title: Comparison of transcriptomes of PARP inhibitor-treated and -untreated isogenic human BRCA-wildtype ovarian cancer PDX

city: Denver

first: Benjamin

last: Bitler

state: CO

department: OB-GYN

institute: The University of Colorado

summary: Background: PARP inhibitors (PARPi) kill cancer cells by stalling DNA replication and preventing DNA repair, resulting in a critical accumulation of DNA damage. Resistance to PARPi is a growing clinical problem in the treatment of high grade serous ovarian carcinoma (HGSOC). Acetylation of histone H3 lysine 14 (H3K14ac) and associated histone acetyltransferases (HATs) have known functions in DNA repair and replication, but their expression and activities have not been examined in the context of PARPi-resistant HGSOC. Results: Using mass spectrometry profiling of histone modifications, we observed altered H3K14ac enrichment in PARPi-resistant HGSOC cells relative to isogenic PARPi-sensitive lines. By RT-qPCR and RNA-Seq, we also observed altered expression of numerous HATs in PARPi-resistant HGSOC cells and a PARPi-resistant PDX model. Knockdown of HATs only modestly altered PARPi response, although knockdown and inhibition of PCAF significantly increased resistance. Pharmacologic inhibition of HBO1 severely depleted H3K14ac but did not affect PARPi response. However, knockdown and inhibition of BRPF3, which is known to interact in a complex with HBO1, did reduce PARPi resistance. Conclusions: This study demonstrates that severe depletion of H3K14ac does not affect PARPi response in HGSOC. Our data suggest that bromodomain functions of HAT proteins such as PCAF, or accessory proteins such as BRPF3, may play a greater role in PARPi response than acetyltransferase functions.

overall_design: GTFB-PDX1009 is a BRCA-wildtype human HGSOC that develops significant ascites disease burden. Patient ascites cells were injected via intraperitoneal route into NSG mice and allowed to develop for 7 days. Mice were then treated once daily for 21 days by IP injection of vehicle control or 50 mg/kg olaparib. After the treatment regimen, disease burden was allowed to develop/recur for 2 months. At this point, mice were sacrificed and necropsied. Solid tumors and ascites were collected. The ascites cells were used for RNA-seq analysis. We hypothesized that olaparib treatment would select for a PARP inhibitor-resistant population of cells. We then used RNA-seq to compare the transcriptomes of the treated/selected cells vs. the untreated/unselected groups.

middle: G

first: Zachary

last: Watson

first: Hyunmin

last: Kim


title: Loss of heterozygosity of K-RAS in pancreatic cancer cell lines

status: Public on Dec 22 2021

city: New York

first: Richard

middle: A

last: Friedman

state: NY

address: 1130 St. Nicholas Av

department: Medicine

country: USA

institute: Columbia University Irving  Medical Center

type: Expression profiling by high throughput sequencing

summary: iWT-KRAS-MIA PaCa-2 cells were treated with doxcycline and compared to untreated control. The PaCa-2 line, which harbors a G12C mutation in KRAS, has lost its WT KRAS allele with amplification of the mutant allele.These cells express WT KRAS mRNA and protein upon induction with doxcycline in a dose-dependent manner.

overall_design: Gene expression of iWT-KRAS-MIA -PaCa-2, a modification of the  PaCa-2 a human pancreatic cell line which expresses extra WT-KRAS upon induction with doxycycline.

first: Han

last: Yan

first: Chih-Chieh

last: Yu

first: Stuart

last: Fine

first: Ayman

middle: L

last: Youssof

first: Ye-Ran

last: Yang

first: Jun

first: Dillon

middle: C

last: Karg

first: Edwin

last: Cheung

first: Haoqiang

last: Ying

first: Emily

middle: I

last: Chen

first: Ji

last: Luo

first: Yi

last: Miao

first: Wanglong

last: Qiu

first: Gloria

middle: H

last: Su


title: Single cell RNA sequencing of mouse brain hematopoietic cells after brain ischemia model

city: Tsukuba

first: Akira

last: Shibuya

state: Ibaraki

address: 1-1-1 Tennodai

department: Immunology

country: Japan

institute: University of Tsukuba

summary: We performed scRNA-seq of brain CD45+ hematopoietic cells from the brain of Cd300a-fl/fl and Cd300a-fl/fl Lys2-Cre mice 0, 1 and 3 hrs after the reperfusion of middle cerebral artery occlusion (MCAO).

overall_design: Brain PI-CD45+ hematopoietic cells were sorted by flow cytometry from Cd300a-fl/fl and Cd300a-fl/fl Lys2-Cre mice 0, 1 and 3 hrs after the reperfusion of MCAO. Three brains were pooled per each sample. Re-staining of PI of sorted samples were measured by flow cytometry, and the viability of all samples were more than 85%. Single cells were processed through the Chromium Single Cell Platform using the Chromium Single Cell 3’ Library and Gel Bead Kit v3 (10X Genomics, PN-1000092) and the Chromium Single Cell B Chip Kit (10X Genomics, PN-1000074) as per the manufacturer’s protocol. The sequencing of libraries was performed at Macrogen Japan (Kyoto, Japan) using an Illumina HiSeq X. Processed files were generated from Cell Ranger (10X Genomics) pipeline.

first: Kazumasa

last: Kanemaru

first: Chigusa

last: Oda

first: Yuta

last: Nakazawa

first: Fumie

last: Abe


title: Gene eRandomized Controlled Trial of the Gastrin Receptor Antagonist Netazepide in Patients with Barrett’s Esophagus

first: Richard

last: Friedman

summary:
A randomized, double-blind, placebo-controlled trial of netazepide (YF476) in patients with BE without dysplasia was performed.
Gene expression before and after treatment with netazepide and with a placebo was  measured with RNASeq

overall_design:
Gene expression of Barrett's esophogus of patients treated with netazapeptde 25 mg or placebo once daily for 12 weeks before and after treatment.
aa.pla.t1 placebo arm, before treatment with placebo, n=9.
ab.pla.t2 placebo arm,  after treatment with placebo, n=9.
ba.trt.t1 treatment arm, before treatment with netazapeptide, n=10.
bb.trt.t12treatment arm,  after treatment with treatment with netazapeptide, n=10.

first: Julian

last: Abrams

first: Armando

last: Del Portillo

first: Caitlin

last: Hills

first: Griselda

last: Compres

first: Bin

last: Cheng

first: John

middle: M

last: Poneros

first: Charles

middle: J

last: Lightdale

first: Massimiliano

last: di Pietro

first: Rebecca

last: Fitzgerald

first: Antonia

middle: R

last: Sepulveda

first: Malcolm

last: Boyce

first: Timothy

last: Wang


title: Amplification of Germ Cells through Parthenogenesis Faithfully Maintain Genomic Imprinting [RNA-seq]

city: Munich

first: Chenglei

last: Tian

address: Ingolstädter Landstraße 1

country: Germany

institute: Helmholtz Munich

summary: We perform E9.5 PGC RNA-seq for indentify the germ cell marker genes and imprinting genes expression.

overall_design: Two biological repetitions of E9.5 PGC by RNA-seq to indentify the germ cell marker genes and imprinting genes expression (compared with published E12.5 PGC and Inner cell mass RNA-seq data).


status: Public on Dec 21 2021

city: Jinan

first: Yujing

last: Sun

state: Shandong

address: 44 Wen Hua Xi Road

department: Department of pathology

country: China

institute: School of Basic Medical Sciences, Shandong University

summary: The long intergenic non-coding RNA linc01133 is reported to be oncogenic in various malignancies. However, the role and mechanism of linc01133 in regulating gastric cancer growth is still not clear. In the present study, we found that linc01133 was significantly up-regulated in gastric cancer tissues compared to non-tumorous gastric tissues. Linc01133 over-expression significantly correlated with tumor size and tumor differentiation in gastric cancer patients. The expression of linc01133 was regulated by c-Jun and c-Fos collaboratively. In both in vitro and in vivo studies, linc01133 was shown to promote gastric cancer cell growth. Linc01133 localized in the cytoplasm and functioned as an endogenous competing RNA of miR-145-5p to up-regulate the expression of YES1，which was proved to be the target gene of miR-145-5p. By promoting YES1-dependent YAP1 nuclear translocation, linc01133 up-regulated the expression of the key cell cycle regulators CDK4, CDK6 and cyclin D1 to promote G1-S phase transition. Thus, our study unveiled the function and mechanism of linc01133 regulating cell cycle progression in gastric cancer.

overall_design: RNA-sequencing of HGC27 cells with and without linc01133 overexpression, no replication

first: Peng

last: Gao


title: Sequencing of serum exosomal lncRNAs in lung adenocarcinoma

first: chuanxin

last: wang

address: 247 Beiyuan Street, Jinan 250033, Shandong, China,

institute: The Second Hospital of Shandong University

type: Non-coding RNA profiling by high throughput sequencing

summary: lncRNA sequencing for 10 mixed samples.

overall_design: Examining 2 conditions, each with 5 replicates.


title: Amplification of Germ Cells through Parthenogenesis Faithfully Maintain Genomic Imprinting [RRBS]

type: Methylation profiling by high throughput sequencing

summary: We asked whether oocyte number could be amplified through parthenogenesis of mouse oocytes, without requiring creation of a paternal genome and a genetically unique genome. Parthenotes develop to a blastocyst-like stage, and from this parthenogenetic ESCs (pESCs) can be derived with high efficiency. Like ESCs, pESCs maintain unlimited self-renewal and pluripotency, as well as germline competence. Further, we demonstrate that their expression pattern of imprinted maternal genes resembles that observed in oocytes. pESCs can be directed to differentiate into primordial germ cell-like cells (PGCLCs) and form oocytes that produce fertile pups and reconstitute ovarian endocrine functions. The transcriptome and imprinting pattern of PGCLCs differentiated from pESCs more closely approximate those of in vivo produced embryonic PGCs, than PGCLCs produced from ESCs. Parthenogenesis offers a promising route for deriving PGCLCs and amplifying oocytes by faithfully maintaining maternal genes, without fertilization.

overall_design: We performed RRBS assay to analysis the DNA methylation between female ESC and pESC, and also analysis the DNA methylation among E9.5PGC, E12.5PGC, female ESC-PGCLC and pESC-PGCLCs


title: Amplification of Germ Cells through Parthenogenesis Faithfully Maintain Genomic Imprinting

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

overall_design: Refer to individual Series


title: Retinal pigment epithelium-specific CLIC4 mutant is a mouse model of dry age-related macular degeneration

first: Ching-Hwa

last: Sung

state: New York

address: 1300 York Ave

institute: Weill Cornell Medicine

summary: Purpose: Bulk RNA-seq was perform to profile changes in 3-month-old mouse RPE cells caused by CLIC4 deficinecy

overall_design: RNAs  of RPE cells isolated from 3-month-old knokcout (KO), Control (Ctrl), and CreCtrl mice followed by deep sequencing using Illumina NovaSeq6000 sequencer with pair-end 50 cycles. We use two pairs (KO vs Ctrl), three pairs (KO vs. WT), and two paris (KO vs. Cre-Ctrl)

first: Jen-Zen

last: Chuang

last: Yang

first: Nobuyuki

last: Nakajima

first: Cheng

last: Fu


title: IBD HIBEC and HILEC transcriptome compendium

city: Milano

first: Luca

last: Massimino

state: MI

address: Via Olgettina, 58

department: Gastroenterology

country: Italy

institute: Ospedale San Raffaele

summary: The study aims to investigate the differences between Human intestinal blood and lymphatic endothelial cells, HIBECs and HILECs, respectively, isolated from colon and ilea of IBD and control patients.

