title: Disruption of Mitochondria-Associated Endoplasmic Reticulum Membrane (MAM) integrity identified in FUS‑mutant oligodendroglial progeny from human induced pluripotent stem cells

city: Leuven

first: Yingli

last: Zhu

address: ON4 Herestraat 49

institute: KULeuven

summary: Amyotrophic lateral sclerosis (ALS) is a rapidly progressive, fatal neurodegenerative disorder, characterized by selective loss of motor neurons (MNs). A number of causative genetic mutations underlie the disease, including mutations in the fused in sarcoma (FUS) gene, which can lead to both juvenile and late onset ALS. Although ALS results from MN’s death, there is evidence that dysfunctional glial cells, including oligodendroglia, contribute to neurodegeneration. Here, we used human induced pluripotent stem cells (hiPSCs) with a FUSR521H or a FUSP525L mutation and their isogenic controls to generate oligodendrocyte progenitor cells (OPCs) by inducing SOX10 expression from a TET-On SOX10 cassette as described previously in our lab. Mutant and control iPSC differentiated efficiently into OPCs. RNA sequencing identified a myelin sheath-related phenotype in mutant OPCs. Subsequent lipidomics studies demonstrated defects in myelin lipids, with a reduction of phospholipids in mutant OPCs. Among them, phosphatidylcholine (PC) and phosphatidylethanolamine (PE) are the most two abundant phospholipid in mammalian cells and regulate the mitochondria-associated ER membrane (MAM) integrity. Interestingly, FUSR521H mutant OPCs displayed a decrease in the PC/PE ratio, known to be associated with maintaining membrane (mitochondria) integrity. Proximity ligation assay further indicated that MAM was diminished in mutant OPCs. Moreover, mutant OPCs displayed stronger activation of unfolded protein response markers, a hallmark of increased susceptibility to endoplasmic reticulum (ER) stress, when exposed to thapsigargin, and exhibited impaired mitochondrial respiration and reduced Ca2+ signaling from ER Ca2+ stores. Taken together, these results demonstrate that FUS mutant OPCs display defects in lipid metabolism associated with MAM disruption manifested by impaired mitochondrial metabolism with increased susceptibility to ER stress and suppressed physiological Ca2+ signaling events.

overall_design: RNA Samples of FUS‑mutant oligodendroglia from human induced pluripotent stem cells were analyzed

first: Katrien

last: Neyrinck

first: Jonathan

last: De Smedt

first: Catherine

middle: M

last: Verfaillie


title: Single cell transcriptome signature of hepatocellular carcinoma (HCC) at different stages along with tumor development

status: Public on Jan 01 2024

city: Shanghai

first: YuLi

middle: yang

last: Li

address: Dongan Road

department: 2Department of Immunology

country: China

institute: Fudan University

type: Expression profiling by high throughput sequencing

summary: HCC possesses a highly heterogeneous tumor micro-environment with dynamic populations and interaction along tumor progression. We used single cell RNA sequencing (scRNA-seq) to analyze cell composition and features of HCC. A subgroup of tumor-associated macrophages characterized with distinctive metabolic activity was identified. We further excavated its functional phenotype as well as interaction with other components and investigated the underlying molecular mechanism.

overall_design: Liver dissection of the gene-induced HCC models at four different timepoints including 0 week (HCC-NL), 4 week (HCC-ES), 8 week (HCC-MS), and 12 week (HCC-LS) along tumor development were conducted and prepared into single-cell suspensions and analyzed using scRNAseq.

first: Yuli

last: Lin


title: Role of PLK1 in epithelial-mesenchymal transition in human melanoma

city: Madison

first: Nihal

last: Ahmad

state: Wisconsin

address: WIMR II, 1111 Highland Ave

department: Dermatology

country: USA

institute: University of Wisconsin

summary:
Polo-like kinase 1 (PLK1), a serine/threonine kinase and important cell cycle regulator, is overexpressed in melanoma and its expression has been associated with poor disease prognosis. PLK1 has been shown to interact with NUMB, a NOTCH antagonist. However, the exact role of PLK1-NUMB-NOTCH axis in epithelial-mesenchymal transition (EMT) and melanoma progression is unclear.
In this study, Affymetrix microarray analysis was performed to determine differentially expressed genes following doxycyclin induced shRNA-mediated knockdown of PLK1 in human melanoma cells that showed significant modulations in EMT and metastasis related genes.

overall_design: Total RNA (biological triplicates) was isolated using an RNeasy Plus Mini Kit (QIAGEN, CA) from experimental groups. The RNA samples were hybridized to the Affymetrix Human Transcriptome Array (HTA) 2.0 microarrays and processed by the UW-Madison Gene Expression Center in the UW Biotechnology Center for Affymetrix GeneChip Services.

first: Gagan

last: Chhabra

first: Chandra

middle: K

last: Singh

first: Mary

middle: A

last: Ndiaye


title: Biogenerated oxygen-related environmental stressed apoptotic body targets endothelial cells

first: Qiuyu

last: Zhao

address: 639 Zhi Zao Ju Road,

institute: Shanghai Ninth People’s Hospital, Shanghai JiaoTong University School of Medicine

summary: he dynamic balance of hypoxia and oxidative stress constitutes the oxygen-related microenvironment in injured tissues. Oxygen homeostasis is highly variable; therefore, it is not a therapeutic target for injured tissue repair. We found an enrichment and extensive apoptosis of mesenchymal stem cells (MSCs) infused intravenously in the wound microenvironment with co-existing hypoxia and oxidative stress. Apoptotic bodies (ABs), generated from in situ apoptosis, significantly promotes angiogenesis. We improved the derivation pathway of ABs by simulating oxygen homeostasis in injured tissues, with cobalt chloride-induced hypoxia or hydrogen peroxide-induced oxidative stress in MSCs. Oxygen-related environmental stressed ABs, derived from environments of hypoxic and oxidative stress, were identified and loaded onto hydrogel microspheres for accurate regulation of endothelial cells (ECs) vascularization. These ABs directly targeted ECs; oxidative stress ABs (Oxi-ABs) have a 2.5- and 4-fold higher tube-forming ability than hypoxic and normoxic ABs, respectively. miRNA microarray analysis revealed that different oxygen-stressed ABs deliver similar miRNAs, which leads to the broad upregulation of EC phosphokinase activity. Finally, local delivery of Oxi-ABs-loaded hydrogel microspheres promotes wound healing. Oxi-ABs-loaded hydrogel microspheres achieved controlled AB release, targeting EC by reducing the consumption of early inflammatory cells and adapting to the proliferative phase of wound healing. Thus, the biogenerated apoptotic bodies responding to oxygen-related environmental stress can target ECs to promote vascularization.

overall_design: After co-culturing with PBS or 100 μM CoCl2 or 50 μM H2O2 for 24 h, MSC were induced to apoptosis and miRNA expression was measured in the harvested apoptotic vesicles. Three independent experiments were performed for each individual culture condition, using a different batch of cells for each experiment.

first: Bolun

last: Lu


title: Effect of interleukin-10 on primary oligodendrocyte progenitor cells under hemin administration

status: Public on Jan 03 2024

city: Beijing

first: Weihua

last: Wu

address: No.10 Xitoutiao, You'anmenWai, Fengtai District

department: Department of Biochemistry and Molecular Biology

institute: Capital Medical University

summary: To investigate the gene expression changes in primary  oligodendrocyte progenitor cells generated from wild-type mouse pups after hemin or hemin+interleukin-10 or hemin+interleukin-10+stattic treatment

overall_design: Ttreat wild-type mouse primary  oligodendrocyte progenitor cells with veh or hemin (50 μM) or hemin  (50 μM)+interleukin-10 (10 ng/mL) or hemin (50 μM)+interleukin-10  (10 ng/mL)+stattic (2.5 μM) for 12 h


title: Monotherapy of the DNA Hypomethylating Agent, NTX301 Targets Ovarian Cancer

city: Chicago

first: Daniela

last: Matei

address: 420 E. Superior St.

institute: Northwestern University Feinberg School of Medicine

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

overall_design: Refer to individual Series


title: Monotherapy of the DNA Hypomethylating Agent, NTX301 Targets Ovarian Cancer (RNA-Seq)

summary: DNA methylation causes silencing of tumor suppressor and differentiation-associated genes being linked to chemoresistance and stemness. Previous studies demonstrated that hypomethylating agents (HMA) re-sensitize ovarian cancer (OC) cells to chemotherapy, however, translation to the clinic has been slow. NTX 301 (PinotBio) is a novel, highly potent and orally bioavailable HMA, in early clinical development. We assessed the anti-tumor effects of NTX301 in OC models and studied the underlying molecular mechanisms by using cell proliferation, stemness and ferroptosis assays, RNA sequencing and validation. OC cells (SKOV3, IC50=5.089nM; OVCAR5 IC50=3.664nM) were highly sensitive to NTX301 (p<0.05) compared to immortalized fallopian tube epithelial cells (FT-190) (IC50=103.3nM). Treatment with NTX301 induced significant downregulation of the DNA methyltransferases (DNMTs) 1-3 expression in SKOV3 and OVCAR5 cells compared with decitabine (p<0.05). Treatment with low dose NTX301 (100nM) induced significant transcriptomic reprogramming with 15,000 differentially expressed genes (DEGs) compared to DMSO (p<0.05). Among the NTX301 down-regulated DEGs, Gene Ontology (GO) enrichment analysis identified pathways related to regulation of alcohol, cholesterol, and fatty acid biosynthetic process and molecular functions related to aldehyde dehydrogenase (ALDH) and oxidoreductase activity, known features of cancer stem cells (CSCs). Indeed, treatment with low dose NTX301 (100nM) reduced the ALDH(+) cell population and expression of stemness associated transcription factors (Oct4 and Sox2).  Additionally, stearoyl-Coenzyme A desaturase 1 (SCD), a key enzyme that regulates unsaturated fatty acid homeostasis in the lipogenesis process was found among the top DEGs downregulated in response to NTX301 (fold change=9.36, FDR<0.05). As blockade of SCD had been shown to induce ferroptosis, oxidized lipids levels were measured by C11-BODIPY staining. NTX301 treatment increased the levels of oxidized lipids compared to DMSO and this was blunted by deferoxamine, indicating that NTX301 promoted cell death via ferroptosis. NTX301 induced ferroptosis was successfully rescued by addition of oleic acid (200mM) into the culture medium, supporting that it was mediated through SCD depletion.  In vivo, monotherapy with NTX301 (1mg/kg), significantly inhibited subcutaneous OC and PDX xenograft tumor growth (p<0.05). Decreased SCD levels and increased oxidized lipids were recorded in NTX301-treated xenografts. In conclusions, NTX301 is a potent HMA active in OC models. Our findings point to a new mechanism by which epigenetic blockade disrupts lipid homeostasis and promotes cancer cell death.

