate. The sequence reads that passed quality filters were analyzed at the transcript isoform level . qRT–PCR validation was performed using TaqMan and SYBR Green assays to confirm miR-21 knockout.
Results: Using an optimized data analysis workflow, we mapped about 60 million sequence reads per sample to the mouse genome (build mm10) and identified 22,836 transcripts in the SMCs of WT and miR-21fl/flmTmG/Myh11ERT2 mice. RNA-seq data revealed ~300 genes differentially expressed in miR-21 knockout SMCs after atherosclerosis. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized pathways that may contribute to SMC function during atherosclerosis progression.
Conclusions: Our study represents the first detailed analysis of SMC lacking of miR-21 transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.

overall_design: Aortic SMC mRNA profiles of 20-week western diet feeding wild type (WT) and miR-21fl/flmTmG/Myh11ERT2 mice


title: RNA-Seq mapping enabled quantitative analysis of gene expression differences between normal and ded1-ref in Zea mays (Maize)

summary: The goals of this study are to compare NGS-derived transcriptomes (RNA-Seq) derived from ded1-ref mutant and normal maize endosperm tissue. Homozygous ded1-ref mutants exhibit a seed defect. Maize Ded1 encodes a transcription factor. This study characterizes DEGs between ded1-ref and normal endosperm and serves to identify direct target genes together with DAP-seq data.

overall_design: Endosperm mRNA profiles from 12 days after pollination (DAP) normal and ded1-ref kernels in the Zea mays W22 background were generated by deep RNA-Seq. Total RNA was prepared for 4 biological replicates of paired ded1-ref and normal sibling pools. 2x100bp PE sequencing performed on an Illumina HiSeq2000.

first: A

last: Settles

first: W

last: Barbazuk

first: Ruth

last: Davenport

first: Janaki

last: Mudunkothgi


title: Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and miR-21-/- Smooth Muscle Cell Transcriptomes

summary:
Purpose: Next-generation sequencing (NGS) has revolutionized systems-based analysis of cellular pathways. The goals of this study are to investigate the effect of miR-21 knockout in aortic smooth muscle cell transcriptome profiling (RNA-seq) to understand how miR-21 regulates SMC function at basal level.
Methods: Aortic smooth muscle cell (SMC) mRNA profiles of 12-week-old wild-type (WT) and SMC specific miR-21 knockout (miR-21fl/flmTmG/Myh11ERT2) mice were generated by deep sequencing, in triplicate. The sequence reads that passed quality filters were analyzed at the transcript isoform level . qRT–PCR validation was performed using TaqMan and SYBR Green assays to confirm miR-21 knockout.
Results: Using an optimized data analysis workflow, we mapped about 30 million sequence reads per sample to the mouse genome (build mm9) and identified 16,014 transcripts in the SMCs of WT and miR-21fl/flmTmG/Myh11ERT2 mice. RNA-seq data revealed >400 genes differentially expressed in miR-21 knockout SMCs. Hierarchical clustering of differentially expressed genes uncovered several as yet uncharacterized pathways that may contribute to SMC function.
Conclusions: Our study represents the first detailed analysis of SMC lacking of miR-21 transcriptomes, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. Our results show that NGS offers a comprehensive and more accurate quantitative and qualitative evaluation of mRNA content within a cell or tissue. We conclude that RNA-seq based transcriptome characterization would expedite genetic network analyses and permit the dissection of complex biologic functions.