overall_design: n=54 biopsy-derived endothelial cells, n=10 CD HIBEC, n=8 UC HIBEC, n=10 healthy HIBEC, n=10 CD HILEC, n=8 UC HILEC, n=8 healthy HILEC

first: Federica

last: Ungaro

first: Luigi

last: Lamparelli

first: Silvio

last: Danese


title: Combinatorial Immunotherapy Induces Tumor Infiltrating CD8+ T Cells with Distinct Functional, Migratory, and Stem-Like Properties III

city: San Diego

first: Wenjing

last: Yanf

state: CA

address: 10770 Science Center Drive

department: Oncology

institute: Pfizer

summary: This study reveals supeior efficiacy of triple combination treatment (TCT) based on anti-PD-(L)1 and anti-4-1BB/OX40 and describes immunological mechanisms underlying synergism between the treatment components

overall_design: Using bulk RNA seq of TILs isolated from isotype- and TCT-treated mice sorted based on expression of PD-1 and KLRG1 into 4 distinct populations, we established a gene expression profile of key CD8+ T cell populations induced by the TCT treatment. Using scRNA-Seq of TILs isolated from isotype- and TCT-tretaed mice we defined a sunctional, phenotypic and developmental T cell heterogeneity within treated and non treated tumors.

first: Natalija

last: Budimir


title: Combinatorial Immunotherapy Induces Tumor Infiltrating CD8+ T Cells with Distinct Functional, Migratory, and Stem-Like Properties II

overall_design: Using scRNA-Seq of TILs isolated from isotype-treated mice we defined a functional and phenotypic  T cell heterogeneity within non treated tumors.


title: Context-specific roles of diphthamide deficiency in hepatocellular carcinogenesis

city: Taipei

first: Wei-Ling

last: Tu

state: Beitou

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

country: Taiwan

institute: National Yang Ming Chiao Tung University

summary: In our current study, we observed that in hepatic carcinogen diethylnitrosamine (DEN)-induced liver tumor mouse model, diphthamide-deficient mouse developed less liver tumor incidence than control mouse. To explore the detailed mechanism, RNA-seq experiment was conducted to compare gene transcriptome profiles of control and diphthamide-deficient livers during HCC progression.

overall_design: Comparison of liver tissue transcriptomes in control (2 samples) and Dph1 KO livers (2 samples) at 1-day (1D) post-DEN treatment

first: Chun-Ming

last: Chen

first: Ya-Tung

last: Shih


title: CBX8 together with SET facilitates ovarian carcinoma growth and metastasis by suppressing the transcription of SUSD2

city: Tianjin

first: Chenghao

last: Xuan

address: 22 Qixiangtai Road

institute: Tianjin Medical University

summary: CBX8 promotes cell proliferation and migration through downregulating the expression of SUSD2.The mRNAs from SKOV3 cells expressing control shRNAs or CBX8 shRNAs were extracted and subjected to RNA-seq.

overall_design: Examination of gene expression changes in CBX8 knockdown and control SKOV3 cells.

first: Yang

last: Duan


title: Comprehensive Atlas of the Mouse Urinary Bladder

city: Farmington

first: Dylan

last: Baker

state: CT

address: 10 Discovery Dr

institute: The Jackson Laboratory

summary: We report the application of single-cell based sequencing for high throughput profiling of the mouse urinary bladder. By utilizing multiple dissociation techniques and library preparation techniques we generated an atlas comprising 43,119 cells including major cell types absent from previous reports. We found that previous single-cell profiling of the mouse bladder lacked a major cell type in the detrusor smooth muscle and incorrectly annotated other cell types such as mesothelial cells. Using the atlas, we elucidated aspects of bladder biology including urothelial differentiation, the identity of interstitial cells of Cajal, detrusor smooth muscle control and immune distributions. Finally, we combine the single-cell based sequencing with spatial transcriptomics and imaging mass cytometry to add spatial context to transcriptomic profiling.

overall_design: Single cell and nucleus sequencing of the mouse urinary bladder paired with spatial transcriptomics and bulk RNA sequencing.

first: Paul

last: Robson


title: Combinatorial Immunotherapy Induces Tumor Infiltrating CD8+ T Cells with Distinct Functional, Migratory, and Stem-Like Properties


title: Combinatorial Immunotherapy Induces Tumor Infiltrating CD8+ T Cells with Distinct Functional, Migratory, and Stem-Like Properties I

overall_design: Using scRNA-Seq of TILs isolated from isotype- and TCT-tretaed B16-inoculated mice we defined a functional, phenotypic and developmental T cell heterogeneity within treated and non treated tumors.


title: Impaired TFAM expression promotes mitochondrial damage to drive fibroblast activation and fibrosis in systemic sclerosis

city: Erlangen

first: Thuong

middle: Minh

last: Trinh

address: Ulmenweg 18

institute: University of Erlangen-Nuremberg

summary:
Objectives: The transcription factor TFAM is controlling the transcription of core proteins required for mitochondrial homeostasis. The aim of the current study was to investigate changes in TFAM expression in systemic sclerosis (SSc), to analyze mitochondrial function and to evaluate the consequences for fibroblast activation.

Methods: The expression of TFAM was analyzed by immunofluorescence and Western blot. The effects of TFAM knockout were investigated in cultured fibroblasts and in bleomycin-induced skin and lung fibrosis and in TβRIact-induced skin fibrosis.

Results: The expression of TFAM was downregulated in fibroblasts in SSc skin and in cultured SSc fibroblasts. The downregulation of TFAM was associated with decreased mitochondrial number and accumulation of damaged mitochondria with release of mtDNA, accumulation of deletions in mtDNA, metabolic alterations with impaired OXPHOS and release of the mitokine GDF15. Chronic, but not acute exposure of normal fibroblasts to TGFβ mimicked the finding in SSc fibroblasts with downregulation of TFAM and accumulation of mitochondrial damage. Downregulation of TFAM promotes fibroblast activation with upregulation of fibrosis-relevant GO-terms in RNASeq. Mice with fibroblast-specific knockout of TFAM are prone to fibrotic tissue remodeling with fibrotic responses even to NaCl instillation and enhanced sensitivity to bleomycin injection and TβRIact-overexpression. TFAM knockout fosters SMAD3 signaling to promote fibroblast activation.

Conclusions: Alterations in the key mitochondrial transcription factor TFAM in response to prolonged activation of TGFβ and associated mitochondrial damage induce transcriptional programs that promote fibroblast-to-myofibroblast transition and drive tissue fibrosis.

overall_design: Gene expression profile of TFAM siRNA-treated human dermal fibroblasts stimulated with TGFβ, and non-targeting siRNA-treated human dermal fibroblasts stimulated with TGFβ as controls

first: Xiang

last: Zhou

first: Cuong

last: Tran

last: Trinh-Minh

first: Jörg

last: Distler


title: Early-life thermal stress mediates long-term alterations in hypothalamic microglia

city: Rishon LeZion

first: Tatiana

last: Kisliouk

address: 68 HaMaccabim Road

department: Animal Sciences

country: Israel

institute: ARO


title: iChiffon triggers global histone H3 acetylation and expression of developmental genes in Drosophila embryos [chiffon]

city: West Lafayette

first: Vikki

middle: Marie

last: Weake

state: IN

address: 175 South University Street

department: Biochemistry

institute: Purdue University

summary: Purpose: The goal of this study was to identify the differential expressed genes in the Drosophila embryo when chiffon is disrupted.

overall_design: Method: RNA extracted from Drosophila embryos (stage 12-14) was used to generate cDNA libraries using NuGen Ovation RNA-seq System V2. Samples were sequenced using Ilumina HiSeq150 next generation sequencer (four biological replicates per genotype: chiffon null mutants flies, and the rescue of different transgenes: chiffon FL, chiffon C, chiffon WF24, and a WT control.

first: Eliana

last: Torres-Zelada


title: Chiffon triggers global histone H3 acetylation and expression of developmental genes in Drosophila embryos [ada2b]

summary: Purpose: The goal of this study was to identify the differential expressed genes in the Drosophila embryo when ada2b is disrupted.

overall_design: Method: RNA extracted from Drosophila embryos (stage 12-14) was used to generate cDNA libraries using NuGen Ovation RNA-seq System V2. Samples were sequenced using Ilumina HiSeq150 next generation sequencer (four biological replicates per genotype: ada2b null mutants flies, and the rescue of different transgenes: Ada2b_PA, Ada2b_PB, A34908116da2b_PA+Ada2b_PB, and a WT control.


title: Chiffon triggers global histone H3 acetylation and expression of developmental genes in Drosophila embryos [ChIP-seq]

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Purpose: The goal of this study was to identify H3K14ac levels in chiffon mutant vs WT

overall_design: Method: Drosophila embryos (stage 12-14) was used to ChIP H3K14ac and H3. 1ug of Drosophila chromatin was mixed to 50ng of yeast chromatin (spike-in) and ChIP was performed. Then we generated ChIP libraries using Ovation Ultralow library system (NuGEN) with unique dual indices for multiplexing. Samples were sequenced using Ilumina HiSeq150 next generation sequencer (three biological replicates per genotype: chiffon null mutants flies, and WT: chiffon mutant expressing the transgene chiffon FL.


title: Transcriptomics of MLN-expressing cells in human duodenum organoids

city: Cambridge

first: Fiona

last: Gribble

state: Cambridgeshire

address: Box 289 Addenbrooke's Hospital, Hills Road

department: Medical Research Laboratories

country: United Kingdom

institute: University of Cambridge

summary: We purified two populations of human duodenal MLN-expressing enteroendocrine cells (Venus+ and Venus-) from mature differentiated hMLN-Venus organoids by FACS and identified transcripts enriched in endocrine cell lineages.

overall_design: Comparative transcriptomic study of 2x cell populations by bulk RNAseq, with 4x biological replicates.

first: Emily

last: Miedzybrodzka

first: Christopher

last: Smith


title: Chiffon triggers global histone H3 acetylation and expression of developmental genes in Drosophila embryos


title: Nrf2 depletion sensitizes pancreatic cancer cells to gemcitabine via aldehyde dehydrogenase 3a1 repression.

city: Sendai

first: Shin

last: Hamada

state: Miyagi

address: 1-1 Seiryo-machi, Aobaku

department: Division of Gastroenterology

institute: Tohoku University Graduate School of Medicine

summary: The central regulator of oxidative stress responses, Nrf2, is attracting great interest as a therapeutic target in various cancers. Explorations of novel Nrf2 inhibitors have been reported, which suggest future clinical applications. Halofuginone, the derivative of natural plant alkaloid, has been reported to deplete Nrf2 rapidly in cancer cells with Nrf2 activation. In the current study, we identified that halofuginone sensitizes pancreatic cancer cells to gemcitabine, both in vitro and in vivo settings. We identified downregulation of Aldh3a1 in mouse pancreatic cancer cell lines after halofuginone treatment. The Nrf2 inducer diethyl maleate treatment upregulated Aldh3a1, and Nrf2 deleted pancreatic cancer cell lines failed to show induction. Knockdown of Aldh3a1 also sensitized cancer cells to gemcitabine, suggesting contribution of Aldh3a1 to gemcitabine resistance as a downstream target of Nrf2. Current study confirmed the therapeutic benefits of halofuginone in pancreatic cancer.

overall_design: Mouse cancer cell (K5, K6) were treated with control (DMSO) (K5C, K6C) or halofuginone at 100 nM for 24 hours (K5H, K6H). Gene expression profiles were compared between K5C and K5F, K6C and K6F, respectively.