overall_design: We assessed the anti-tumor effects of NTX301 in ovarian caner models and studied the underlying molecular mechanisms by using RNA sequencing and HumanMethylation 450 array.

first: Yinu

last: Wang

first: Xiaolei

last: Situ

first: Horacio

last: Cardenas

first: Ellie

last: Siu

first: Sayedabdulrazzaq

last: Alhunayan

first: Russell

last: Keathley

first: Eilidh

last: Obrien

first: Jian-Jun

last: Wei

first: Yuying

last: Tan

first: Edward

last: Tanner

first: Chinmayee

middle: P

last: Dessai

first: Ji-Xin

last: Cheng


title: Methylation profiling of  choroid plexus carcinoma cell lines

city: Cairo

first: Shahenda

last: El-Naggar

address: 1 Sekket El Emam, El Madbah El Kadeem Yard, Sayeda Zeinab

department: Research Department

country: Egypt

institute: Children’s Cancer Hospital Egypt 57357

summary: Genomic DNA was extracted from two choroid plexus carcinoma cell lines using the GeneJET genomic DNA (Thermo Fisher).  DNA was quantified using the DENOVIX Fluorometer. The quality of the extracted DNA samples was  assessed by the Illumina FFPE QC kit (Illumina). Bisulfite conversion of the DNA was performed  using the EZ DNA methylation kit (Zymo). Bisulfite-converted DNA was hybridized to the Illumina  Infinium Human Methylation EPIC array bead chip and scanned using the Illumina iScan microarray  scanner (Illumina) according to the manufacturer protocol.

overall_design: In total, we have 2 samples of primary choroid plexus carcinoma cell lines used for the Illumina Infinium 850K EPIC Methylation array.

first: Nada

last: Amer


title: LT-HSC RNA-seq of 5-FU treated mice

city: Hangzhou

first: lu

last: huan

address: 866 Yuhangtang Road

institute: Zhejiang University medical center

summary: Mice were pre-conditioned with 5-FU, and then treated with Ferumoxytol (FMT) to determine the regeneration of HSCs

overall_design: Mice were pre-conditioned with 5-FU (250 mg/kg body weight), and then treated with PBS (5-FU+Saline) or FMT (5-FU+FMT),  LT-HSCs were sorted to perform RNA-seq. Control represented the untreated mice.

first: Qiwei

first: Huan

last: Lu

first: Pengxu

last: Qian


title: Transcriptional profiles of peripheral eosinophils in COPD and asthma - a preliminary study

city: Warszawa

first: Katarzyna

last: Mycroft

address: ul. Banacha 1a

department: Department of Internal Medicine, Pulmonary Diseases and Allergy

country: Poland

institute: Medical Univesity of Warsaw

summary: The role of eosinophilic inflammation in chronic obstructive pulmonary disease (COPD) pathogenesis is unknown and is probably different than in asthma. The molecular processes underlying the differences between eosinophils from asthma and COPD have not been studied. The study group included 5 patients with asthma and 4 patients with COPD. The RNA-Seq data analysis identified 26 differentially expressed genes between COPD and asthma (according to adjusted p-value). In total, 6 genes were up-regulated (CCL3L1, CCL4L2, SERPINB2, PRSS21, GPR82) and 20 were down-regulated (e.g. JUN, IFITM3, DUSP1, GNG7, ZNF107, BCL6) in peripheral eosinophils of COPD patients compared to asthma. Biological processes associated with down-regulated genes were cell differentiation, positive regulation of RNA metabolic process. The genes associated with signaling of IL-4 and IL-13 pathway were down-regulated in COPD eosinophils compared to asthma. Peripheral eosinophils from COPD and asthma patients present different transcriptomic profiles suggesting their different function in pathobiology of both obstructive airway diseases.

overall_design: The objective of this study was to compare the transcriptomic profiles of blood eosinophils in COPD and asthma.Eosinophils were isolated from peripheral blood drawn from stable mild-to-moderate COPD and asthma patients. RNA was isolated from eosinophils and was prepared and sequenced using an NGSelect RNA. The prepared libraries were then quality-checked, pooled, and sequenced on an Illumina platform.

first: Małgorzata

last: Proboszcz

first: Magdalena

last: Paplińska-Goryca

first: Rafał

last: Krenke

last: Górska


title: BET inhibitor JQ-1 combined with cardamonin can inhibit the invasion and metastasis of triple-negative breast cancer cells

first: Xinrui

last: Wang

address: 197 Ruijin Rd II

institute: Rui-Jin Hospital affiliated to Shanghai Jiao Tong University School of Medicine

summary: Purpose: The BET family protein BRD4 is an important anti-tumor target and is highly expressed in breast cancer. However, BET inhibitors are susceptible to drug resistance. The aim of this study is to explore the mechanism of BET inhibitor JQ-1 combined with cardamonin in the anti-invasion and metastasis of triple-negative breast cancer. Methods: triple-negative breast cancer cells mRNA of DMSO (Control), JQ-1,cardamonin and combined groups were progressed with deep sequencing, in triplicate through Illumina sequencing platform (HiSeqTM 2500). Genes with adjusted P-value < 0.01 and |foldChange| > 2 were defined as differentially expressed genes (DEGs) and subjected to the following GO and KEGG enrichment. qRT-PCR was performed to validate several essential genes using QuantStudio 5 (Thermo Fisher) and SYBR Green assays. Results: As an inhibitor of mTOR, Cardamonin(CAR) can inhibit the proliferation of tumors. Research have found that BET inhibitor can affect the function of a large number of genes by regulating the epigenome, which plays an important role in a variety of diseases. In this study we explored the JQ1 which is a BRD4 protein inhibitor can inhibit the proliferation and metastasis of MDA-MB-231 cells in triple-negative breast cancer cell lines, and this phenomenon can be significantly enhanced when CAR is combined with JQ-1, and the same results are shown in mouse tumor formation experiments and triple-negative breast cancer organoids. At the same time, RNA-seq was performed in MD1-MB-231 cells to explore the changes of specific molecular signaling pathways. We have mapped over 40 million sequence reads to the MDA-MB-231 cell line genome in each specimen.Compared with the control group, CAR and JQ1, the combination group has significantly enriched gene expression. KEGG and GO enrichment analysis indicate that the combination group showed a series of biological processes such as macroautophagy, negative regulation of cell growth, lipid catabolic process, which led to decreased cell viability and decreased proliferation and metastasis ability. Conclusions: In this study, we demonstrated that JQ-1 combined with cardamomin could inhibit the invasion and metastasis of triple-negative breast cancer MDA-MB-231 cells, induce cell cycle arrest and promote cell apoptosis. The results of RNA-Seq were consistent with our in vitro and in vivo experiments, which demonstrated that the combination of JQ-1 and cardamonin effectively inhibited the proliferation, invasion and metastasis of MDA-MB-231 cells.

overall_design: mRNA expression profiles of DMSO,JQ-1,CAR, and Combination groups.

first: Liping

last: Zhu

first: Zhoujie

last: Ye


title: Monotherapy of the DNA Hypomethylating Agent, NTX301 Targets Ovarian Cancer (Methylation)

first: Daniela

last: Matei

first: Yinu

first: Xiaolei

last: Situ

first: Horacio

last: Cardenas

first: Ellie

last: Siu

first: Sayedabdulrazzaq

last: Alhunayan

first: Russell

last: Keathley

first: Eilidh

last: Obrien

first: Jian-Jun

last: Wei

first: Yuying

last: Tan


title: The dysregulated inclusion of BOLA3 exon 3 promoted by HNRNPC accelerates the progression of esophageal squamous cell carcinoma in esophageal squamous cell carcinoma

first: bo

last: tian

address: No.168, changhai road

institute: changhai hospital


title: CD45+ single cell transcriptomes in the MC38 tumor microenvironment

city: Salt Lake City

first: W. Zac

last: Stephens

state: UT

address: 15 N. Medical Drive, EEJMRB 4208, EEJMRB 4208

department: Pathology

institute: University of Utah

summary: Colorectal cancer (CRC) continues to increase globally, thus identification of mechanisms that prevent cancer are sorely needed. The microbiota has emerged as a key driver of CRC pathogenesis; however, little is known regarding how specific microbes might prevent CRC. Using transplantation of the microbiota from individuals with or without CRC into germfree mice, we identified that the microbiota harbored by non-diseased individuals can reduce tumor formation and Bacteroides uniformis was a potentially protective member of the microbiota. Single cell sequencing of CD45+ immune cells within the MC38 tumor shows a change in T and NK cell populations with B. uniformis treatment. T cell deficient mice were still protected from CRC in response to B. uniformis treatment, while NK cell depletion eradicated microbe-mediated protection. B. uniformis is reduced in individuals with CRC and thus our data identify B. uniformis as a microbe enriched in healthy individuals that can reduce tumor formation through enhanced NK cell activity.

overall_design: Sorted CD45+ cells from the MC38 tumor microenvironment from mice with or without B. uniformis oral gavage supplementation. Cells from 10 male or 10 female mice were pooled for each sample.