overall_design: Aortic SMC mRNA profiles of 12-week old wild type (WT) and miR-21fl/flmTmG/Myh11ERT2 mice


title: DAP-seq was used to generate genome-wide DNA:TF interaction maps for ZmMYB73/DED1

first: Mary

last: Galli

first: Andrea

last: Gallavotti


title: Physiological and transcriptomic effects of hexafluoropropylene oxide dimer acid in Caenorhabditis elegans during development

status: Public on Jun 09 2022

city: San Diego

first: Goran

last: Bozinovic

address: 3030 Bunker Hill St

institute: Boz Life Science Research and Teaching Institute

summary: Per- and polyfluoroalkyl substances (PFAS) are chemicals that are relatively resistant to degradation. While such features are desirable in a variety of consumer and industry products, such as firefighting foams and non-stick cookware coating, some PFAS, including perfluorooctanoic acid (PFOA), are toxic and bioaccumulate. Hexafluoropropylene oxide dimer acid (HFPO-DA) is an emerging PFAS developed to replace PFOA and has not been extensively studied. To evaluate the potential toxicity of HFPO-DA with a cost- and time-efficient approach, we exposed C. elegans larvae to 4×10-9–4 g/L HFPO-DA in liquid media and performed assays measuring developmental, behavioral, locomotor, and transcriptional effects at various exposure levels. After 48 hours of 1.5–4 g/L HFPO-DA exposure, acute developmental toxicity was observed as developmental delay; statistically significantly delayed (p < 0.05) progeny production was observed in worms exposed to 2–4 g/L HFPO-DA. After 48 hours of 4×10-9–0.4 g/L exposure, no significant behavioral or locomotor effect was observed relative to the 0 g/L control group. Statistically significant differential gene expression was identified with over 99% confidence via the R-package NOISeq in all fourteen 48-hour 1.25×10¬-5–4 g/L HFPO-DA exposure groups, except for 6.25×10¬-5 g/L. Among 10298 genes analyzed, 2624 differentially expressed genes (DEGs) were identified in the developmentally delayed 4 g/L group only, and 78 genes were differentially expressed in at least one of the thirteen exposure groups testing 1.25×10¬-5–2 g/L HFPO-DA exposures. Genes encoding for detoxification proteins such as cytochrome P450 enzymes and UDP glucuronosyltransferases are upregulated in 0.25–4 g/L acute exposure groups. In the lower exposure concentration groups, statistically significant gene expression changes were also observed, though these DEGs did not share any biological functions, except for six ribosomal protein-coding genes. While our transcriptional data is insufficient for conclusive mechanistic explanation, the statistically significant gene expression differences detected at 1.25×10¬-5 g/L, the lowest concentration tested for transcriptional changes, is concerning and calls for further targeted analyses that focus on low-dose HFPO-DA exposure effects.

overall_design: Gene expression profiling analysis of shallow RNA-seq data for C. elegans 48 hour exposure to hexafluoropropylene oxide dimer acid (HFPO-DA); four replicates of 15 experimental conditions (14 different concentrations of HFPO-DA plus a control) were generated, with each condition containing around 1000 worms

first: Zuying

last: Feng

first: Flannery

last: McLamb

first: Jeanne

middle: P

last: Vu

first: Sylvia

last: Gong

first: Richard

last: Gersberg


title: Expression data of wild-type and mutant worms treated with Cry5Ba

city: Wuhan

first: feng

last: chen

address: No.1,Shizishan Street · Hongshan District

institute: Huazhong Agricultural University

summary:
Microarrays data of wild-type and mutant worm(RB1884) were used to determin which genes in the nematode were expressed depednet on mnp-1
To identify the genes that are key factors involved in MNP-1 mediated protection against Cry5Ba

overall_design: L4 stage RB1884 and N2 worms were fed with Cry5Ba/BL21 or BL21 at 20℃ for 4h, the worms washed by M9 buffer for 2-3 times were collected into 1.5 ml tubes, which were stored at -70°C after addition of 150 µl Trizol solution. Microarray production, hybridization, and scanning were performed with Affymetrix C. elegans fene 1.0 ST. The default background subtraction and normalization settings were used. Spots were filtered based on foreground to background ratio; values less than 1.2 were flagged. Log (base 2) values were exported.