title: MHC-independent αβ T cells replace epidermal and dermal γδ T cells in Tcrd-/- mice

city: Hannover

first: Christoph

last: Binz

state: Niedersachsen

address: Carl-Neuberg-Straße 1

department: Institut für Immunologie

institute: Medizinische Hochschule Hannover

summary: In Tcrd-/- mice, dendritic epidermal T cells (DETC) are known to be replaced by a DETC-population expressing αβ TCR. Here, the TCR-repertoire from Tcrd-/- mouse skin T cells was compared to the one from WT mice. The analysis revealed the existence of T cells expressing MHC-independent αβ TCR and replacing the missing γδ T cells in Tcrd-/- mouse epidermis and dermis.

overall_design: For TCR-sequencing the desired populations were sorted from WT or Tcrd-/- mice by flow cytometry as living CD45+CD3+TCRβ+ (WT dermal, Tcrd-/- dermal, Tcrd-/- pLN), living CD45+CD3hiTCRβhi (Tcrd-/- DETC), living CD45+CD3hiTCRγδhi (WT DETC) and living CD45+CD3+TCRβ+CD103+ or CD103hi (Tcrd-/- dermal CD103+, Tcrd-/- dermal CD103hi). Each sample is a pool from equivalent populations from several (5 to 18) animals. RNA was isolated from these cells and was analyzed by 5´RACE and PCR amplification followed by illumina-sequencing of Tcrb or Tcrg transcripts.

first: Anja

last: Bubke

first: Inga

last: Sandrock

first: Immo

last: Prinz


title: Transcriptomic analysis of total plantaris muscles RNA extract before and after one week of overload

city: Paris

first: Franck

last: Letourneur

address: 22 rue mechain

country: France

institute: INSERM

summary: Adult skeletal muscle is a plastic tissue that can adapt its size to workload. Here, we show that RhoA within myofibers is needed for overload-induced hypertrophy by controlling satellite cell fusion to the growing myofibers without affecting protein synthesis. At the molecular level, we demonstrate that, in response to increased workload, RhoA controls in a cell autonomous manner Erk1/2 activation and the expressions of extracellular matrix (ECM) regulators such as Mmp9/Mmp13/Adam8 and of macrophage chemo-attractants such as Ccl3/Cx3cl1. Their decreased expression in RhoA mutant is associated with ECM and fibrillar collagen disorganization and lower macrophage infiltration. Moreover, Mmps inhibition and macrophage depletion in controls phenocopied the lack of growth of RhoA mutants. These findings unravel the implication of RhoA within myofibers, in response to increase load, in the building of a permissive microenvironment for muscle growth and for satellite cell accretion through ECM remodeling and inflammatory cell recruitment.

overall_design: The aim of the analysis is to compare plantaris muscle controls and plantaris muscles mutants lacking RhoA (RhoA loss occurs in the myofibers) before and after compensatory hypertrophy induction. RhoA deletion occurs in vivo, in adult myofibers once that the mice were injected with tamoxifen. The mice were submitted to overload procedure (cut of gastrocnemius and soleus tendons in order to achieve plantaris muscle hypertrophy). One week after surgery mice were sacrificed and plantaris muscles were collected. Sham operated mice, control and mutant, were used as controls. RNA was isolated by plantaris muscles using TRIzol™ protocol. Microarray analysis will help to understand the molecular pathways involved in muscle growth defect observed in absence of RhoA upon hypertrophy. Isolated RNA from 4 mutant sham operated muscles, 4 mutant overloaded muscles, 3 controls sham operated muscles and 4 controls overloaded muscles were used to perform trasncriptomic arrays.


title: RNA-Seq data from A549 cells

city: Beijing

first: Qianchen

last: Li

address: Beijing

institute: PKU

summary: We report the transcription factor HOXB9 directly increases the expression of many target genes by performing RNA sequencing using HOXB9 stable knockdown A549 Lung adenocarcinoma cell line and HOXB9(WT/T133A) cell line.

overall_design: Examination of A549 cells stably infected with lentivirus carrying PLVX-HOXB9(WT/T133A) or HOXB9 KO

first: T

last: Wang

first: Q

first: H

last: Zhang

first: J

last: Zhan


title: Azacitidine as a potential therapeutic drug in pyridoxine-refractory female X-linked sideroblastic anemia

city: Kyoto

first: Yoshinori

last: Yoshida

address: 53, Kawahara-cho, Shogoin, Sakyo-ku

institute: Kyoto University

summary: X-linked sideroblastic anemia (XLSA) is associated with mutations in the erythroid-specific δ-aminolevulinic acid synthase (ALAS2) gene. Apart from pyridoxine-responsive patients, treatment for XLSA is mainly supportive. Female XLSA often represents a later onset of severe anemia, mostly due to the acquired skewing of X-chromosome inactivation. We successfully generated active wild-type and active mutant ALAS2 induced pluripotent stem cell (iPSC) lines from the peripheral blood cells of an affected mother and two daughters in a family with pyridoxine-resistant XLSA due to a heterozygous ALAS2 missense mutation (R227C). The erythroid differentiation potential was severely impaired in active mutant iPSC lines compared with active wild-type iPSC lines. Most of the active mutant erythroblasts revealed an immature morphological phenotype, and some showed dysplasia and perinuclear iron deposits. Additionally, globin and HO-1 expression and heme biosynthesis in active mutant erythroblasts were severely impaired compared with active wild-type erythroblasts. Furthermore, genes associated with erythroblast maturation and karyopyknosis showed a significantly lower expression in active mutant erythroblasts, recapitulating the maturation defect. Notably, the erythroid differentiation ability and hemoglobin expression of active mutant iPSC-derived hematopoietic progenitor cells (HPCs) were improved by the administration of δ-aminolevulinic acid (ALA), confirming the suitability of the cells for drug testing. The administration of a DNA demethylating agent, azacitidine, reactivated the silent wild-type ALAS2 allele in active mutant iPSC-derived HPCs and ameliorated the erythroid differentiation defects, suggesting that azacitidine is a potential novel therapeutic drug for female XLSA. Our patient-specific iPSC platform provides novel biological and therapeutic insights for XLSA.

overall_design: RNA was extracted from XLSA patient-derived iPSCs and CD34+ cells on day 8, CD43+CD34+CD38-Lin- cells on day 15, and CD235a+ erythroblasts on day 34 from active wild-type iPSC lines and active mutant iPSC lines sorted by FACS. RNA was also extracted from MACS-sorted CD235a+ primary erythroblasts in patient bone marrow mononuclear cells and healthy donor bone marrow mononuclear cells.

first: Yuki

last: Morimoto

first: Kazuhisa

last: Chonabayashi

first: Chikako

last: Okubo

first: Megumi

last: Narita


title: Disruption of the histone H3 cupric reductase activity prevents global transcriptional rewiring when YFH1 is diminished

city: Los Angeles

first: Siavash

middle: K

last: Kurdistani

address: 615 Charles E Young Dr South

department: Biological Chemistry

institute: UCLA

summary: Disruptions to iron-sulfur (Fe-S) clusters, essential cofactors for a broad range of proteins, cause widespread cellular defects resulting in human disease. An underappreciated source of damage to Fe-S clusters are cuprous (Cu1+) ions. Since histone H3 enzymatically produces Cu1+ to support copper-dependent functions, we asked whether this activity could become detrimental to Fe-S clusters. Here, we report that histone H3-mediated Cu1+ toxicity is a major determinant of cellular Fe-S cluster quotient in the budding yeast. Inadequate Fe-S cluster supply, due to diminished assembly as occurs in Friedreich’s Ataxia, causes substantial growth defects and numerous transcriptional responses. Decreasing Cu1+ abundance, through attenuation of histone cupric reductase activity via the H3H113N mutation, prevented the widespread transcriptional rewiring. Our findings reveal a novel interplay between chromatin and mitochondria in Fe-S cluster homeostasis.

overall_design: mRNA profiles of yeast with the histone H3 H113N mutation and/or YFH1 shutoff grown in fermentative medium

first: Oscar

last: Campos


title: Next Generation Sequencing Facilities Quantitative analysis of hiPSC derived neural stem cells, early neural progenitors, neural progenitors and neural progenitors after exposure to Idebenone and Bezafibrate Transcriptomes

city: Warsaw

first: Pawel

last: Gaj

address: Zwirki i Wigury 101

country: Poland

institute: Warsaw Genomics

summary: The mechanisms of neural differentiation of human induced pluripotent stem cells (hiPSCs) have been widely investigated, however the genes that play key regulatory role, especially involved in mitochondrial function are still unidentified. In this report transcriptome analysis of hiPSC differentiating into neural stem cells (NSC), early neural progenitors (eNP) and neural progenitors (NP) were performed. Gene Ontology and Network Analysis was implemented considering especially mitochondrial genes at all developmental stages and upon stimulation by idebenon (IDB) and bezafibrate (BZ) at NP stage.   The genes strongly up-regulated during the process of differentiation in all tested stages belong mainly to: WNT; Cadherin and Gonadotropin-releasing hormone receptor signaling pathways. The interaction network shows that the transcription factors including HOXD3, ASCL1, HES5, and ZEB1 are possible key players in this process. Mitochondrial unfolded protein response (UPRmt), ROS defense, and pyruvate metabolism were the strongest up-regulated mitochondrial processes in all analyzed stages of neural differentiation. IDB and BZ treatment at NP stage induced different set of nuclear encoded mitochondrial genes involved in respiratory electron transport chain and fatty acid metabolic process respectively. Thus PARP agonist-mediated stimulation of mitochondrial genes influence the process of neural differentiation despite of mitochondrial biogenesis, but through the specific receptor pathways.

overall_design: Examination of 3 stages of hiPSC neural differentation (neural stem cells - NSC, early neural progenitor - eNP, neural progenitor - NP) and response of hiPSC derived- neural progeniotors (NP) to Idebenone (IDB) or Bezafibrate (BZ). Three independant experiments of hiPSC neural differentation and NP treatment was performed. Analyzed 1 replicate from 3 independent neural differentation experiments and 1 replicate from 3 experiments related with drug exposure.

first: Lonora

last: Buzanska

first: Piotr

middle: P

last: Stepien

first: Krystian

last: Jazdzewski

first: Justyna

last: Augustyniak

first: Monika

last: Kolanowska


title: microRNA-155 controls iNKT cell development and lineage differentiation by coordinating multiple regulating pathways

city: Detroit

first: Qing-Sheng

last: Mi

state: Michigan

address: 1 Ford Place

department: Department of Dermatology

institute: Henry Ford Health System

summary: The development of invariant natural killer T (iNKT) cells requires a well-attuned set of transcription factors, but how these factors are regulated and coordinated remains poorly understood. MicroRNA-155 (miR-155) is a key regulator of numerous cellular processes that affects cell development and homeostasis. Here, we found that the miR-155 was highly expressed in early iNKT cells upon thymic section, and then its expression is gradually downregulated during iNKT cell development. However, the mice with miR-155 germline deletion had normal iNKT cell development. To address if downregulated miR-155 is required for iNKT cell development, we made a CD4Cre.miR-155 knock-in (KI) mouse model with miR-155 conditional overexpression in the T cell lineage.  Upregulated miR-155 led to interruption of iNKT cell development, diminished iNKT17 and iNKT1 cells, augmented iNKT2 cells, and these defects were cell-intrinsic. Furthermore, defective iNKT cells in miR-155KI mice resulted in the secondary innate-like CD8 T cell development. Mechanistically, miR-155 modulated multiple targets and signaling pathways to fine tune iNKT cell development. MiR-155 modulated Jarid2, a critical component of a histone modification complex, and Tab2, the upstream activation kinase complex component of NF-kB, which function additively in iNKT development and in promoting balanced iNKT1/iNKT2 differentiation.  In addition, miR-155 also targeted Rictor, a signature component of mTORC2 that controls iNKT17 differentiation. Taken together, our results indicate that miR-155 serves as a key epigenetic regulator, coordinating multiple signaling pathways and transcriptional programs to precisely regulate iNKT cell development and functional lineage, as well as secondary innate CD8 T cell development.