first: W

middle: Z

first: Allison

middle: W

last: Weis


title: Transcriptomic analysis of adult ovaries and testes exposed to the cocktails of pharmaceuticals ibuprofen (IBU), 2hydroxyIBU (2hIBU), diclofenac (DCF) and 17a ethinyl-estradiol (EE2) in mice

city: Montpellier

first: Brigitte

last: Boizet Bonhoure

address: 141 rue de la Cardonille

department: Human Genetic Institute

country: France

institute: CNRS UMR9002

summary: The endocrine disrupting pharmaceuticals, non-steroidal anti-inflammatory drugs (NSAIDs) and 17α-ethinyl-estradiol (EE2), are among the most relevant molecules found in aquatic ecosystems, surface and drinking water due to their incomplete removal by wastewater treatment plants. Exposure to therapeutic doses has a negative impact on gonadal development and fertility in rodent models; however, the effects of their chronic exposure at lower doses are not known. In this study, we investigated the impact of chronic exposure to a mixture containing ibuprofen, 2hydroxy-ibuprofen, diclofenac, and EE2 at two environmentally relevant doses (added to the drinking water from fetal life, 8.5 dpc, until sexual maturity) on the reproductive tract in F1 exposed mice and their F2 offspring. In adult F1 and F2 animals, chronic exposure to low environmental doses of IBU, 2hydroxy-IBU, DCF, and EE2 mixtures, affected reproductive organ maturation, estrous cyclicity (in females), spermiogenesis (in males), and some sperm parameters in F2 males. Transcriptomics further revealed significant changes in gene expression patterns and associated pathways, underlying the modified mechanisms on testis and ovarian physiology and on germ cells, the precursors of gametes. Consequently, fertility of F1 male and female animals exposed to the higher dose of pharmaceuticals and that of their F2 offsprings, measured through the total number of pups and the time between litters, was affected after 5 months of age.  This suggested that exposure to these drug cocktails has an inter-generational impact.

overall_design: Two mixtures containing environmentally relevant doses of IBU/2hIBU + DCF + EE2 (D1 and D2) were added or not (control; C) to the drinking water of pregnant mice and their progeny from 8.5 days post-coitum (dpc) until animal sacrifice (8 week-old for females and 10 week-old for males): F0 males and females from different litters were mated to minimize inbreeding and mated females were divided in three groups: control (no exposure), IBU/2hIBU+DCF+EE2 dose 1 (D1), and IBU/2hIBU+DCF+EE2 dose 2 (D2) (n= 6-7 females per exposure group). The IBU, 2hIBU, DCF and EE2 doses were calculated on the basis of their mean concentrations (5, 40, 10 and 1-2 ng/L, respectively) and maximum concentrations (50-100, 85-100, 50 and 20-50 ng/L, respectively) found in environmental drinking water samples. Thus, the calculated doses in the animal drinking water were: i) D1: IBU 11.3 ng/L/2hIBU 90 ng/L + DCF 22.5 ng/L + EE2 2.25 ng/L; and ii) D2: IBU 113 ng/L/2hIBU 225 ng/L + DCF 112 ng/L + EE2 45 ng/L. The control group was exposed to diluted ethanol (0.001%). Animals were reared in polyphenylsulfone (PPSU) cages in controlled environmental conditions (light/darkness: 12h/12h, 23°C) and drinking water was put in PPSU bottles wrapped in aluminum foil, and were fed with SAFE D131 that does not contain fish proteins, soy and alfalfa. Municipal tap water was provided ad libitum.

last: Boizet-Bonhoure

first: Pascal

last: Philibert


title: Perturbation of the circadian clock contributes to radiotherapy-induced salivary gland hypofunction

city: shenzhen

first: yang

last: kaibin

address: No.1066 Xueyuan Road, Nanshan District

institute: shenzhen university

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Salivary gland hypofunction is a common adverse effect during and after radiotherapy of head and neck cancers, resulting in the dry mouth syndrome called xerostomia. Previous studies suggested that the functionality of the salivary gland is under the regulation of the circadian clock, however, the extent and scope of this regulation remains unexplored. Here, we profiled the diurnal fluctuation of gene expression in the mouse submandibular salivary gland. We further analyzed the regulatory role of key circadian transcription factors Bmal1, Nr1d1 (Rev-erba), and Dbp, which revealed a wide range of potential down-stream target genes. The circadian clock was disrupted upon irradiation, as revealed by gene expression analysis. We propose that the mechanism of salivary gland hypofunction in radiotherapy involves perturbation of the circadian clock.

overall_design: For RNA-seq, we euthanized mice aged 6-8 weeks using the neck dislocation method, cut the skin along the neck to expose the submandibular gland, and then remove the gland tissue. Total RNAs were extracted using the Trizol method. Library construction and sequencing were performed on the DNBSEQ-T7 platform at the Geneplus Co, Shenzhen, China. Final data were presented as FPKM value for each gene. Tissue samples were collected at ZT3, ZT9 and ZT15 (10:00 am, 4:00 pm, and 10:00 pm). For Cut-taq, the mouse submandibular gland was minced and digested with Worthington Collagenase I at 0.2% concentration for 25-30 minutes at 37°C. The sample was filtered through a Falcon 40um filter, and the remaining tissue was incubated with 1x Gibco Trypsin/EDTA at 37°C for 20 minutes. After filtration, the sample was resuspended in PBS and centrifuged at 500 g for 5 minutes to obtain the supernatant. Standard operations were performed according to the instructions ofTD901-TD902 from Hyperactive In-Situ Chip Library Prep Kit for IIIumina. The 150 bp paired-end DNA sequencing was performed on the Illumina NovaSeq 6000 system (Haplox Genomics Center, Jiangxi, China).

first: Zhicao

last: Yue

first: Shiting

last: Song

first: Kaibin

last: Yang


title: The aberrant inclusion of BOLA3 exon 3 promoted by HNRNPC acts as an oncogenic driver in esophageal squamous cell carcinoma [11pairs RNA-seq]

summary: Aberrant RNA splicing events produce transcripts to facilitate ESCC progression, yet how this splicing process is abnomally regulated remains elusive. Here, we unveil a alternative splicing axis of BOLA3 transcripts and its regulator HNRNPC in ESCC.The long isoform of BOLA3 (-L) with exon3 is highly expressed in ESCC and associated with poor prognosis. Functional assays demonstrate the pro-tumorigenic function of BOLA3-L in ESCC cells. HNRNPC binds to BOLA3-L pre-mRNA and promotes exon3 inclusion forming BOLA3-L. HNRNPC knockdown suppresses the tumorigenesis of ESCC cells, phenocopying BOLA3-L knockdown, which is significantly rescued by BOLA3-L overexpression. High expression of HNRNPC is correlated with the exon-3 inclusion of BOLA3 in ESCC and associated with poor prognosis. BOLA3-L maintains mitochondrial structural and functional stability, exerting pro-oncogenic effects through the WNT/β-catenin pathway. E2F7 acts as a key transcription factor to regulate the transcriptional upregulation of HNRNPC and promotes the inclusion of BOLA3 exon3.Taken together, our findings provide insights into how alternative splicing contributes to ESCC progression, by investgating a HNRNPC-BOLA3 splicing axis.

overall_design: twenty two samples (including eleven tumor samples and eleven paracancerous samples) were used to detect the differential gene expression.Total RNA was extracted using the TRIzol reagent (Invitrogen, CA, USA) according to the manufacturer’s protocol. RNA purity and quantification were evaluated using the NanoDrop 2000 spectrophotometer (Thermo Scientific, USA). RNA integrity was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). Then the libraries were constructed using VAHTS Universal V6 RNA-seq Library Prep Kit according to the manufacturer’s instructions.The libraries were sequenced on an llumina Novaseq 6000 platform and 150 bp paired-end reads were generated. About xxxxx raw reads for each sample were generated. Raw reads of fastq format were firstly processed using fastp1 and the low quality reads were removed to obtain the clean reads.

first: Bo

last: Tian

first: Yan

last: Bian

first: Chu-Ting

last: Yu

first: Ye

last: Gao

first: Luo-wei

last: Wang

first: Lei

last: Xin

first: Han

last: Lin


title: Double-stranded RNA triggers a distinct integrated stress response in the early embryo

city: Qingdao

first: tong

last: lu

address: 72, binghai rd

institute: ShanDong University


title: The aberrant inclusion of BOLA3 exon 3 promoted by HNRNPC acts as an oncogenic driver in esophageal squamous cell carcinoma [shBOLA3-L RNA-seq]

first: bo

last: tian

overall_design: eight samples (including four normal control samples and four BOLA3-L isoform knockdown samples) were used to detect the differential gene expression.Total RNA was extracted using the TRIzol reagent (Invitrogen, CA, USA) according to the manufacturer’s protocol. RNA purity and quantification were evaluated using the NanoDrop 2000 spectrophotometer (Thermo Scientific, USA). RNA integrity was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). Then the libraries were constructed using VAHTS Universal V6 RNA-seq Library Prep Kit according to the manufacturer’s instructions.The libraries were sequenced on an llumina Novaseq 6000 platform and 150 bp paired-end reads were generated. About xxxxx raw reads for each sample were generated. Raw reads of fastq format were firstly processed using fastp1 and the low quality reads were removed to obtain the clean reads.


title: Double-stranded RNA triggers a distinct integrated stress response in the early embryo [dsRNA tp53]

summary: Gene functional study and the emerging mRNA therapy technology heavily rely on in vitro transcribed (IVT) mRNA, but the double-stranded RNA (dsRNA) by-product generated during IVT is often recognized by cells as a marker of virus replication, leading to the integrated stress response (ISR).The biological effects of dsRNA in undifferentiated cells differ significantly from those of differentiated counterparts.Here, we found that dsRNA triggered an unprecedented ISR in zebrafish early embryos, leading to developmental delay, severe cell necrosis, and lethal malformations.Mechanistically, dsRNA stimulation caused a PKR-eIF2α independent global translation inhibition and a burst of p53 signaling. In turn, p53 activated Interferon Stimulated Genes (ISGs) and other defense-related genes in the absence of the IFN system. Unexpectedly, p53 does not promote cell death; instead, it plays a protective role in weakening the damage of translation repression by compromising protein turnover via enhanced ISGylation.This study thus unveils a distinct ISR upon dsRNA stimulation, which may help perform faithful gene overexpression in early embryos and shed light on the strategies of undifferentiated cells against RNA virus infection.