first: Cuiyun

last: Pang

first: Wei

last: Zhou


title: CONSTANS Imparts DNA Sequence Specificity to the Histone Fold NF-YB/NF-YC Dimer

city: Milan

first: Matteo

last: Chiara

address: via celoria 26

department: Biosciences

institute: University of Milan

summary:
Nuclear Factor Y (NF-Y) is a heterotrimeric transcription factor that binds CCAAT elements. The NF-Y trimer is composed of a Histone Fold Domain (HFD) dimer (NF-YB/NF-YC) and NF-YA, which confers DNA sequence specificity. NF-YA shares a conserved domain with the CONSTANS, CONSTANS-LIKE, TOC1 (CCT) proteins. We show that CONSTANS (CO/B-BOX PROTEIN1 BBX1), a master flowering regulator, forms a trimer with Arabidopsis thaliana NF-YB2/NF-YC3 to efficiently bind the CORE element of the FLOWERING LOCUS T promoter.
Using saturation mutagenesis, electrophoretic mobility shift assays, and RNA-sequencing profiling of co, nf-yb, and nf-yc mutants, we identify CCACA elements as the core NF-CO binding site. CO physically interacts with the same HFD surface required for NF-YA association, as determined by mutations in NF-YB2 and NF-YC9, and tested in vitro and in vivo. The co-7 mutation in the CCT domain, corresponding to an NF-YA arginine directly involved in CCAAT recognition, abolishes NF-CO binding to DNA

overall_design: Comparative gene expression profiling analysis of co, nf-yb, and nf-yc  Arabidopsis thaliana mutant seedlings

first: Nerina

last: Gnesutta

first: Roderick

last: Kumimoto

first: Swadhin

last: Swain

first: Chamindika

last: Siriwardana

first: David

last: Horner

first: Ben

middle: F

last: Holt III

first: Roberto

last: Mantovani


title: Does a hypersensitive immune response to helminth infection at duodenum make Merino sheep susceptible to diarrhoea?

city: Perth

first: Shamshad

middle: Ul

last: Hassan

state: Western Australia

address: 35 Stirling Highway

country: Australia

institute: The University of Western Australia

summary:
Purpose:To explore the underlying biological mechanisms relating to development and control of diarrhoea, we compared sheep that were prone to develop diarrhoea with sheep that were not prone.
Methods:Transcriptomes in the tissues of Merino sheep where the parasites were located were analyzed using RNASeq. Functional enrichment of DEGs using GO terms in biological processes and KEGG pathways were studied. PPI (protein-protein interaction) network and sub-network were studied using clusterONE plugin in Cytoscape.
Result and conclusion: Our results indicate that suppression of ‘immune response’, ‘antigen processing and presentation’, and a list of biological functional terms related to ‘suppression in immune tolerance and activation of repair processes associated with tissue damage, including ‘extracellular matrix organization’, ‘collagen fibril organization’, ‘tissue morphogenesis’, ‘circulatory system development’, ‘morphogenesis of an epithelium’, and ‘focal adhesion’ were associated with high-diarrhoea phenotype. The genes with important roles in the responses to helminth infection could be targeted in breeding programs to prevent diarrhoea.

overall_design: Four biological replicates in each of High-diarrhoea (HD) and Low-diarrhoea (LD) group were used. LD group was used as a control group.

middle: U

first: Eng

last: Chua

first: Erwin

last: Paz

first: Parwinder

last: Kaur

first: Chin

last: Tay

first: Johan

middle: C

last: Greeff

first: Shimin

last: Liu

first: Graeme

last: Martin


title: Genes regulated by SMAD3 in HCC cells

first: Xuewu

last: Zhang

address: Tongji Hospital, Jiefang Avenue

institute: Huazhong University of Science and Technology

summary: To identify genes regulated by SMAD3, we have employed whole transcriptome microarray expression profiling to identify genes downstream of SMAD3. Total RNAs from SMAD3-overexpression (7721-SMAD3 vs 7721-VEC) or SMAD3-knockdown (LM3-shSMAD3 vs LM3-CON) cell lines were measured.

overall_design: Total RNAs were extracted from 7721-SMAD3, 7721-VEC, LM3-shSMAD3, LM3-CON stable cells, three repeated experiments were perfomred for each groups.

first: Xiaoping

last: Chen