overall_design: perform an unbiased RNA-Seq analysis of sorted NK1.1- iNKT cells and  NK1.1+ iNKT cells from miR-155KI and WT littermate mice

first: Jie

first: Kai

last: Li

first: Xilin

first: Guihua

first: Tingting

last: Liu

first: Xiaojun

last: Wu

first: Stephen

last: Brown

first: Li

last: Zhou


title: RNA Binding Protein SRSF3 confers an essential role in megakaryocyte maturation and platelet production

city: Clayton

first: Minna-Liisa

last: Anko

state: VIC

address: 15 Innovation Walk

department: Department of Anatomy and Developmental Biology

country: Australia

institute: Monash University

summary: RNA sequencing was performed to identify gene expression changes following SRSF3 depletion in mouse bone marrow megakaryocytes and peripheral blood platelets.

overall_design: Low (8N) or high (≥16N) ploidy bone marrow megakaryocytes and peripheral blood platelets were isolated from adult Pf4Cre-Srsf3fl/fl and control mice and RNA-sequencing was performed.


title: AMPK activation enhances apoptotic priming and BCL2 dependence via PERK phosphorylation

first: Franck

last: Letourneur

summary: gene expression profiling comparing the effect of a direct AMPK activator (GSK621) in human AML cells (MOLM-14 cell line) transduced with a non-targeted CRISPR/Cas9 RNA guide (SCR) or a EIF2AK3 (PERK) - targeting RNA guide

overall_design: PERK WT and PERK KO MOLM-14 cells were cultured 6h with 30mM GSK621 before RNA extraction

first: Jerome

last: Tamburini


title: Melatonin promotes Arabidopsis root growth in an IAA dependent manner [72h]

city: Wuhan

first: Zhulong

last: Chan

state: Hubei Province

address: No. 1, Shizishan St, Nanhu

department: College of Horticulture and Forestry Sciences

institute: Huazhong Agricultural University

summary:
Purpose: We aimed to compare transcriptomic changes after melatonin (MT) and IAA treatments in Arabidopsis and dissected cross-talk between MT and IAA
Methods: A total amount of 1 μg RNA was used for generation of sequencing libraries using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample. After cluster generation, the library preparations were sequenced on an Illumina Hiseq2000 platform and paired-end reads were generated. Clean reads were obtained by removing low quality reads, reads containing adapter and ploy-N from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated. Index of the Arabidopsis genome was built using Bowtie v2.2.3 and paired-end clean reads were aligned to the reference genome using TopHat v2.0.12. HTSeq v0.6.1 was used to count the reads numbers mapped to each gene. And then FPKM (Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced) of each gene was calculated based on the length of the gene and reads count mapped to this gene. Differential expression analysis of drought stress versus control condition was performed using the DESeq R package (1.18.0).
Results:In total, six samples with two biological replicates per genotype/treatment combination were used for RNA sequencing analysis. At least 2 G clean bases were generated for each sample. Comparative analysis identified coregulated genes by melatonin and IAA in Arabidopsis seedlings

overall_design: In this study, transcriptome changes after 1nM IAA and 10nM MT were compared at 72h after treatment. The rosette leaves from control and treated plants were collected for RNA isolation. Three treatments in duplicate were used in this study.

last: Yang


title: Transcriptomic analysis revealed adverse effect of melatonin on root growth

summary:
Purpose: We aimed to compare transcriptomic changes after high concentration melatonin treatment in Arabidopsis
Methods: A total amount of 3 μg RNA was used for generation of sequencing libraries using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample. After cluster generation, the library preparations were sequenced on an Illumina Hiseq platform and 125 bp/150 bp paired-end reads were generated. Clean reads were obtained by removing low quality reads, reads containing adapter and ploy-N from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated. Index of the Arabidopsis genome was built using Bowtie v2.2.3 and paired-end clean reads were aligned to the reference genome using TopHat v2.0.12. HTSeq v0.6.1 was used to count the reads numbers mapped to each gene. And then FPKM (Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced) of each gene was calculated based on the length of the gene and reads count mapped to this gene. Differential expression analysis of drought stress versus control condition was performed using the DESeq R package (1.18.0).
Results:In total, eight samples with two biological replicates per genotype/treatment combination were used for RNA sequencing analysis. At least 2 G clean bases were generated for each sample. Comparative analysis revealed different networks which were modulated by melatonin and IAA in Arabidopsis seedlings

overall_design: In this study, wild type Arabidopsis plants were grown in MS plate for 7 days.Plants were then treated with 0,100 and 1000μM melatonin for 12h. The rosette leaves were then collected for RNA isolation and deep sequencing.

first: Jun

last: You


title: Double-strand GFP RNA injection reveals no off-target effect in Nasonia vitripennis

city: Wageningen

first: Julien

last: Rougeot

state: Gerlderland

address: Droevendaalsesteeg 1  Building 107

department: Plant Science Group

country: Netherlands

institute: Wageningen University and Research

summary: The aim of this experiment is to assess the off-target effect of RNAi in Nasonia vitripennis at transcriptome level by comparing the effect of a non-specific RNAi against GFP together with water or not injected controls.

overall_design: RNAseq of dsGFP RNA, water-injected, or non-injected controls each in triplicates.

first: Yidong

last: Wang

first: Eveline

last: Verhulst


title: Microgravity and space radiation exert either opposite or synergic effects on single molecular pathways

city: Padova

first: Mattia

last: Forcato

address: Via Ugo Bassi, 58/B

institute: University of Padova

summary:
Microgravity and space radiation (SR) are two highly influential factors affecting humans in space flight. Many health problems reported by astronauts derive from endothelial dysfunction and impaired homeostasis. Here we describe the adaptive response of human, capillary endothelial cells to space. Reference samples on ground and at 1g onboard allowed discrimination between the contribution of microgravity and SR within the combined response to space.
Cell softening and reduced motility occurred in space, with loss of actin stress fibers and a greater distribution of microtubules and intermediate filaments in compensation. The frequency of primary cilia also increased. DNA repair mechanisms were indeed activated. Transcriptomics highlighted the opposing effect of microgravity from SR on specific molecular pathways: radiation, unlike microgravity, stimulated pathways for endothelial activation (hypoxia, inflammation), DNA repair and apoptosis, promoting an ageing-like phenotype; microgravity, unlike SR, activated pathways for metabolism and pro-proliferation phenotype. Thus, microgravity and SR should be considered separately to tailor effective countermeasures to protect astronauts’ health.

overall_design: To identify transcriptional changes induced by microgravity, space radiation, and their combination together with other stimuli in space flight, we used a model of human endothelial cells of microvascular origin (HMEC-1). Space samples were exposed to environmental microgravity (g) or set inside the centrifuge reproducing 1g; on ground samples were used as ground control.

first: Debora

last: Angeloni

first: Silvio

last: Bicciato

first: Ivana

last: Barravecchia


title: Transcriptomic response of bermudagrass to heat stress treatments for 2 and 12h

summary:
Purpose: We aimed to dissect heat response of bermudagrass and identify heat stress responsive genes.
Methods: A total amount of 3 µg RNA was used for generation of sequencing libraries using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample. After cluster generation, the library preparations were sequenced on an Illumina Hiseq platform and 125 bp/150 bp paired-end reads were generated. Clean reads were obtained by removing low quality reads, reads containing adapter and ploy-N from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated. Index of the Arabidopsis genome was built using Bowtie v2.2.3 and paired-end clean reads were aligned to the reference genome using TopHat v2.0.12. HTSeq v0.6.1 was used to count the reads numbers mapped to each gene. And then FPKM (Fragments Per Kilobase of transcript sequence per Millions base pairs sequenced) of each gene was calculated based on the length of the gene and reads count mapped to this gene. Differential expression analysis of drought stress versus control condition was performed using the DESeq R package (1.18.0).
Results:In total, six samples with two biological replicates per genotype/treatment combination were used for RNA sequencing analysis. At least 2 G clean bases were generated for each sample. Comparative analysis identified genes modulated by short term heat (2h) and long term heat (12h) treatments.

overall_design: In this study, 10-day-old bermudagrass seedlings were treated with 42 °C temperature for 2 and 12h. The leaves of control and heat treated bermudagrass plants were then collected for RAN isolation.


title: Maintenance of meiotic crossover against reduced DSB formation in fission yeast lacking histone H2A.Z

city: New York

first: Shintaro

last: Yamada

address: 1275 York Avenue

department: MOLECULAR BIOLOGY PROGRAM

institute: MEMORIAL SLOAN-KETTERING CANCER CENTER

type: Genome binding/occupancy profiling by genome tiling array

summary: Meiotic crossover (CO) recombination is a process in which programmed DNA double-strand breaks (DSBs) around hotspots are repaired by homologous chromosomes (homologs), and thus exchanges DNA molecules between two homologs. CO is critical for most sexually-reproducing organisms because it promotes accurate chromosome segregation and creates genetic diversity. Therefore, faithful accomplishment of CO formation must be ensured, but the bases of the regulation are not fully understood. Our previous study using fission yeast has revealed that mutants lacking the conserved histone H2A.Z are defective in DSB formation but maintain CO frequency at three loci tested. Here, we tested five additional sites to show that H2A.Z-lacking cells exhibit normal and increased CO frequency at two and three loci, respectively. Detailed interrogation on one of the intervals revealed that CO increase in the mutant would be genetically separable from DSB reduction, and regulated at least partly at the recombination intermediate level. In addition, through genetic analyses of model regions containing a hotspot flanked by two markers as well as genome-wide analyses, we propose that, in the absence of H2A.Z, CO arose from non-hotspot DSBs may compensate for DSB reduction at hotspots. These observations provide clues to further our understanding on CO control.

overall_design: S. pombe pat1-114 rad50S rec12+-FLAG cells in wild type, or pht1 deletion background were induced to enter meiosis by the pat1 haploid meiosis system, and harvested five hours after the induction. ChIP were performed using anti-FLAG antibodies. Input DNA was purified from pat1-114 cells in wild type background and harvested four hours after the induction.

first: Takatomi

first: Da

middle: Q

last: Ding

first: Yurika

last: Fujita

first: Emi

last: Takaya

first: Yasushi

last: Hiraoka

first: Hiroshi

last: Murakami

first: Kunihiro

last: Ohta


title: Molecular characterization of t(2;14)(q22;q32) and  t(6;14)(q25;q32) translocations in mixed myeloid/T lymphoid leukemia patients.

city: Leuven

first: Zeynep

last: Kalender Atak

address: Herestraat 49

country: Belgium

institute: KULeuven

summary: We characterized two recurrent translocations, i.e. t(2;14)(q22.3;q32) and t(6;14)(q25;q32), in FLT3 positive mixed myeloid/T-cell lymphoid leukemias (MLPA). Both translocations involved BCL11B at 14q32. In cases with t(2;14), the 2q22.3 breakpoint disrupted ZEB2 and produced a ZEB2-BCL11B fusion, a significant ZEB2-BCL11B over-expression with concomitant BCL11B wild type (wt) down-regulation. In cases with t(6;14), the 6q25 breakpoints fell in a 100Kb region where no genes have been mapped thus it not appeared to produce a fusion transcript. As long and short BCL11B isoforms were overexpressed in cases with t(6;14), this could be linked to loss of negative regulatory elements or juxtaposition to strong enhancers/promoters to the intact gene. As BCL11B and ZEB2-BCL11B have identical motifs for DNA and protein interactions, we hypothresized that common downstream target genes might be deregulated. Indeed, gene expression profiling analysis identified a specific signature that characterized this subgroup of MLPA and was unlike any known AML and T-ALL. Exome sequencing did no revealed common variations, thus it was used to investigate 252 candidate genes known to be involved in solid cancer or leukemias. Recurrent mutations in 13 genes, 5 of which were new somatic mutations of DNMT3A, EP300, GNAS, FAT4, and PRRX1 were identified.