overall_design:
To investigate the impact of the dsRNA-induced stress response, we compared the transcriptomes between uninjected and dsRNA-injected zebrafish MZtp53 embryos at 6 hpf
Comparative gene expression profiling analysis of RNA-seq data for MZtp53 uninjected embryos and its injected dsRNA embryos

first: Tong

last: Lu

first: Ming

last: Shao


title: The aberrant inclusion of BOLA3 exon 3 promoted by HNRNPC acts as an oncogenic driver in esophageal squamous cell carcinoma [shHNRNPC RNA-seq]

overall_design: eight samples (including four normal control samples and four HNRNPC knockdown samples) were used to detect the differential gene expression.Total RNA was extracted using the TRIzol reagent (Invitrogen, CA, USA) according to the manufacturer’s protocol. RNA purity and quantification were evaluated using the NanoDrop 2000 spectrophotometer (Thermo Scientific, USA). RNA integrity was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). Then the libraries were constructed using VAHTS Universal V6 RNA-seq Library Prep Kit according to the manufacturer’s instructions.The libraries were sequenced on an llumina Novaseq 6000 platform and 150 bp paired-end reads were generated. About xxxxx raw reads for each sample were generated. Raw reads of fastq format were firstly processed using fastp1 and the low quality reads were removed to obtain the clean reads.


title: H3K27ac profiling in liver sinusoidal endothelial cells through chromatin immunoprecipitation and sequencing

city: Rochester

first: Huihuang

last: Yan

state: Minnesota

address: 200 First Street SW

institute: Mayo Clinic

summary: Portal hypertension (PHTN) is a severe complication of liver cirrhosis. It is associated with intrahepatic sinusoidal remodeling induced by sinusoidal resistance and angiogenesis. Collagen type IV (COL4), a major component of basement membrane (BM), forms in liver sinusoids upon chronic liver injury. However, the cellular source and transcriptional regulation of COL4, as well as how it progresses in liver diseases is unknown. Here, we examined how COL4 is produced and how it regulates sinusoidal remodeling in PHTN. RNA-seq analysis on human cirrhotic livers revealed an increase in COL4 expression, which was further confirmed via immunofluorescence (IF) staining. scRNA-seq analysis identified Liver sinusoidal endothelial cells (LSECs) as the predominant source of COL4 expression in mouse liver, which was upregulated in a TNFα-NFκB dependent manner. Epigenetic repression of enhancer-promoter interaction silences COL4 gene expression via dCas9-KRAB-mediated epigenome editing approach. LSEC-specific COL4 mutation or repression abrogated sinusoidal resistance and angiogenesis, thereby alleviated sinusoidal remodeling and PHTN. Our findings reveal LSECs as a major source of COL4 in the liver which plays a prominent role in sinusoidal remodeling and PHTN.

overall_design: Chromatin immunoprecipitation DNA-sequencing (ChIP-seq) for histone modification H3K27ac in liver sinusoidal endothelial cell for identifying potential regulatory elements

first: Sheng

last: Cao

first: Can

last: Gan

first: Vijay

middle: H

last: Shah


title: The transcriptome profiling of multi-cell types by the droplet based full-length single-cell transcript sequencing

city: Guangzhou

first: Yicong

last: Xu

state: Guangdong

address: No. 135, Xingang Xi Road

institute: Sun Yat-sen University

summary: We developed a scalable Tn5-based Full-Length In situ Transcriptome sequencing method (scFLIT-seq). This method combines the advantages of in situ reverse transcription of methanol-fixed cells, single-cell separation and barcode ligation via the 10X scATAC-seq protocol and microfluidic platform, and Tn5-mediated fragmentation and tagging of cDNA. Various cells treated with different concentrations of methanol were used to validate the efficacy of this method. The efficacy at the single-cell level of this method was validated on a mixed cell population of HEK293T and NIH/3T3 cells.

sample_organism:
Homo sapiens
Mus musculus

platform_organism:
Homo sapiens
Mus musculus

overall_design: The reverse transcription efficiency analysis of the FLIT-seq under different conditions in multi-cell types.

first: Tao

first: Ziyang

last: An

first: Jin


title: mRNA cleavage by 21-nt phasiRNAs dependent on AGO1d is important for temperature-sensitive male sterility in rice

city: Shenzhen

first: Chuanlin

last: Shi

address: Dapeng

institute: Chinese Academy of Agricultural Sciences

summary: Temperature-sensitive male sterility is one of the core components for hybrid rice breeding based on two-line system. We previously found that knockout of ARGONAUTE 1d (AGO1d) caused temperature-sensitive male sterility in rice by influencing phased small interfering RNA (phasiRNA) biogenesis and function. However, the specific phasiRNAs and their targets underlying the temperature-sensitive male sterility in ago1d mutant remain to be determined. Here, we demonstrate that the ago1d mutant displays normal female fertility but complete male sterility at low temperature. Through a multi-omics analysis of small RNA, degradome, and transcriptome, we found that 21-nt phasiRNAs account for the greatest proportion of the 21-nt small RNA (sRNA) species in rice anthers and are sensitive to low temperature and markedly down-regulated in ago1d mutant. Moreover, we found that 21-nt phasiRNAs are essential for the mRNA cleavage of a set of fertility- and cold tolerance-associated genes, such as EDT1, TDR, OsPCF5 and OsTCP21, directly or indirectly determined by AGO1d-mediated gene silencing. The loss-of-function of 21-nt phasiRNAs can result in up-regulation of their targets and causes varying degrees of defects in male fertility and grain setting. Our results highlight the essential functions of 21-nt phasiRNAs in temperature-sensitive male sterility in rice and suggest their promising application in two-line hybrid rice breeding in the future.

overall_design: Anthers at S8-9 of wild type and ago1d mutant grown at 28 °C and 22 °C were collected with three biological replicates. The degradome libraries were constructed as previously described (German et al., 2009) with minor modifications. Total RNA was isolated from each sample and purified with oligo(dT) beads. The purified poly(A) RNAs containing a 5′ monophosphate of the 3′ cleavage product of the mRNA were ligated to 5’-adaptor. First-strand cDNA was reversely transcribed using 3′-adapter random primers, and size selection was performed with AMPureXP beads. Then the cDNAs were amplified with PCR by the following conditions: initial denaturation at 95 °C for 3 min; 15 cycles of denaturation at 98 °C for 15 sec, annealing at 60 °C for 15 sec, and extension at 72 °C for 30 sec; and then final extension at 72 °C. The final products were sequenced (single-end 50-bp) on an Illumina Hiseq2500 platform by LC-BIO (Hangzhou, China). Degradome sequencing data were preprocessed by removing adapters and then mapped to the Oryza sativa v.7.0 genome sequences (MSU Rice Genome Annotation Project) using bowtie. Target prediction and validation of the 3,116 sRNAs with the degradome sequencing was conducted by CleaveLand4 and GSTAr which are available at https://sites.psu.edu/axtell/software/cleaveland4/. The validated targets were defined with mismatched score ≤ 7 (specifically, mismatched query bases or target-bulged bases, are penalized 1. G-U wobbles are penalized 0.5. These penalties are double within positions 2-13 of the query.), category ≤ 2 (category 0: >1 read, equal to the maximum on the transcript, when there is just 1 position at the the maximum value; category 1: >1 read, equal to the maximum on the transcript, when there is >1 position at maximum value; category 2: >1 read, above the average depth, but not the maximum on the transcript; category 3: >1 read, but below or equal to the average depth of coverage on the transcript; category 4: Just one read at that position) and read counts >1 in wild type at both 28 °C and 22 °C.

last: shi

first: Wenli

last: Zou

first: Lianguang

last: Shang


title: RNA-seq analysis of peripheral nerve in Mtmr2 null model of CMT4B1.

first: John

last: Svaren

state: WI

address: 1500 Highland Ave.

department: Comparative Biosciences

summary: The goal of the experiment was to assess gene expression  in sciatic nerve of Mtmr2 null mice, which is a model of the genetic neuropathy known as CMT4B1, and response to niacin treatment.

overall_design: For each condition, four independent biological replicates were prepared (3 males and 1 female/group).

first: Shuang

last: Liu

first: Phu

last: Duong

first: Alessandra

last: Bolino


title: T-cell instructed monocyte activation is a key pathological feature in Ankylosing Spondylitis and provides novel therapeutic opportunities

city: Oxford

first: Feng

state: Oxfordshire

address: Botnar Research Centre, Windmill Road, Headingtong

country: United Kingdom

institute: University of Oxford

summary: Activated T-cells potently stimulated monocytes from patients with AS to produce IL-1β and IL-23.  These in turn act back on T cells to enhance Th17 responses. T-cell activated monocytes are heterogeneous but include a subset of “super proinflammatory monocytes” not induced by LPS activation and for enriched for AS risk genes, including ERN1. Importantly, we identify a population of CD14+ myeloid cells highly enriched in AS synovial fluid strikingly similar gene expression profile high levels of CCL3, CCL4, IL1B, TNF, CXCL2, strongly suggesting T cell activation in vivo.  a T-cell instructed monocyte activation can be inhibited by genetic or chemical suppression of ERN1.