overall_design: Four  patients with t(2;14)(q22;q32) and one with t(6;14)(q25;q32) translocations were compared with 14 AML patients (five NPM+/FLT3+, five NPM+/FLT3- and  four PML-RARA/FLT3+).

first: Paolo

last: Gorello

first: Kim

last: De Keersmaecker

first: Roberta

last: La Starza

first: Lucia

last: Brandimarte

first: Caterina

last: Matteucci

first: Idoya

last: Lahortiga

first: Danika

last: Di Giacomo

first: Valentina

last: Pierini

first: Enrico

last: Gottardi

first: Filomena

last: Daraio

first: Massimo

last: Negrini

first: Barbara

last: Crescenzi

first: Barba

last: Gianluca

first: Luigina

last: Luciano

first: Achille

last: Ambrosetti

first: Nicoletta

last: Testoni

first: Donatella

last: Raspadori

first: Iwona

last: Wlodarska

first: Peter

last: Vandenberghe

first: Stein

last: Aerts

first: Jan

last: Cools

first: Cristina

last: Mecucci


title: Tauroursodeoxycholic acid prevents acute graft-versus-host disease in mice by regulating intestinal apoptosis and inflammation

city: Freiburg

first: Geoffroy

last: Andrieux

address: Breisacherstr 153

institute: University clinics Freiburg

summary:
The effect of tauroursodeoxycholic acid on the development of acute graft-versus-host disease was tested in a murine allogeneic stem cell transplantation model.
TUDCA was shown to reduce acute GVHD by reducing intestinal antigen presentation and following apoptosis and increasing the viability of intestinal epithelial cells.

overall_design: Small intestine of mice receiving TUDCA or vehicle treatment after transplantattion was isolated on day 14 and analyzed by Affymterix arrays.

first: P

last: Apostolova

first: G

first: M

last: Boerries


title: Cicatricial Alopecia pathogenesis: Uniqueness or similarities between lymphocytic subtypes Lichen planopilaris (LPP) and Central centrifugal cicatricial alopecia (CCCA)

city: Cleveland

first: Pratima

last: Karnik

state: Ohio

address: 10900 Euclid Ave., BRB547A

department: Dermatology

institute: Case Western Reserve University

summary: Microarray analyses of LPP and CCCA was undertaken to decipher differential pathophysiological pathways leading to disease initiation and progression

overall_design: Scarring alopecia consists of a collection of disorders characterized by destruction of hair follicles, replacement with fibrous scar tissue, and irreversible hair loss.  Alopecia affects men and women worldwide and can be a significant source of psychological stress and depression for affected individuals.  The purpose of this study was to explore Gene expression profiles in scalp tissue samples from normal control subjects (n=1(comprising 10 pooled samples)) and in matched samples obtained from affected (n=3) and unaffected (n=3) areas of the scalp in patients with Central centrifugal cicatricial alopecia (CCCA)  as well with matched samples obtained from affected (n=7) and unaffected (n=7) areas of the scalp in patients with Lichen planopilaris (LPP) .

first: Paradi

last: Mirmirani

first: Vera

last: Price

first: Patrick

last: Leahy

first: Roopa

middle: S

last: Subbaiah


title: RNA-seq analysis in mouse cortical extracts with deletion of STI1 TPR1 domain

city: Jerusalem

first: Hermona

last: Soreq

state: Israel

address: The Edmond J. Safra Campus, Givat Ram

institute: The Hebrew University of Jerusalem

summary: Long RNA-sequencing was performed on 5 cortex samples from STI1 wt/wt and 5 STI1 dTPR/dTPR mice. RIN values for these samples ranged between 8.2 and 8.7. Illumina sequencing yielded an average of 797,963 reads per sample, with 93.69% reads mapping rate to the mouse genome. We first verified the complete absence of STI1 exons 2 and 3 in the dTPR/dTPR samples compared to the wt/wt control tissues where these axons were both expressed. Principal component analysis and sample distance matrix analyses did not segregate between the two groups, indicating minimal differences between them. Furthermore, thorough RNA sequencing analysis did not reveal any significant changes in transcripts passing FDR correction (padj<0.05). Briefly, there were 85 gene transcripts (representing 18 unique genes) that passed p.value<0.05 and whose levels were changed by more than 1.5 fold increase (or 0.75 decrease)

overall_design: Examination of transcriptome changes in control and transgenic mice lacking STI1-TPR1 domain

first: Marco

last: Prado

first: Gilli

last: Moshitzky


title: The human dental apical papilla promotes spinal cord repair through a paracrine mechanism

status: Public on Dec 20 2021

city: Bruxelles

first: Anne

last: des Rieux

address: Drève de la Ferme, 92

institute: UCLouvain

summary: Traumatic spinal cord injury is an overwhelming condition that strongly and suddenly impacts the patient’s life and her/his entourage. There are currently no predictable treatments to repair the spinal cord, while many strategies are proposed and evaluated by researchers throughout the world. One of the most promising avenues is the transplantation of stem cells, although its therapeutic efficiency is limited by several factors, among which cell survival at the lesion site. In our previous study, we showed that the implantation of a human dental apical papilla, residence of stem cells of the apical papilla (SCAP), supported functional recovery in a rat model of spinal cord hemisection.

overall_design: In this study, we employed protein multiplex, immunohistochemistry, cytokine arrays, RT-qPCR and RNAseq technology to decipher the mechanism by which the dental papilla promotes repair of the injured spinal cord. We found that the apical papilla reduced inflammation at the lesion site, had a neuroprotective effect on motoneurons and increased the apoptosis of activated macrophages/ microglia. This therapeutic effect is likely driven by the secretome of the implanted papilla since its known to secrete an entourage of immunomodulatory or pro-angiogenic factors. Therefore, we hypothesize that the secreted molecules were mainly produced by SCAP, and that by anchoring and protecting them, the human papilla provides a protective niche ensuring that SCAP could exert their therapeutic actions. Therapeutic abilities of the papilla were demonstrated in the scope of spinal cord injury but could very well be beneficial to other types of tissue.


title: Cholesterol deprivation drives DHEA biosynthesis in human adrenals

city: Bern

first: Emanuele

last: Pignatti

address: Freiburgstrasse 15

country: Switzerland

institute: University of bern

summary: We report the transcriptome profiling obtained by serum starving NCI-H295R cells

overall_design: Differential expression between NCI-H295R cells in normal growth media, and cells deprived of serum


title: Mitochondrial dysfunction induces oxidative stress and an impaired ubiquitin proteasome system to trigger the pathogenesis of Frontal Fibrosing Alopecia (FFA)

summary:
Mitochondrial dysfunction is implicated in myriad diseases. However, there is no single platform that facilitates mapping of mitochondrial pathways and processes essential for health and those that are defective in disease. Here, we manually curated 1008 mitochondrial proteins to create a database of 31 mitochondrial pathways and processes and named the database as ‘MitoGyan’. ‘Gyan’ in Sanskrit means knowledge. We also developed a mitochondria associated metabolic pathways visualization tool called MitoCyc within HumanCyc. Using these novel resources and an integrated ‘Omics’ approach, we demonstrate that mitochondrial dysfunction occurs early in a rare, inflammatory hair disease called Frontal Fibrosing Alopecia (FFA). Transmission electron microscopy and mass spectrometry show morphological and functional changes in the mitochondria of FFA hair follicles. Our data suggests that mitochondrial dysfunction induces oxidative stress and an impairment of the ubiquitin proteasome system (UPS) in FFA. We propose that mitochondria-targeted antioxidants provide a new therapeutic strategy for FFA.
Microarray analyses of FFA was undertaken to decipher pathophysiological pathways of FFA leading to disease intitiation and progression

overall_design: Scarring alopecia consists of a collection of disorders characterized by destruction of hair follicles, replacement with fibrous scar tissue, and irreversible hair loss.  Alopecia affects men and women worldwide and can be a significant source of psychological stress and depression for affected individuals.  The purpose of this study was to explore Gene expression profiles in scalp tissue samples from normal control subjects (n=10(pooled)) and in matched samples obtained from affected (n=3) and unaffected (n=2) areas of the scalp in patients with lymphocytic Frontal Fibrosing Alopecia (FFA).

first: Sausan

last: Azzam

first: Mary

last: Consolo

first: Sujatha

last: Mohan

first: Selam

last: Shah

first: Calvin

last: Chi

first: Sreejith

last: Panicker

first: Daniela

last: Schlatzer

first: Hisashi

last: Fujioka

first: Bernard

last: Tandler


title: Next Generation Sequencing Facilitates Quantitative Analysis of the transcriptomes of RAW264.7 cell in 3D and 2D culture

city: JINAN

first: zenjun

last: li

address: 440 Jiyan Road, Huaiyin District

institute: Shandong Cancer Hospital and Institute, Shandong First Medical University and Shandong Academy of Medical Science

summary:
Purpose: the goal of this study is to detect and compare the transcriptome expression of mouse macrophages cultured in 3D and 2D environments, and find the effect of 3D culture on macrophages compared with traditional 2D culture.
Methods: Total RNA was extracted from purified and untreated RAW264.7 cells from 3D and 2D culture systems using TRIzol. The RNA samples were analyzed using Whole Genome Oligo Microarrays. After RNA had been hybridized to the microarray, it was washed and scanned, and data were extracted using Agilent Feature Extraction Software. Gene expression data were generated using Affymetrix GeneChip Human Genome U133 Plus 2.0 on an Affymetrix 3000 instrument running Gene‑Chip operating software.
Result: RNA-sequencing (RNA-seq) revealed that after 24 h of culture under the same conditions, 3D-cultured macrophages showed significant differences in gene expression. RNA-seq detected 18580 genes in the 2D group and 15777 genes in the 3D group, among which 6762 were differentially expressed (|log2(fold change)| >= 1, padj < 0.05) in both 3D and 2D groups. A total of 5949 genes were downregulated and 813 were upregulated.
Conclusion: Our study represents the first detailed analysis of the effect of 3D culture on mouse macrophages, and the results showed that compared with traditional 2D culture, the gene expression of macrophages under 3D culture was significantly changed.These findings are therefore worthy of further investigation and verification, and provide novel avenues for future research in cytology and macrophages.

overall_design: the mRNA profiles of mouse macrophages RAW264.7 in 3D and 2D culture

first: Haoxin

last: Shi

first: Dong

last: Li


title: Comparative transcriptome characterization of paired primary gastric cancer and ovarian metastasis

city: Shanghai

first: Shenglin

last: Huang

address: 270 Dongan

institute: Fudan University

summary: To understand the molecular mechanism underlying ovarian metastasis of gastric cancer, we performed RNA-seq of paired primary tumor, normal mucosa and ovarian metastasis of four GC patients by the Illumina sequencing platform with 150-bp paired-end, followed by functional enrichment analyses of differentially expressed genes between three sample sets. A total number of 15,493 protein-coding genes were detected, the majority of which were the annotated human reference genes. Using a threshold of fold change >2 and adjusted P value <0.05, a total number of 3023 differentially expressed genes were detected between different sets of samples. Among them, 479 and 609 protein-coding genes were up- and down-regulated in ovarian metastases over primary tumors, respectively. Functional enrichment analyses revealed the significant enrichment of immune system, tumor microenvironment, metabolism and sex hormone-related pathways in the ovarian metastasis of gastric cancer. In conclusion, comparative transcriptome characterization of paired primary and ovarian metastatic tumor profiled the genome-wide molecular expression and unveiled functionally enriched pathways underlying this specific type of distant metastasis in GC.

overall_design: RNA profiling of triple-matched primary gastric cancer, normal mucosae and ovarian metastasis by deep sequencing using Illumina NovaSeq .

first: Jianpeng

last: Gao

first: Dazhi

last: Xu


title: PRDM16 Functions as A Compact Myocardium-Enriched Transcription Factor Required to Maintain Compact Myocardial Cardiomyocyte Identity in Left Ventricle (Spatial Transcriptomics)

city: La Jolla

first: Ju

last: Chen

state: California

address: 9500 Gilman Drive 0613

institute: University of California San Diego

summary: The loss of compact myocardial cardiomyocyte (compact CM) identity suggests that PRDM16 deficiency may dramatically alter the cellular composition in left ventricular (LV) compact myocardium. To test this hypothesis, we performed single-cell RNA-seq (scRNA-seq) to examine gene dysreulation in Prdm16cKO heart. We also performed Visium Spatial Transcriptomics (ST) to facilitate mapping CM clusters to their original locations in the heart. As a result, we were able to demonstrate gene misregulation in Prdm16cKO mouse mainly occurred in LV compact CM.

overall_design: Two pairs of Prdm16cKO/WT E13.5  mouse hearts and two pairs of Prdm16cKO/control E15.5  mouse hearts were used for Visium Spatial Transcriptomics (ST).

first: Tongbin

last: Wu

first: Zhengyu

last: Liang


title: Transcriptome_LPS_macrophage_ProNa

city: Beijing

first: Yujun

address: Yumingyuan West Road

institute: China Agricultrual University

summary: ProNa supplementation for LPS-challenged macrophages


title: PRDM16 Functions as A Compact Myocardium-Enriched Transcription Factor Required to Maintain Compact Myocardial Cardiomyocyte Identity in Left Ventricle (scRNA-seq)

overall_design: Two E13.5 WT heart samples and two E13.5 Prdm16cKO heart samples (each sample included 3-4 individual hearts) were used for scRNA-seq.


title: PRDM16 Functions as A Compact Myocardium-Enriched Transcription Factor Required to Maintain Compact Myocardial Cardiomyocyte Identity in Left Ventricle (bulk RNA-seq)

summary: Gene expression profiling in whole heart, left ventricle (LV) and right ventricle (RV) of WT and Prdm16 conditional knockout (Prdm16cKO) mouse (Prdm16flox/flox; Xmlc2-Cre) at embryonic day 13.5 (E13.5).

overall_design: Three pairs of whole hearts, four pairs of  isolated LV and four pairs of isolated RV from Prdm16cKO and littermate control mice were subject to RNA-seq analysis to identify differentially expressed genes (DEGs) in whole heart, LV and RV of Prdm16cKO mice.


title: Metabolic and transcriptional changes across osteogenic differentiation of mesenchymal stromal cells

city: Reykjavik

first: Ólafur

last: Sigurjónsson

address: Menntavegur 1

department: Department of Engineering

country: Iceland

institute: University of Reykjavik

summary: Mesenchymal stromal cells (MSCs) are multipotent post-natal stem cells with applications in tissue engineering and regenerative medicine. MSCs can differentiate into osteoblasts, chondrocytes, or adipocytes, with functional differences in cells during osteogenesis accompanied by metabolic changes. The temporal dynamics of these metabolic shifts have not yet been fully characterized and are suspected to be of importance for therapeutic applications, such as osteogenesis optimization. Here, our goal was to characterize the metabolic shifts that occur during osteogenesis. We longitudinally profiled five key extracellular metabolites (glucose, lactate, glutamine, glutamate, and ammonia) from MSCs from four donors to classify osteogenic differentiation into three metabolic stages, defined by changes in the uptake and secretion rates of the metabolites in cell culture media. We used a combination of untargeted metabolomic analysis, targeted analysis of 13C-glucose labelled intracellular data, and RNA-sequencing data to reconstruct the gene regulatory network and further characterize cellular metabolism. The metabolic stages identified in this proof-of-concept study provide a framework for more detailed investigations aimed at identifying biomarkers of osteogenic differentiation and small molecule interventions to optimize MSC differentiation for clinical applications.

overall_design: Human bone marrow-derived MSCs from four donors were purchased from Lonza (Basel, Switzerland). Cells were cultured in osteogenic differentiation media. Cell cultures were sampled at specified time points for expression analysis

first: Olafur

middle: E

last: Sigurjonsson

first: Ottar

last: Rolfsson

first: James

middle: T

last: Yurkovich

first: Snaevar

last: Sigurðsson

first: Aristotelis

last: Kotronoulas

first: Adrian

last: Lopez Garcia de Lomana

first: Sarah

last: McGarrity

first: Thora

middle: B

last: Sigmarsdottir


title: PRDM16 Functions as A Compact Myocardium-Enriched Transcription Factor Required to Maintain Compact Myocardial Cardiomyocyte Identity in Left Ventricle (ChIP-seq)

summary: Identification of genome-wide PRDM16 binding sites in E13.5 whole mouse heart, as well as in isolated left ventricle (LV) and right ventricle (RV).

overall_design: Whole heart (two WT replicates), isolated LV (two WT replicates) and RV (two WT replicates) tissue samples were used in PRDM16 ChIP-seq experiments. Prdm16cKO samples and input samples were used to evaluate the specificity of  PRDM16 antibody in ChIP-seq. Each whole heart sample combined three individual hearts of the same genotype, each isolated LV or RV samples combined 10-12 individual WT LVs or RVs.


title: Transcriptomic profiles of splenic Gr-1+ cells from IDO-KO hosts and WT hosts after bone marrow transplantation

city: Seoul

first: Woo-Jeong

last: Shon

address: 103 Daehak-ro, Jongno-gu,

department: College of Medicine

country: South Korea

institute: Seoul National University

summary: Tryptophan-catabolizing enzyme indoleamine 2,3-dioxygenase 1 (IDO1) has also immunological function to suppress T cell activation in inflammatory circumstances, including  graft-versus-host disease (GVHD), a fatal complication after allogeneic bone marrow transplantation (allo-BMT). Although the mononuclear cell expression of IDO1 has been associated with improved outcomes in GVHD, the underlying mechanisms remain unclear. Herein, we used IDO-deficient (Ido1-/-) BMT to understand why myeloid IDO limits the severity of GVHD. Hosts with Ido1-/- BM exhibited increased lethality, with enhanced proinflammatory and reduced regulatory T cell responses compared with wild type (WT) alloBMT controls. Despite the comparable expression of the myeloid-derived suppressor cell (MDSC) mediators, arginase-1, inducible nitric oxide synthase, and IL-10, Ido1-/- Gr1+CD11b+ cells from allo-BMT or in vitro BM culture showed compromised immunesuppressive functions and were skewed toward the Ly6ClowLy6Ghi subset, compared with the WT counterparts. Importantly, Ido1-/-Gr-1+CD11b+ cells exhibited elevated levels of reactive oxygen species (ROS) and neutrophil numbers. These characteristics were rescued by human IDO1 with intact heme-binding and catalytic activities, and were recapitulated by the treatment of WT cells with the IDO1 inhibitor L1-methyl tryptophan. ROS scavenging by Nacetylcysteine reverted the Ido1-/-Gr-1+CD11b+ composition and function to an MDSC state, as well as improved the survival of GVHD hosts with Ido1-/- BM. In summary, myeloid-derived IDO1 enhances GVHD survival by regulating ROS levels and limiting the ability of Gr1+CD11b+ MDSCs to differentiate into proinflammatory neutrophils. Our findings provide a novel mechanistic insight into the immune-regulatory roles of the metabolic enzyme IDO1.

overall_design: To investigate the molecular pathways regulated by myeloid IDO1, we compared the transcriptomic profiles of splenic Gr-1+ cells from IDO-KO hosts and WT hosts on day 7 posttransplantation. Gr-1+ cells from syngeneic non-GVHD hosts were included as controls.

first: Ji

last: Ju

first: Minho

last: Jung

first: Young-Kwan

last: Lee

middle: Y

last: Lim

first: Joo

last: Kim

first: Jun

last: Chang

first: Chang

last: Min

first: Dong-Sup

first: Kyungho

last: Choi

first: Dong-Mi

last: Shin

first: Eun


title: PRDM16 Is a Compact Myocardium-Enriched Transcription Factor Required to Maintain Compact Myocardial Cardiomyocyte Identity in Left Ventricle

first: Ju

last: Chen

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

overall_design: Refer to individual Series


title: TNF-receptor 2 signaling enhances suppressive abilities in human circulating T follicular regulatory cells

city: Fukuoka

first: Yuichiro

last: Semba

address: 3-1-1 Maidashi

institute: Kyushu University

summary:
Objective: T follicular regulatory (Tfr) cells express CXC chemokine receptor type 5. They migrate into germinal centers and suppress T follicular helper (Tfh) and B cells for regulating antibody production. Tumor necrosis factor (TNF) inhibitors are used in inflammatory diseases; however, autoantibody and anti-drug antibody production is challenging. As TNF receptor 2 (TNFR2) signaling is important for suppressive functions of regulatory T cells, we investigated the role of TNFR2 on Tfr cells.
Methods: Tfr, Tfh, and naive B cells were obtained from human peripheral blood. Tfr cells were stimulated with MR2-1 (anti-TNFR2 agonistic antibody) and their proliferation, Forkhead box P3 (Foxp3) expression, and surface molecules were evaluated by flow cytometry. Tfh/B-cell proliferation and B-cell IgM production and differentiation in co-cultures with MR2-1-stimulated Tfr cells were examined.
Results: TNFR2 expression was higher on Tfr cells than conventional T cells. MR2-1 altered the gene expression profile of Tfr cells. Cell proliferation and Foxp3 expression were induced by MR2-1. Inducible costimulator and program death-1 were dramatically upregulated in MR2-1-stimulated Tfr cells and significantly suppressed Tfh/B-cell proliferation, IgM production, and B-cell differentiation. Tfr cells treated with MR2-1 migrated according to the CXCL13 gradient.
Conclusion: TNFR2 signaling in Tfr cells regulates Tfh and B-cell population. Aberrant antibody production during TNF inhibitor treatment might be associated with TNFR2 signaling inhibition in Tfr cells, suggesting TNFR1-specific inhibition as a better therapeutic strategy than pan-TNF inhibition. TNFR2-treated Tfr cells may serve as prospective candidates for cell-based therapy of autoimmune diseases.

overall_design: Transcriptome analysis of Tfr cells and Treg cells with or without TNFR2 stimulation.

first: Shotaro

last: Kawano

first: Hiroki

last: Mitoma

first: Jumpei

last: Nogami

first: Hiroaki

last: Niiro


title: Genome-wide STAT5 occupancy profiling in LPS induced CD127 expressing human monocytes

first: Bin

last: Zhang

address: Zheng Yutong Building room D311

institute: Tsinghua University

summary: STAT5 functions in human inflammatory monocytes is largely unknown. We identified the anti-inflammatory function of CD127-STAT5 axis in human monocytes. To investigate the mechanism underlying STAT5-mediated anti-inflammatory regulation, thoroughly dissecting the STAT5 genomic distribution in human monocytes would be informative. Therefore, we performed anti-STAT5 ChIP-seq analyses for CD127high monocytes under LPS stimulation to globally characterize the STAT5 occupancy in CD127 expressing human monocytes.