overall_design: PBMCs isolated from AS blood were treated with LPS, T cell activation beads, or no treatment. Cells were then profiled by 10X scRNA-seq

first: Jiaqi

last: Li

first: Claudia

last: Worth

first: Davide

last: Simone

first: Lihua

last: Duan

first: Paul

last: Bowness

first: Liye

last: Chen


title: Intrarenal single cell sequencing of MPO-ANCA associated glomerulonephritis patients reveal novel targetable treatment options

city: Clayton

first: Peter

middle: James

last: Eggenhuizen

state: VIC

address: Monash Medical Centre, 246 Clayton Road

department: Medicine

country: Australia

institute: Monash University

summary: The etiopathogenesis underlying myeloperoxidase anti-neutrophil cytoplasmic antibody associated glomerulonephritis (MPO-AAGN) remains incompletely understood. Furthermore, there are only limited treatment options and treatment resistance of MPO-AAGN is still a common problem. To identify new targeted treatment options, intrarenal single-cell RNA sequencing (scRNA-seq) was applied to kidney biopsies from MPO-AAGN patients and control health kidney tissues to define the transcriptomic landscape at single-cell resolution. Intrarenal scRNAseq was also applied to a pre-clinical mouse model of MPO-AAGN to show that this model of disease can be used to trial new targeted treatments. NF-κB pathway activation was confirmed in a variety of kidney cells in MPO-AAGN patients. Kidney infiltrating immune cells of MPO-AAGN patients were mainly enriched in inflammatory pathways including TNF signaling, IL-17 signaling and NOD-like receptor signaling. These findings were similar in our pre-clinical mouse model of MPO-AAGN. Furthermore, there was an overexpression of inflammasome related genes (AIM2, IFI16) in MPO-AAGN patients. A dynamic gene expression in glomerular resident cells was observed in MPO-AAGN, including increased expression of several genes, including CD9 and SPARC, which were closely related to parietal epithelial hyperplasia and crescent formation and lesion progression. Importantly, overexpression of HSP90AA1 in non-focal mesangial cells and endothelial cells was found and the expression of several chemokines (CCL20, CXCL3, CXCL8, CXCL1, CCL2) were upregulated in non-focal proximal tubule cells. Moreover, MPO-AAGN patients with treatment resistance had higher proportions of kidney infiltrating classical monocytes and CD8+ T cells. Elevated expression of SPARC and LAMA4 in mesangial cells, IL33 in endothelial cells, and CFL1 in several cell clusters (proximal tubule cells, loop of Helen, macrophages) were observed in MPO-AAGN patients with treatment resistance when compared with patients who achieved remission after induction therapy. These results offer new insight into the pathogenesis of the progression and treatment resistance MPO-AAGN. We have identified new therapeutic targets for MPO-AAGN that can be tested in a pre-clinical model of disease.

overall_design: Single cell suspensions of mouse kidney were prepared from MPO-AAGN and healthy wild-type mice and scSeq was performed.

first: Yong

last: Zhong

first: Wei

last: Lin

first: Elean

middle: S

last: Tay

first: Ting

last: Meng

first: Peng

last: Jin

first: Xiang

last: Ding

middle: J

first: Poh-Yi

last: Gan

first: Rong

last: Tang

first: Joshua

middle: D

last: Ooi

first: Xiangcheng

last: Xiao


title: Comparative analysis of peripheral blood immunoinflammatory landscapes in patients with acute cholangitis and its secondary septic shock using single-cell RNA sequencing

city: Hefei

first: He

middle: Ms.

last: Zhang

address: Jixi Road 218

institute: Anhui medical university

summary: The immune inflammatory responses play an important role in the occurrence and development of septic shock. We performed a detailed transcriptomic analysis of peripheral blood mononuclear cells (PBMCs) to understand the differences of immunocyte subtypes in immunoresponse and inflammation between common infections and septic shock secondary to acute cholangitis (AC). Through the screening and analysis of sequencing data, we found that there were significant differences in cell subtypes and molecular characteristics between AC and septic shock. Furthermore, we demonstrated that proNeus played a crucial role in the immunoinflammation of septic shock. Our study revealed the differentiation trajectory of neutrophils, and explored the mutual effect between main cells. In this regard, it provided a reference for accurately evaluating the pathological severity of patients with septic shock and discovering the targets for therapy.

overall_design: Peripheral blood samples were collected from three patients with acute cholangitis and three patients with septic shock secondary to acute cholangitis. GEXSCOPETM Single Cell RNA Library Kit was used to barcode single cells, capture mRNA and generate cDNA libraries for scRNA-seq. Then pooled for sequencing on a Illumina Nova-seq 6000 platform with 150 bp paired-end reads

last: Wang

first: Yuntian

last: Xu

first: Mingchao

last: Pei

first: Yun

last: Zheng


title: Single-cell RNA-sequencing of tumor-infiltrating cells from intra-tibial injection of LLC1 cells in murine model.

city: hangzhou

first: Zhaoming

last: Ye

address: 88 jiefang road

institute: The Second Affiliated Hospital, Zhejiang University School of Medicine

summary: We injected LLC1 cells into tibia to construct a murine model to study bone microenvironment in cancer bone metastasis. We used single cell RNA sequencing (scRNA-seq) to analyze the cellular heterogeneity of tumor-infiltrating cells.

overall_design: LLC1 cells were injected intra-tibially into CAG-ZsGreen mice. 14 days later tumor-bearing tibia were dissected and enzymatically dissociated into single cell suspension. Tumor-infiltrating cells were isolated by Fluorescence-activated cell sorting (FACS) according to the presence of ZsGreen signal and analyzed using scRNAseq.

first: Hao

last: Zhou

first: Wenkan

first: Junjie

last: Gao


title: Single-cell RNA-sequencing of osteocyte-derived mitochondria recipient cells in the intra-tibial injection of LLC1 cells murine model.

summary: LLC1 cells were injected into the tibia of DMP1-Cre PhAM mice to study the intercellular mitochondrial transfer in the bone microenvironment. We used single-cell RNA sequencing (scRNA-seq) to analyze the cellular heterogeneity of osteocyte-derived mitochondria recipient cells.

overall_design: LLC1 cells were injected intra-tibially into DMP1-Cre PhAM mice. 14 days later tumor-bearing tibia were dissected and enzymatically dissociated into single cell suspension. Osteocyte-derived mitochondria recipient cells were isolated by fluorescence-activated cell sorting (FACS) according to the presence of Dendra2 signal and analyzed using scRNAseq.


title: CDAHFD induced NASH mouse model

city: Shanghai

first: Jiangtao

last: Fu

address: Guohe Road 325

institute: Naval medical university

summary: Non-alcoholic steatohepatitis (NASH) is the most significant cause of chronic liver disease worldwide, with limited therapeutic options. In this experiment, a choline-deficient amino acid-defined high fat diet (CDAHFD)  were used to construct a mouse NASH model. After 16 weeks of CDAHFD  diets, liver samples were collected. We want to further confirm that the elevated EFHD2 is specifically expressed in infiltrated macrophages/monocytes in NASH.

overall_design: Animal experiments were conducted using CDAHFD induced NASH models in C57BL/6J mice.Prepare mouse liver as a single-cell suspension.

first: Pei

first: Yi

last: Liu


title: Nuclear actin tunes ERK activity to promote somatic cell reprogramming into pluripotency

city: New Haven

first: Shangqin

last: Guo

state: CT

address: 10 Amistad ST

department: Cell Biology

institute: Yale University

summary: Yamanaka reprogramming is a stochastic process resulting in only a small fraction of somatic cells successfully converting into iPSCs. The molecular and cellular basis underlying this stochasticity remains elusive. Here we demonstrate that this stochasticity can be eliminated when ERK activity is tuned within a narrow range. This tuning can be accomplished by one tenth the concentration of MEK inhibitor in the 2i media. In the absence of pharmacologic ERK inhibition, cells fine tune ERK by morphological changes, growing taller by allocating more actin into their nucleus. A minimal cell height of 10 m is required for pluripotency, a cell geometric feature underlying the “dome-shaped” colony morphology of naïve pluripotent stem cells. Nuclear actin tunes ERK activity by binding to and immobilizing TFII-I, a transcription factor that binds and mediates ERK’s nuclear activation. This work uncovers a mechanistic link for how cell morphology controls cell identity. Insights into these fundamentally coupled processes provide effective approaches to overcome the stochasticity in reprogramming into pluripotency.

overall_design: We performed bulk RNA-seq of control (EV), Actin overexpressed (Actin)  and NLS-actin OE (NLA-Actin) in mouse MEF and day 4 reporgramming cells. We also performed bulk RNA-seq of green positive or red positive reprogramming cells with or without NLS-actin under day 10 and day 14.

first: Jian

first: Qiao

last: Wu


title: Effect of depletion of Fer on gene expression of BEAS-2B cells

city: Chongqing

first: Wang

last: Jiaqi

address: No. 30, Gaotanyan Street

institute: Army Medical University

summary: To investigate the function Fer in the tumorigenesis, we established BEAS-2B cell lines in which Fer has been knocked down by shRNA. We then performed gene expression profiling analysis using data obtained from RNA-seq of 2 different cells.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for BEAS-2B CTL cells and Fer KD cells.

first: Yan

last: Ruan

first: Jiaqi


title: RediScore: Prospective Validation of a pipeline for Homologous Recombination Deficiency Analysis

city: Athens

first: Georgios

last: Tsaousis

state: Attiki

address: Spaton 52, Ave., Gerakas

department: Bioinformatics

country: Greece

institute: Genekor Medical SA

summary: SNP arrays and appropriate bioinformatic algorithms were used to calculate GIS in 29 Ovarian cancer patients with known GIS status using a validated NGS assay.

overall_design: Affymetrix SNP arrays were performed according to the manufacturer's directions on DNA extracted from patients with ovarian cancer tumor tissue

first: Katerina

last: Tsantikidi

first: Eirini

last: Papadopoulou

middle: N

last: Nasioulas


title: Single-cell RNA-sequencing of intratibial LLC cancer model in DMP1-Cre Mfn2-/- mice, DMP1-Cre Rhot1-/- mice, and wild-type mice.

summary: We injected LLC1 cells into the tibia of DMP1-Cre Mfn2-/- mice, DMP1-Cre Rhot1-/- mice, and wild-type mice to study the changes in the cellular composition of the cancer microenvironment after knockout of Mfn2 or Rhot1 in osteocytes.

overall_design: LLC1 cells were injected intra-tibially into DMP1-Cre Mfn2-/- mice, DMP1-Cre Rhot1-/- mice, and wild-type mice. 14 days later tumor-bearing tibia were dissected and enzymatically dissociated into single cell suspension. Live cells were analyzed using scRNAseq (3 tumors from 3 mice mixed 1:1:1 for each group).

last: Zhang


title: Distinct roles for COMPASS core subunits Set1, Trx, and Trr in the epigenetic regulation of Drosophila heart development

city: Baltimore

first: Hangnoh

middle: Joseph

last: Lee

state: MD

address: 670 W Baltimore St

institute: University of Maryland School of Medicine

summary: This study investigated the role Complex of Proteins Associated with Set1 (COMPASS) in heart development. These are evolutionarily highly conserved multiprotein complexes required for H3K4 methylation. In Drosophila, their core subunits are Set1, Trx, and Trr. We studied flies deficient for either of these three subunits their hearts. These flies showed high lethality at eclosion (emergence of adult flies from their pupal case) and significantly shortened lifespans for the adults that did emerge. Furthermore, their hearts showed reduced H3K4 monomethylation (H3K4me1) and dimethylation (H3K4me2), and significantly altered gene expression patterns. The data revealed that each of the COMPASS core subunits (Set1, Trx, and Trr) control methylation of different sets of genes with distinct temporal activity: Set1 throughout, whereas Trr was active early and Trx at later stages of heart development. We found that many metabolic pathways were active early in development and throughout, while muscle and heart differentiation processes were methylated during later stages of development. Taken together, the findings demonstrate the importance of COMPASS complexes during heart development, therewith supporting their implication in congenital heart diseases.

overall_design:
To investigate the gene expression patterns regulated by Set1, Trx, and Trr. We employed the Drosophila UAS-Gal4 system combined with RNAi knockdown. We used twist (twi)-Gal4 to tissue knockdown Set1, trx, or trr specifically in the Drosophila heart.
We then performed gene expression profiling analysis using data obtained from RNA-Seq of three different samples for each genotype.
Comparative gene expression profiling analysis of RNA-Seq data for wildtype and its knock-down (RNAi) derivatives (Set1, trx, and trr).

first: Junyi

last: Zhu

first: Zhe

last: Han


title: BCAS2 regulates oocyte meiosis by participating in alternative mRNA splicing

city: Beijing

first: xiaohong

last: Yao

address: No. 2 Yuanmingyuan West Road, Haidian District, Beijing

institute: China Agricultural University

summary: Oocyte meiosis is an important factor affecting female reproduction. Breast cancer amplified sequence 2 (BCAS2) is a component of the spliceosome. Previous reports have shown that BCAS2 is critical in male germ cell meiosis, oocyte development, and early embryo genome integrity. However, the role of BCAS2 in oocyte meiosis has not been reported. We used Stra8-GFP-Cre mice to knock out BCAS2 during the pachytene phase of oocytes. The results of fertility tests showed that the cko mice were infertile. Morphological analysis showed that the number of primary follicles of 2M ovary was significantly reduced and follicle development was blocked. Further analysis showed that the number of primordial follicles decreased and follicle development slowed from 7dpp ovaries. Sequencing revealed that DNA damage in oocytes could not be repaired from 5dpp. There was an abnormality in meiosis, some oocytes could not reach the diplotene stage of meiosis, and more oocytes could not develop to the dictyate stage. AS analysis reveals that abnormal variable splicing of Dazl and Diaph2 Oogens-related genes in cKO mice, with involvement of the PRP19/CDC5l complex.

overall_design: BCAS2 was knocked out in ovarian oocytes, and 1dpp ovaries were taken to analyze and compare the gene expression profiles of RNA-Seq data between WT and knockout groups.

first: Xiaohong

first: Jiali


title: Single-cell RNA-sequencing of PBMCs from SAVI patients reveals disease-associated monocytes with elevated integrated stress response

city: Paris

first: Mickael

middle: Mathieu

last: Menager

address: 24 Boulevard du Montparnasse

institute: Institut Imagine

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

overall_design: Refer to individual Series


title: Multi-omics integration identifies progenitor-state specific PBRM1-PIAS1 cooperation in repressing differentiation in epithelial tissue

city: Evanston

first: Xiaomin

last: Bao

state: IL

address: 633 Clark Street

department: Molecular Biosciences

institute: Northwestern University

summary: Here we determine the gene targets controlled by PBRM1-PIAS1 cooperation in regulating epidermal differentiation.

overall_design: RNA-seq for PBRM1 shRNA knockdown and PIAS1 shRNA knockdown was performed on undifferentiated and differentiated keratinocytes against controls. ChIP-seq for PBRM1 and PIAS1 was performed in undifferentiated and differentiated keratinocytes. CUT&RUN for BRD7 was performed in undifferentiated keratinocytes in control versus PBRM1 shRNA knockdown conditions.

first: Patric

last: Ho


title: Dual-active oxadiazoles targeting the Mycobacterium tuberculosis ESX-1 secretion system and boosting ethionamide activity

city: Cologne

first: Raphael

last: Gries

address: Robert-Koch-Straße 21

country: Germany

institute: University Hospital Cologne

summary: To find potential regulatory targets of the anti-virulence compound S3  in Mycobacterium tubercuolsis.

overall_design: We checked for differentially expressed genes in Mycobacterium tuberculosis after 4h exposure to compound S3.

first: Michael

last: Dal Molin

first: Jason

last: Chhen

first: Edeltraud

last: van Gumpel

first: Lindsay

last: Sonnenkalb

first: Christian

last: Utpatel

first: Jan

last: Rybniker


title: Single-cell RNA-sequencing of PBMCs from SAVI patients reveals disease-associated monocytes with elevated integrated stress response II

summary: Gain-of-function mutations in STING1, that codes for the Stimulator of Interferon Gene (STING), result in a severe autoinflammatory disease termed STING-associated vasculopathy with onset in infancy (SAVI). Although elevated type I interferon (IFN) production is thought to be the leading cause of the symptoms observed in patients, STING can induce a set of pathways, and their role in the onset and severity of SAVI remains to be elucidated. To address this point, we compared a single-cell RNA sequencing (scRNA-seq) dataset of peripheral blood mononuclear cells (PBMCs) from SAVI patients to a dataset of healthy PBMCs treated with recombinant IFN-β. We revealed a loss of mucosal associated invariant T cells and CD56bright natural killer cells in SAVI patients, not replicated in IFN-β-treated PBMC. Patient T cells are in an activated state associated with senescence and apoptosis, dependent on type I IFNs. Inferring cell to cell communication, from scRNA-seq predicted monocytes as potential drivers of this T cell phenotype and was supported by plasma cytokines measurement, with high CCL3, CCL4 and IL-6. Furthermore, scRNA-seq clustering identified a patient-specific subset of monocytes, highly inflammatory and expressing a strong integrated stress response (ISR). It also pinpointed to a patient with lower ISR, allowing us to identify a secondary mutation in PERK, that was recently shown to be activated by STING to trigger the ISR. Finally, based on the identification of this patient-specific subset of monocytes and the exploration of IFN-β stimulated PBMCs from healthy donors, we developed a strategy to propose a transcriptomic signature specific of STING activation and independent of type I IFN. Altogether, these results provide a deeper understanding of SAVI at the cellular and molecular levels.

overall_design: 7 CTRL, 5 untreated SAVI patients, and 4 SAVI patients treated with a JAK-inhibitor (including 2 samples for the first patient)

first: Camille

last: de Cevins

first: Laure

last: Delage

first: Maxime

last: Batignes

first: Marine

last: Luka

first: Anne

last: Remaury

first: Tinhinane

last: Fali

first: Cecile

last: Masson

first: Bénédicte

last: Hoareau

first: Catherine

last: Meunier

first: Melanie

last: Parisot

first: Mohammed

last: Zarhrate

first: Quentin

last: Riller

first: Brieuc

last: Pérot

first: Victor

last: Garcia-Paredes

first: Francesco

last: Carbone

first: Luc

last: Canard

first: Charlotte

last: Boussard

first: Etienne

last: Crickx

first: Jean-Claude

last: Guillemot

first: Marie-Louise

last: Frémond

last: Neven

first: Galina

last: Boldina

first: Franck

last: Augé

first: Alain

last: Fischer

first: Michel

last: Dider

first: Fredéric

last: Rieux-Laucat

first: Mickaël

last: Ménager


title: Stabilisation of b-Catenin-WNT signalling by USP10 in APC-truncated colorectal cancer drives cancer stemness and enables super-competitor signalling

city: Berlin

first: Michaela

last: Reissland

address: Ihnestr. 63

department: Genome Regulation

institute: Max Planck Institut for molecular genetics

summary: The contribution of deubiquitylating enzymes to b-Catenin stabilisation in intestinal stem cells and colorectal cancer (CRC) is poorly understood. Here, we report the deubiquitylase USP10 as APC-truncation- specific regulator enhancer of b-Catenin stability, potentiating WNT signalling pathway in CRC and cancer stem cells. Mechanistically, interaction studies in various CRC cell lines and in vitro binding studies, together with computational modelling, revealed that USP10 binding to b-Catenin is mediated via its USP10 unstructured N-terminus and requires truncated in the absence of full-length APC. Notably, loss of USP10 in CRISPR engineered intestinal organoids reduces tumorigenic properties of CRC cells and organoids, and blocks the super competitor-properties of APC-mutated intestinal cells, elicits induces the expression of differentiation genes, and opposes the APC-truncated phenotype in an intestinal hyperplasia model of D.melanogaster. Taken together, our findings reveal USP10s role in intestinal tumourigenesis by stabilising b-Catenin, leading to aberrant WNT signalling, enhancing cancer cell stemness and implicate the DUB USP10 as a cancer specific therapeutic vulnerability in Apc truncated CRC.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for APC:p53:Kras shNTC and shUsp10-1,-2 organoids.

first: Oliver

last: Hartmann

first: Saskia

last: Tauch

first: Cristian

last: Prieto-Garcia

first: Clemens

last: Schulte

first: Daniel

last: Solvie

first: Sinah

last: Loebbert

first: Anne-Claire

last: Jacomin

first: Marina

last: Pesic

first: Jeroen

last: Bugte

first: Christina

last: Schuelein-Voelk

first: Carmina

middle: T

last: Fuss

first: Nikolet

last: Pahor

first: Carsten

last: Ade

first: Victoria

last: Buck

first: Michael

last: Potente

first: Vivian

last: Li

first: Gerti

last: Beliu

first: Armin

last: Wiegering

first: Eliya

last: Bitman-Lotan

first: Tom

last: Grossman

first: Mathias

last: Rosenfeldt

first: Martin

last: Eilers

first: Hans

last: Maric

first: Maurice

last: Madelon

first: Florian

last: Greten

first: Ivan

last: Dikič

first: Amir

last: Orian

first: Peter

last: Gallant

first: Markus

middle: E

last: Diefenbacher


title: Environmentally relevant concentrations of niclosamide disrupt antioxidant defense, histology, and the liver and intestine transcriptome of Chinese soft-shelled turtle

city: Changsha

first: Hao

last: Wu

address: Shuanghe Road 728, Kaifu District, Changsha

institute: Hunan Fisheries Science Institute

summary: To investigate the cellular responses of the Chinese soft-shelled turtle after niclosamide exposure, a total of 45 healthy juvenile turtles randomly selected and divided into three groups containing one control group (without NIC exposure, CG) and two experimental groups (exposed to 10 and 50 μg/L of NIC, respectively).