overall_design: PBMC were isolated from buffy coat of healthy blood donors by Ficoll gradients, and CD14+ monocytes were isolated from PBMC with anti-CD14 magnetic beads (Miltenyi Biotec). CD14+ monocytes were treated with LPS for 6 h. Stimulated cells were firstly stained for surface CD127, and then were collected for FACS-based sorting of CD127high monocytes. Sorted cells were used to perform anti-STAT5 ChIP-seq experiment to globally characterize the STAT5 occupancy in CD127 expressing human monocytes.


title: Transcriptome profiling of Mantle Cell Lymphoma and healthy controls using RNA-seq

city: Aarhus N

first: Morten

last: Veno

state: DK

address: Aabogade 15

country: Denmark

institute: Omiics ApS

summary: The aim of this study was to profile transcriptome data, in particular circRNA expression, in lymph node tumour specimens from patients with Mantle Cell Lymphoma at diagnosis

overall_design: We performed total RNA sequencing using total RNA from lymph node tumour specimens of 14 patients with MCL, total RNA from two reactive lymph node specimens from healthy controls, and total RNA isolated from Naive B cells of peripheral blood of four healthy controls.

first: Lasse

last: Sommer Kristensen

first: Kirsten

last: Gønbæk

first: Mette

last: Dahl


title: 35 samples

city: Boston

first: Irene

last: Ghobrial

state: MA

address: 4 Blackfan Circle, 2nd Floor, Suite 240

department: Medical Oncology

country: USA

institute: Dana-Farber Cancer Institute

summary:
35 samples

Keywords: Expression profiling by high throughput sequencing

overall_design:
35 samples

>>>Submitter states that raw data will be submitted to dbGaP due to patient privacy concerns<<<

first: Gad

last: Getz


title: Transcriptome analysis of islets from diabetes-resistant and diabetes-prone obese mice reveals novel gene regulatory networks involved in beta cell compensation and failure

city: Sydney

first: Ruby

middle: CY

last: Lin

state: NSW

address: 176 Hawkesbury Road, Westmead

institute: The Westmead Institute for Medical Research

summary: We carried out genome-wide transcriptome analysis of islets to investigate the gene expression landscape of ob/ob and db/db mouse models at 6 and 16 weeks of age. The purpose of this study is to determine the changes in the islet transcriptomic landscape that mediate the processes of beta cell compensation and failure in ob/ob and db/db mice.

overall_design:
Islets of C57BL/KsJ db/db and C57BL/6J ob/ob mice, and their age-matched lean control mice were studied at 6 and 16 weeks of age.
Mice were anesthetized and their islets isolated and handpicked under a stereomicroscope for extraction of RNA.

first: Jeng

last: Chan

first: Mohammed

last: Bensellam

first: D

last: Laybutt


title: Transcriptomic and lipidomic analysis reveal the mechanism of phosphorus-induced lipid class remodeling in Nannochloropsis sp. PJ12

city: Zhoushan

first: Jianhua

last: Liu

address: 1 Zheda Road, Lin-cheng street

institute: Ocean College, Zhejiang University

summary: Phytoplankton lipids, such as microalgae lipids, are important compounds of increasing  interest in bioenergy, food, pharmacy, aquaculture and ecology for their high molecular diversity. There is a taxonomically diverse lipid response under P stress with unresolved questions related to the diversified mechanism behind the lipid responses. A marine microalgae with high EPA content was isolated, named Nannochloropsis sp. PJ12. We reveal a mechanism of phosphorus-induced lipid class remodeling in Nannochloropsis sp. PJ12 based on highly corresponding transcriptome and lipidome data. Phosphorus- deprivation leads to the rapid reduction of phospholipids (PL) and synthesis of the betaine lipids (BL). Phosphorus-complement recovers the content of PL and BL to the original level. The changes are mediated mainly by a glycerophosphoryldiester phosphodiesterases on the transcriptome level. To adapt to low phospholipids, the transcription levels of gene encoding P transporter were upregulated. When Nannochloropsis sp. PJ12 was once again under phosphorus-complement, some of gene encoding P transporter continue to increase on the transcription levels. The novel phospholipid-remodeling scheme opens new avenues for metabolic engineering of lipid composition in algae.

overall_design: Examination of transcriptional profiling of PJ12 cells in response to phosphorus replete, phosphorus deprivation and phosphorus replenishment

first: Jibei

last: Liang

first: Fang

last: Wen


title: The Transcriptome Landscape change of Human Granulosa Cells under gonadotropins Revealed by Small bulk Cell RNA Sequencing

first: Zhiqiang

last: Yan

address: No.5 Yiheyuan Road Haidian District

institute: Peking University

summary:
STUDY QUESTION: Can superovulation dose of gonadotropins (Gns) affect the transcriptome of granulosa cells and follicular fluid (FF) hormone levels?

SUMMARY ANSWER: Cell cycle- and meiosis-associated genes were down-regulated by Gns stimulation, whereas immune- and cytokine-associated genes were up-regulated. Hormone levels were also affected by Gns stimulation.

WHAT IS KNOWN ALREADY: Superovulation treatment had adverse effects on maturation, spindle function, and chromosomal aneuploidy in oocytes.

STUDY DESIGN, SIZE, DURATION: One leading pre-ovulatory follicle per subject was used from three natural-cycle and four Gns-stimulated patients.

PARTICIPANTS/MATERIALS, SETTING, METHODS: Granulosa cells and FF were collected from the same leading follicle from each patient. RNA was extracted from granulosa cells and subjected to deep sequencing and analysis. Follicle-stimulating hormone (FSH), estradiol (E2), androstenedione (AND), testosterone (T), luteinizing hormone (LH), and progesterone (P) levels in FF were measured by immunoassays. Student  s t test was used for statistical analysis.

MAIN RESULTS AND ROLE OF CHANCE: A total of 715 genes were up-regulated, and 287 genes were down-regulated, in the Gns-stimulated group relative to the Control group. Gene Ontology analysis revealed that the down-regulated genes were enriched in cell cycle and meiosis pathways, primarily those associated with follicle or oocyte maturation and quality. On the other hand, the up-regulated genes were enriched in functions related to immunity and cytokine Ccytokine receptor interactions. In stimulated vs. natural-cycle follicles, E2 and LH levels were significantly reduced (P < 0.001).

first: Cuiling

last: Lu

first: Huailiang

last: Feng

first: Jie

last: Qiao


title: Nanopore long-read transcriptomics profiling reveals gene expression signatures of mouse tumor endothelial cells 2H-11

status: Public on Dec 19 2021

city: Changchun City, Jilin Province

first: Yuanyuan

last: Tian

address: The Xinmin Street No. 26

institute: The Key Laboratory of Pathobiology, Ministry of Education, The College of Basic Medical Sciences, Jilin University

summary:
Purpose: Nanopore sequencing is a new generation of Nanopore-based single-molecule real-time electrical signal sequencing technology for full-length transcriptome sequencing. We use Oxford Nanopore Technology (ONT) three-generation full-length transcriptome sequencing to detect all genetic structural changes in the transcriptome of tumor endothelial cells (TECs) 2H-11 and normal endothelial cells (NECs) SVEC4-10.
Methods: The mRNA profiles of three samples of mouse tumor endothelial cells (TECs) 2H-11 and three samples of mouse normal endothelial cells (NECs) were generated by Oxford Nanopore Technologies long read processing, using PromethION. Differential expression analysis of six samples was performed using the DESeq2 R package (1.6.3). Genes with a FDR < 0.01 and foldchange ≥ 2 found by DESeq2 were assigned as differentially expressed. Gene Ontology (GO) enrichment analysis of the differentially expressed genes (DEGs) was implemented by the GOseq R packages. KOBAS software was used to test the statistical enrichment of (DEGs) in KEGG pathways.
Results: In mouse tumor endothelial cells 2H-11,1847 genes are up-regulated and 1202 genes are down-regulated. According to the Gene ontology (GO) enrichment analysis of differentially expressed genes (DEGs), we found that different functional trends related to metabolic processes, developmental processes, localization, immune system processes, and locomotion are the main reasons for the differences. DEGs are mainly enriched in signal pathways related to cancer, immunity and metabolism, involving Pathways in cancer, Antigen processing and presentation, Proteoglycans in cancer, Focal adhesion, MAPK signaling pathway, Protein digestion and absorption, ECM-receptor interaction, PI3K-Akt signaling pathway and Glutathione metabolism. We also obtained the structural variation of transcripts such as alternative splicing, gene fusion, and alternative polyadenylation and accurately quantified the expression of the transcript.
Conclusions: Differentially expressed genes (DEGs) in key pathways may be potential diagnostic markers or therapeutic targets of TECs. Our study could help better understand the molecular mechanisms of tumor endothelial cells (TECs) involved in tumorigenesis and development. And our data also provide useful genetic resources for improving the genome and transcriptome annotations of TECs and normal endothelial cells (NECs).

overall_design: The mRNA profiles of three samples of mouse tumor endothelial cells (TECs) 2H-11 and three samples of mouse normal endothelial cells (NECs).

first: Wei

last: Li


title: miR-34a, miR-122, miR-206 and miR-210 targets identification

city: Valbonne - Sophia-Antipolis

first: Kevin

last: Lebrigand

address: 660 route des lucioles

institute: IPMC/CNRS

summary: To identify putative novel specific targets of miR-34a, miR-122, miR-206 and miR-210, we overexpressed these miRNAs in human Hela cells by transfecting them with synthetic pre-miRNAs or a synthetic “negative” pre-miRNA as control (miR-Neg1). RNA samples were harvested at 48 hours post-transfection and 2 independent experiments were carried out.

overall_design: 48 hours post-transfection, 2 independent experiments performed in dye-swap: miR-34a versus miR-Neg1 ; miR-122 versus miR-Neg1 ; miR-206 versus miR-Neg1 ; miR-210 versus miR-Neg1.

first: Monsef

last: Benkirane

first: Bernard

last: Mari

first: Pascal

last: Barbry


title: Early-life stress of Cntnap2 heterozygous mice demonstrates a gene by environment interaction model for developmental brain disorders

city: Lincoln

first: Qi

last: Zhang

state: NE

address: 354C Hardin NW

department: Statistics

institute: University of Nebraska Lincoln

summary: We examined whether adult heterozygous mice (Cntnap2+/-; phenotypically normal) subjected to an early-life stress would have social-related deficits and disruptions in 5-hydroxymethylcytosine (5hmC), similar to the Cntnap2-/-.

overall_design: Profiled genome-wide 5hmC and RNA expression in the striatum of prenatal stressed and non-stressed mice that were wild type or heterozygous for the Cntnap2 gene

first: Reid

last: Alisch

first: Ligia

last: Papale


title: Inflammatory fibroblasts mediate resistance to neoadjuvant therapy in rectal cancer

status: Public on Dec 18 2021

city: Frankfurt

first: Florian

last: Buettner

state: Germany

address: Theodor-Stern-Kai 7

department: DKTK Frankfurt

institute: DKFZ

summary: Standard cancer therapy targets tumor cells while not considering the possible collateral damage on the tumor microenvironment that could impair therapy response. Employing patient-derived tumor organoids and primary stroma cells or a novel murine rectal cancer model, we show that tumor cell-derived IL-1a polarizes cancer-associated fibroblasts (CAFs) towards a pro-inflammatory phenotype causing an elevated oxidative DNA damage mediated by reactive nitrogen species (NOS) and subsequently sensitizing iCAFs to a p53-mediated therapy-induced senescence, which triggers changes in matrisome composition culminating in chemoradiotherapy resistance and disease progression. Consequently, IL-1 inhibition, prevention of iCAF senescence or senolytic therapy sensitizes mice to irradiation. Importantly, rectal cancer patients with poor response to radiotherapy are characterized by an increased number of CAFs, a dominant inflammatory CAF gene signature as well as lower IL-1 receptor antagonist (IL-1RA) serum levels. Low baseline IL1RA gene expression predicts poor outcome while IL-1RA serum levels are associated with a single nucleotide polymorphism (SNP) in IL1RA (rs4251961 T/C). Moreover, conditioned supernatant from patient tumor organoids sensitizes fibroblasts to undergo radiation-induced senescence in an IL-1-dependent manner. Collectively, we unravel a critical role for inflammatory CAFs in cancer therapy and identify IL-1 signaling as an attractive target for stroma-repolarization and prevention of CAF senescence.