overall_design: The expression profiling analysis of turtle liver and gut for after niclosmaide exposure


title: Vitamin B12 alleviates myocardial ischemia/reperfusion injury

first: Li

last: Guoxing

state: Chongqing

address: Street of shiyou Road, Yuzhong District, Chongqing, China

institute: Chongqing Medical University

summary: To examine the protective effect of B12 on myocardial ischemia/reperfusion injury and figure out the mechanism of B12.

overall_design: The mice were separated, on random, into 4 groups: Sham, Sham+B12, ischemia/reperfusion model (I/R), I/R+B12. B12 treatment groups at dose of 50 mg/L via drinking water 7 days before I/R procedure. The per day water intake of mice was 4mL. Mice were anesthetized with 2% isoflurane and placed in a supine position on a heating pad (37 ℃) for left thoracotomy under mechanical ventilation. To induce I/R injury, the left anterior descending coronary artery (LAD) of the mice was occluded and then released after 45 min of ischemia, followed by 24 h reperfusion. Sham-operated animals underwent the same procedure without LAD occlusion.After heart specimens were harvested, high-throughput sequencing was then performed.

first: Zhe

last: Cheng

first: Q

last: Yu


title: Single-cell RNA-sequencing of PBMCs from SAVI patients reveals disease-associated monocytes with elevated integrated stress response I

first: Mickael

last: Menager

overall_design: scRNA-seq on PBMCs (from healthy donors) treated with IFN-β at one of several timepoints (0h, 1h, 2h, 4h, 8h, 12h, 24h, 36h)

first: Camille

last: de Cevins

first: Laure

last: Delage

first: Maxime

last: Batignes

first: Marine

last: Luka

first: Anne

last: Remaury

first: Tinhinane

last: Fali

first: Cecile

last: Masson

first: Bénédicte

last: Hoareau

first: Catherine

last: Meunier

first: Melanie

last: Parisot

first: Mohammed

last: Zarhrate

first: Quentin

last: Riller

first: Brieuc

last: Pérot

first: Victor

last: Garcia-Paredes

first: Francesco

last: Carbone

first: Luc

last: Canard

first: Charlotte

last: Boussard

first: Etienne

last: Crickx

first: Jean-Claude

last: Guillemot

first: Marie-Louise

last: Frémond

last: Neven

first: Galina

last: Boldina

first: Franck

last: Augé

first: Alain

last: Fischer

first: Michel

last: Dider

first: Fredéric

last: Rieux-Laucat

first: Mickaël

middle: M

last: Ménager


title: Deregulation of neuronal polarization promotes axonal growth after spinal cord injury (ATAC-Seq)

city: London

first: Franziska

last: Müller

address: Du Cane Road

department: Brain Sciences

institute: Imperial College London

summary: The developmental maturation of a neuron requires the completion of neuronal polarization preceding the formation of a synapse. Whether neuronal polarization marks the decline in growth competence in the injured mammalian adult nervous system remains elusive. Here we show that gene expression and epigenetic signatures associated with the regenerative growth ability of dorsal root ganglia (DRG) sensory neurons are lost during the transition from a non-polarized to a polarized state. The transcriptional co-factor Cited2 was found to be epigenetically upregulated in immature DRG and following a regenerative injury, but it remained unchanged by a non-regenerative spinal cord injury (SCI). Next, Cited2 was overexpressed in DRG neurons in a model of SCI in mice where it promoted sensory axon growth and reversed the gene expression signatures associated with neuronal maturation. Importantly, Cited2 expression stirred the maturation of DRG neurons towards a non-polarized state. Together, these data suggest that the transition from a non-polarized to a polarized state marks the reversible loss of the regenerative ability of a neuron, thus paving the way to targeted repair strategies relying on neuronal de-maturation.

overall_design: AAV-GFP or AAV-Cited2 was injected bilaterally into the sciatic nerve 4 weeks prior to a dorsal column axotomy. 24 hours post-injury, DRG were extracted and a neruonal enriched population processed for ATAC-seq

first: Matt

last: Danzi

first: Ilaria

last: Palmisano

first: Siimone

last: Di Giovanni


title: Phenotypic plasticity for improved light harvesting, in tandem with methylome repatterning in reef-building corals

city: University Park

first: KELLY

middle: Johanna

last: GOMEZ-CAMPO

state: Pennsylvania

address: 212 Mueller Laboratory

department: Biology

institute: PSU

type: Methylation profiling by high throughput sequencing

summary:
Acclimatization through phenotypic plasticity represents a more rapid response to environmental change than adaptation and is vital to optimize organisms’ performance in different conditions. Generally, animals are less phenotypically plastic than plants, but reef-building corals exhibit plant-like properties. They are light-dependent with a sessile and moddular construction that facilitates rapid morphological changes within their lifetime. We induced phenotypic changes by altering light exposure in a reciprocal transplant experiment and found that coral plasticity is a colony trait emerging from comprehensive morphological and physiological changes within the colony. Plasticity in skeletal features optimized coral light harvesting and utilization and paralleled with significant methylome and transcriptome modifications. Network-associated responses resulted in the identification of hub genes and clusters associated to the change in phenotype: inter-partner recognition and phagocytosis, soft tissue growth and biomineralization. Furthermore, we identified hub genes putatively involved in animal photoreception-phototransduction. These findings fundamentally advance our understanding of how reef-building corals repattern the methylome and adjust a phenotype, revealing an important role of light sensing by the coral animal to optimize photosynthetic performance of the symbionts.
This SuperSeries is composed of the SubSeries listed below.


title: Bulk RNAseq data for bladder cancer specimens [UC cohort]

city: New York

first: Li

last: Wang

address: 1 Gustave L. Levy Pl

institute: Icahn school of medicine at Mount Sinai

summary:
When applying deconvolution methods to bulk RNAseq data, a limitation of most prior studies is the lack of paired scRNA-seq and bulk RNA-seq data from the same samples to serve as ground truth for deconvolution. Our UC cohort, containing matched scRNA-seq and bulk RNA-seq data therefore provided a unique opportunity. (The scRNAseq datasets have been published before [PMID: 32111252 PMID: 36129800 PMID: 36099881 PMID: 33837006 ]. Please see linked manuscript for details)
We assembled a scRNA-seq dataset of 100,667 cells from 30 UC tissue samples (20 unique patients). Bulk RNA sequencing was performed on a subset of patients(14) in the single-cell RNA sequencing cohort due to tissue availability.

overall_design: Bulk RNAseq data for bladder cancer specimens from 14 patients.

first: Sudeh

last: Izadmehr


title: Cyclin B1 deficiency in renal tubular epithelial cells promotes kidney ageing and fibrosis in mice

city: Zhanjiang

first: Ji

middle: Xin

last: Tang

address: No.57 South Renmin Avenue, Xiashan District

department: Affiliated Hospital of Guangdong Medical University, Zhanjiang, Guangdong, China.

institute: Mouse kidney

summary: After severe renal injury, renal tubular epithelial cells (TECs) will be arrested in G2/M phase, and the arrested cells will be aging, showing senescence related secretory phenotype (SASP), which mediates renal fibrosis by secreting fibrogenic cytokines. Cyclin B1 is an important protein that promotes G2/M phase transition of cell cycle. Its role in renal fibrosis mediated by G2/M arrest of TECs is unknown. The aim of this study was to investigate the role of cyclin B1 in renal fibrosis induced by TECs G2/M arrest and its mechanism

overall_design: We compared the deletion of cyclin B1 in mouse renal tubular epithelial cells with that in the control group by sequencing genes to explore the role of cyclin B1 in renal fibrosis caused by G2/M cycle arrest of epithelial cells.

first: Yun-Shan

last: Wu

first: Shan

last: Liang

first: Dong-Yi

last: Li

first: Ze-Sen

last: Feng

first: Jun-Hao

last: Wen

first: Dan

first: Chen

last: Yang

first: Ji-Xin

first: Hua-Feng

last: Liu


title: The Hsf1-sHsp cascade has pan-antiviral activity in mosquitoes

city: Nijmegen

first: Jieqiong

last: Qu

address: Geert Grooteplein Zuid 28

department: Department of Medical Microbiology

country: Netherlands

institute: Radboud Institute for Molecular Life Sciences, Radboud University Medical Center

summary: Aedes mosquitoes transmit pathogenic arthropod-borne (arbo) viruses, putting nearly half the world’s population at risk. Blocking virus replication in mosquitoes rather than in humans serves as a promising approach to prevent arbovirus transmission, which requires in-depth knowledge of mosquito immunity. By integrating multi-omics data, we identified that heat shock factor 1 (Hsf1) regulates eight small heat shock protein (sHsp) genes within one topological associated domain. This Hsf1-sHsp cascade acts as an early response against chikungunya virus (CHIKV) infection and shows pan-antiviral activity in three vector mosquitoes, Aedes aegypti, Aedes albopictus, and Anopheles gambiae. We then assessed the baseline expression of sHsp genes in different tissues of female Ae. aegypti using RNA-seq, and we observed a highly dynamic expression pattern of sHsp genes that varied dramatically across different tissues. Interestingly, sHsp genes were expressed at low levels in two main barrier tissues, the midgut and salivary glands, compared to other tissues such as the crop. Importantly, activation of Hsf1 led to a reduced CHIKV infection rate in adult Ae. aegypti mosquitoes, demonstrating Hsf1 as a promising target for the development of novel intervention strategies to limit arbovirus transmission by mosquitoes.

overall_design: We assessed the baseline expression of sHsp genes in different tissues of female Ae. aegypti using RNA-seq. The tested tissues include crop, fat body, leg, malpighian tubules, midgut, ovary, salivary glands, and wings.