overall_design: single-cell RNA-seq of CAFs

last: Greten

first: Adele

last: Nicolas


title: Gene expression profiling of Huh7 and THP1 cells after siRNA knockdown of C5/TRAF1 intergenic transcript

city: Rotterdam

first: Rute

last: Marques

address: Dr Molewaterplein 50

institute: Erasmus MC

summary:
The C5/TRAF1 locus is a well validated genetic risk factor for RA. Upon further investigation of this region we recently discovered that the intergenic region between these two immune-related genes is activelly transcribed in a variety of tissues and is up-regulated in PBMCs, monocytes, B- and T-cells upon immune stimulation.
The present study aimed to characterize the biological function of this newly identified C5/TRAF1 intergenic transcript, by analysing the effect of its knockdow on the gene expression profiles of Huh7 and THP1 cells.

overall_design: siRNA transfections with the intergenic specific siRNA and 2 negative control siRNAs were performed in duplo and independently repeated 4 times per cell line, with different cell culture passages to account for the biological variation. The 4 independent experiments per cell line were divided in two design phases: the Discovery Phase (experiments 1 and 2) and the Validation Phase (experiments 3 and 4).

first: Rene

last: Toes


title: Genome-wide profiling of DNA G-quadruplexes in living neurons

city: Yokohama

first: Sefan

last: Asamitsu

state: Kanagawa

address: 1-7-22, Suehiro-cho, Tsurumi-ku

country: Japan

institute: RIKEN Center for Integrative Medical Sciences

summary: We report a genome-wide mapping of DNA G-quadruplexes and histone modifications in living neurons. We used primary cultured neurons from the fetus mouse cortex. Cultured cells were treated with 2 µM Ara-C at DIV3 to prevent the proliferation of mitotic cells and were harvested at DIV10. For iG4 CUT&Tag, a BG4-expressing plasmid was transfected by electroporation at DIV0.

overall_design: Examination of G4DNAs and four different histone modifications in post-mitotic neurons.

first: Masayuki

last: Takeuchi

first: Kohei

last: Maeda

first: Shingo

last: Usuki

first: Kazuya

last: Matsuo

first: Yasushi

last: Yabuki

first: Norifumi

last: Shioda


title: Comparative analysis of testis transcriptome between a genetic male sterile line (GMS) and its wild-type 898WB in silkworm, Bombyx mori

status: Public on Dec 17 2021

city: Zhenjiang

first: Xinglin

last: Mei

state: Jiangsu

address: No.666, Changhui Road, Dantu New District, Zhenjiang city, Jiangsu Province

institute: Jiangsu University of Science and Technology

summary:
Purpose:The purpose of study these processes, is helpful to understand the mechanisms of genetic sterility of lepidopteran insects and achieve sterile insect technique (SIT) for pest control.
Methods:Intact testes were dissected on day 5 of fifth-instar larvae and immediately frozen in liquid nitrogen for RNA-Seq. The testes from three silkworms were taken as one biological sample, and three samples per strain were prepared as biological replicates.
Results: A total of 1,872 up-regulated genes and 1,823 down-regulated genes were detected in the testis of the GMS mutant. Several genes play important roles in spermatogenesis and testicular development, such as “serine/threonine protein kinase”, “organic cation transporter protein”, “tyrosine protein kinase”,“lncRNAs” and “immune-associated genes”, and so on. GO (Gene ontology) annotation and KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway analysis was performed for functional annotation of the differentially expressed genes (DEGs). KEGG pathway analysis showed that the DEGs annotated to 123 pathways, and 10 pathways were enriched significantly, such as “metabolic pathway”, “biosynthesis of amino acids”, and “phagosome–lysosome pathway” associated with the testis development and spermatogenesis. The qPCR expression results were similar to the results obtained from the RNA-seq data, which showed the sequencing results were reliable.
Conclusions:Differentially expressed genes of the testes between GMS mutant and 898WB were screened by RNA-Seq technology, which provide reliable reference for understanding the mechanism of male sterility in GMS mutant.

overall_design: comparative transcriptome analysis of testes between three biological replicates of the GMS mutant and wild strain 898WB.

first: Mengjie

last: Gao

first: Tianchen

last: Huang

first: Qiaoling

last: Zhao


title: Targeted inhibition of TAK1 phosphorylation at Thr184,184 sites by Tubuloside B to improve rheumatoid arthritis synovium function

city: Shenzhen

first: Yuyang

last: Miao

state: Guangdong

address: Longguan Road 101

department: Longhua Branch

institute: Shenzhen Peoples‘s Hospital

summary: Rheumatoid arthritis (RA) is an intractable chronic inflammatory disease characterized by synovial hyperplasia and consecutive inflammation. Tubuloside B (Tub B) is the major active constituent isolated from Cistanche tubulosa (Schenk) Wight which is a traditional Chinese medicine with anti-inflammatory efficacy. In this study, we aimed to examine the ability of Tubuloside B to treat RA and explore the underlying mechanisms.

overall_design: Briefly, 2 mL cell suspension was seeded into a 6-well plate (1 × 10^6 cells/well) and cultured in DMEM with 10% FBS for 24 h in a CO2 incubator at 37°C. Then, the cell samples were divided into four groups: the mock group was incubated as normal, the model group was stimulated by TNF-α (20 ng/mL) for 24hand the TNF+Tub B-treat group was treated with 60 μM Tub B at the same time as TNF-α stimulation for 24 h,the Tub B-treat group was treated with 60 μM Tub B for 24h

first: Chenyan

last: Shi

first: Dongyu

last: Liu

first: Xiaobin

last: Zeng


title: Mycn regulates intestinal development through ribosomal biogenesis in a zebrafish model of Feingold syndrome 1 [bulk RNA-seq]

city: Hang Zhou

first: Tao

last: cheng

state: Zhe Jiang

address: 886 Yuhangtang Road

department: Institute of genetics

institute: Zhejiang University

summary:
Purpose: To identify differential expressed genes for loss of function of Mycn in zebrafish
Methods: wild-type and mycn mutant zebrafish embryos were collected for RNA sequencing at 48 hpf (hours post fertilisation) and 72 hpf
Results: We found 85 (overlapped) down-regulated genes in mycn mutant embryos at 48 hpf and 72 hpf  (padj < 0.1, log2 fold change < 0)
Conclusions: 85 down-regulated genes were identified

overall_design: comparison of tanscriptome between wild-type and mycn mutant zebrafish embryos at 48 hpf and 72 hpf

last: Cheng

first: Yunfei

last: Li


title: Microarray expression profile and bioinformatics analysis of circular RNAs in the phenotypic switching of rat vascular smooth muscle cells: a preliminary study

city: Xi'an

first: Bo

last: Yuan

address: No. 229, Taibai North Road, Beilin District

institute: Northwest University

summary: In order to further study the role of circular RNA in the phenotypic transformation of vascular smooth muscle cells (VSMCs), the differential expression profile of circRNA in the phenotypic transition of VSMCs induced by platelet-derived growth factor-BB (PDGF-BB) was screened using chip technology. Vascular smooth muscle cells from rat thoracic aorta were induced with 20ng/ml PDGF-BB as the experimental group and compared with the control group. After induction for 24 hours, the differentially expressed circRNA was screened by circular RNA chip.

overall_design: In the experimental group induced by PDGF-BB and the control group, three samples were selected for circular RNA chip screening. According to FC ≥ 1.5; P <0.05, 6 differentially expressed circRNAs were screened out and their differential expression was further verified by reverse transcription quantitative polymerase chain reaction (RT-qPCR). The interaction between these circRNAs and miRNAs was predicted through bioinformatics analysis, and further studies were carried out using GO and KEGG pathway analysis

first: Yuqiang

last: Ji


title: 1,25(OH)2D3 and VDR, Open chromatin

city: Iwaki

first: Yoshiaki

last: Kanemoto

state: Fukushima

address: kaminodai 57, jyobankamiyunagaya-town

institute: Jyoban Hospital

summary: The purpose of this study are to ensure that  whether calcitriol upregulates open chromatin in keratinocytes or not.

overall_design: ATAC Seq in HaCaT WT and VDR KO treated with 1,25(OH)2D3 or not.

first: Rei

last: Amano

first: Takahiro

last: Sawada

first: Jinichi

last: Mori

first: Shigeaki

last: Kato


title: Mycn regulates intestinal development through ribosomal biogenesis in a zebrafish model of Feingold syndrome 1

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

overall_design: Refer to individual Series


title: Single-cell RNA sequencing of repopulated microglia at day 3

city: Hangzhou

first: weiying

last: wu

state: Zhejiang

address: 866 Yuhangtang Rd

summary: To probe potential molecular mechanisms underlying the differences between wild-type and Hk2-cKO microglia during repopulation, we performed the single-cell RNA sequencing (scRNA-seq) of repopulated microglia at Day 3. Microglia were isolated by FACS as CD11b high CD45 low cells from normal brains and Hk2-cKO brains.

overall_design: Hk2 fl/fl and Hk2 cKO repopulated microglia at day3 were isolated by FACS as CD11b high CD45 low cells.

first: Zhihua

last: Gao

first: Yaling

last: Hu

first: Kelei

last: Cao


title: Surface proteomics reveals CD72 as a target for in vitro-evolved nanobody-based CAR-T cells in refractory B-cell malignancies II

city: San Francisco

first: Matthew

last: Nix

state: CA

address: 185 Berry St

department: Laboratory Medicine

institute: UCSF

summary: Alternate strategies are needed for B-cell malignancy patients relapsing after CD19-targeted immunotherapy. Here, integrated cell surface proteomics and epigenetic analysis initially revealed CD72 as an optimal target for poor-prognosis MLL-rearranged B-ALL, which we further found to be expressed widely across B-cell malignancies. Using a recently-described, fully-in vitro system we selected CD72-specific nanobodies, incorporated them into CARs, and demonstrated robust activity against B-cell malignancy models, including CD19 loss. “Antigen escape profiling” modeled membrane proteome changes in the context of CD72 loss while pharmacologic SHIP1 inhibition increased CD72 surface density. We establish CD72-nanobody CAR T’s as a promising therapy for refractory B-cell malignancies.

overall_design: biological duplicates of a cell surfaceome focused CRISPRi dropout screen of essential membrane proteins in the B-ALL cell line SEM, analyzed by Illumina deep sequencing

middle: A

first: Arun

last: Wiita


title: Mycn regulates intestinal development through ribosomal biogenesis in a zebrafish model of Feingold syndrome 1 [single-cell RNA-seq]

summary:
Purpose: To systematically investigate the phenotypes resulting from Mycn loss-of-function at a higher resolution.
Methods:  Approximately 30 mycn mutant or WT zebrafish embryos at 72 hpf (hours post fertilization) were harvested . Libraries were prepared using Chromium Controller and Chromium Single Cell 3’Library & Gel Bead Kit v3 (10x Genomics, PN-1000074) according to the manufacturer’s protocol for 10000 cells recovery.
Results: A total of 25,886 single-cell transcriptomes were collected after stringent quality control measures.
Conclusions: 27 Cell types were identified by intergrating two datasets and 9 subclusters were obtained by reclustering intestinal cells.

overall_design: wild-type or mycn zebrafish embryos at 72 hpf were harvested for scRNA-seq.