first: Ronald

last: van Rij


title: Differential gene expression in the posterior joint capsule for immobility in mice

city: Fukuoka

first: Hirotaka

last: Iura

address: 3-1-1 Maedashi

department: Orthopedic Surgery

country: Japan

institute: Kyushu University

summary: To explore the factors involved in arthrofibrosis, we performed a genome-wide screening for mRNA expressed in the posterior capsule after immobilization in mice. A gene ontology term analysis for differentially expressed genes after 2 weeks immobilization indicated that the genes involved in the extracellular region were particularly enriched.  A heatmap demonstrated that joint immobilization significantly altered the expressions of genes related to the extracellular region. The MA plot indicated that, compared to naïve mice, 1654 genes were upregulated more than 2-fold, and 985 genes were downregulated more than 2-fold in immobilized mice. The MA plot and heatmap visually revealed dynamic changes in the expression of genes in immobilized posterior capsules compared to those in naïve posterior capsules.   Our study represents the first detailed analysis of mouse joint capsule transcriptomes, with biologic replicates, generated by RNA-seq technology. RNA-seq based transcriptome characterization would expedite genetic network analyses and clarify Unresolved pathophysiology of joint contractures.

overall_design: mRNA profiles of posterior joint capsule in 8-week-old wild-type (WT) mice immobilized in casting for 2 weeks.

first: Kazu

last: Kobayakawa

first: Seiji

last: Okada


title: Targeting GPX4-mediated ferroptosis alleviates liver injury in a rat model of intestinal failure-associated liver disease

city: Shanghai

first: Siyang

last: Cheng

address: KongJiang Rd.1665

institute: Xinhua hospital

type: Non-coding RNA profiling by high throughput sequencing

summary: Background: Intestinal failure-associated liver disease (IFALD) is a common complication of long-term parenteral nutrition (PN) that is associated with significant morbidity and mortality. Ferroptosis, as an iron-dependent regulated cell death, has been shown to play an important role in the development of several liver diseases. This study focuses on investigating whether the ferroptosis phenomenon is present in TPN-induced IFALD and further exploring the potential regulatory mechanisms.    Methods: Ferroptosis hallmarkers were measured in children with short bowel syndrome (SBS) who had long-term PN use and caused IFALD. Sprague-Dawley (SD) rats were used to establish a rat model of IFALD with a concomitant ferroptosis inhibition intervention using liproxstain-1. The mir-431 lineage was identified as a potential upstream regulatory mir-RNA by mir-RNA sequencing. HepG2 and 293T cell line was used to demonstrate that mir-431 regulates ferroptosis through the GPX4 pathway at the cellular level in vitro.    Results: Ferroptosis is upregulated in liver of children with IFALD. Liproxstain-1 downregulates ferroptosis in a rat model of IFALD and attenuates hepatic steatosis through the lipid metabolism pathway. In vitro HepG2 and 293T cell experiments reveal that mir-431 affects ferroptosis by regulating GPX4 protein.    Conclusions: Ferroptosis plays an important role in the development of IFALD. Liproxstain-1 inhibits ferroptosis and attenuates hepatic steatosis in a rat model of IFALD through the lipid metabolism pathway. Mir-431 negatively regulates GPX4 protein-induced ferroptosis.

overall_design: Comparative gene expression profiling analysis of MicroRNA-seq data for TPN rats versus Control rats

first: Ying

first: Lu

last: Jiang

first: Wei

last: Cai


title: Therapeutic Blocking of VEGF Binding to Neuropilin-2 Diminishes PD-L1 Expression to Activate Anti-Tumor Immunity in Prostate Cancer

city: Worcester

first: Rui

state: Massachusetts

address: 364 Plantation Street, LRB

department: MCCB

institute: University of Massachusetts Medical School

summary: Prostate cancers (PC) are largely unresponsive to immune checkpoint inhibitors and there is strong evidence that PD-L1 expression itself must be inhibited to activate anti-tumor immunity. Here, we report that neuropilin-2 (NRP2), which functions as a VEGF receptor on tumor cells, is an attractive target to activate anti-tumor immunity in prostate cancer because we demonstrate that VEGF/NRP2 signaling sustains PD-L1 expression. NRP2 depletion increased T cell activation in vitro. Inhibition of the binding of VEGF to NRP2 using a mouse specific anti-NRP2 mAb in a syngeneic model of prostate cancer that is resistant to checkpoint inhibition resulted in significant necrosis and tumor regression compared to both an anti-PD-L1 mAb and control IgG. This therapy also decreased tumor PD-L1 expression and increased immune cell infiltration. We also observed that the NRP2, VEGF-A, and VEGF-C genes are amplified in metastatic castration resistant and neuroendocrine prostate cancer (NEPC) and that NRP2high PD-L1high population in metastatic tumors had a significantly lower AR and higher NEPC scores than other populations. Therapeutic inhibition of VEGF binding to human NRP2 with a high affinity humanized mAb, which is suitable for clinical use, in organoids derived from NEPC patients also diminished PD-L1 expression and caused a significant increase in immune-mediated tumor cell killing consistent with the animal studies. These findings provide justification for the initiation of clinical trials using this novel function-blocking NRP2 mAb in prostate cancer, especially for patients with aggressive primary and metastatic cancers, where blocking NRP2-VEGF signaling shows potential in mitigating the morbidity and mortality associated with these cancers.

overall_design: PC3 cells were sorted into 2 populations: NRP2-high and NRP2-low and analyzed for RNA-seq, in triplicate. Total 6 samples.

first: Mengdie

last: Wang

first: Lihua

last: Zhu

first: Arthur

last: Mercurio


title: Oxidative Stress-initiated One-carbon Metabolism drives Interleukin-10-producing B Cells to Resolute Pneumonia (RNA-Seq)

city: shanghai

last: Fu

address: Dongan road

institute: Fudan university

summary: Metabolic reprogramming for regulatory B cell generation in infectious diseases remains unknown. With a Pseudomonas aeruginosa-induced pneumonia model, we report IL-10-producing B cells (IL-10+B cells) are indispensable for spontaneous remission of the infection. The accumulation of cytosolic reactive oxygen species (ROS) is responsible for the IL-10 producing within B cells. Of note, ROS scavenging downregulated the one-carbon metabolism and the depletion of one-carbon metabolic methyl transfer enzyme serine hydroxymethyl transferase 1 (Shmt1) led to decreased IL-10+ B cells production both in vitro and in vivo. Mechanistically, one-carbon flux enhanced the availability of methyl groups, which altered histone H3 lysine 4 methylation at Il10 loci and led to massive IL-10 production in B cells. Therefore, the metabolic-associated drug ethacrynic acid (EA) was screened out and potentially restored infectious pneumonia accompanied by increased IL-10+ B cells. The results provide a new insight that ROS serve as modulators to resolute inflammation by reprogramming one-carbon metabolism in lung B cells.

overall_design: To screen for alterations involved in ROS-dependent IL-10+B cells, the RNA sequencing analyses were carried out. Splenic B cells were isolated and cultured under correspondent stimulations after 24h (αIgM, αIgM+LPS, αIgM+LPS+NAC, n=3) and the unstimulated group were B cells without any stimulation after isolated (n=3). Total RNA was extracted using RNAsimple Total RNA Kit (TIANGEN, China). RNA sequencing was performed by the Beijing Genomics Institute (BGI, Shenzhen, China).

first: Baichao

last: Yu

first: Qi

first: Zhou

last: Lu

first: Hushan

last: Zhang

first: Dan

first: Feifei

last: Luo

first: Ronghua

first: Luman

first: Yiwei

last: Chu


title: CELLULAR SENESCENCE CONTRIBUTES TO THE PROGRESSION OF HYPEROXIC BRONCHOPULMONARY DYSPLASIA

city: Milwaukee

first: Martin

last: Hessner

address: 8701 Watertown Plank Road

department: Pediatrics

institute: Medical College of Wisconsin

summary:
Oxidative stress (OS), inflammation, and endoplasmic reticulum (ER) stress sequentially occur in the rat model of hyperoxia (HOX) induced bronchopulmonary dysplasia (BPD), and they all increase DNA damage. Tumor suppressors increase after DNA damage, followed by apoptosis or cellular senescence when the damage becomes irreparable. Although cellular senescence contributes to wound healing, its persistence will inhibit growth potential. Therefore, we hypothesized that the persistence of cellular senescence plays a role in BPD progression. We detected evidence of increased cellular senescence in rat and human BPD lungs. Foxo4-p53 binding was increased in BPD rat lungs, and inhibition of this binding attenuated BPD severity, indicating that such binding contributes to BPD's cellular senescence. Treatment with tauroursodeoxycholic acid (TUDCA) decreases ER stress. N-Acetyl-lysyltyrosylcysteine-amide (KYC) reduced toxic oxidant production by myeloperoxidase (MPO) and subsequent OS and inflammation. Both agents effectively decreased cellular senescence. Concomitantly, alveolar complexity and the number of type 2 alveolar cells increased, indicating that MPO-mediated OS and ER stress preceded cellular senescence in BPD rat pup lungs. These data suggest that cellular senescence plays an essential role in BPD progression. Reducing MPO toxic oxidant production, ER stress, and attenuating cellular senescence are potential therapeutic strategies for halting BPD progression.
Using multiomic data to investigate the role of cellular senescence in the development of BPD in rat model.

overall_design: Nursing dams were cared for in a 12-h dark-light cycle and had free access to chow and water. According to the study design, two or four pregnant rats were used for each experiment. The dam and pups were cared for in either >90% oxygen chamber (HOX) or room air (NOX) from postnatal day 1 to day 10 (P1-P10). An oxygen sensor continuously monitored oxygen concentrations (Reming Bioinstruments Co., Redfield, NY). Pups were then recovered in room air between P11-P21. Pups were caged with nursing dams while the dams were alternated between oxygen environments to diminish oxygen toxicity. Lungs obtained at P21 from both NOX and HOX groups were used for transcriptomic studies using Affymetrix chips, histology, and immunoblots.

first: Xigang

last: Jing

first: Shuna

last: Jia

first: Maggie

last: Teng

first: Billy

last: Day

first: Adeleye

last: Afolayan

first: Jason

middle: A

last: Jarzembowski

first: Kirkwood

last: Pritchard

first: Stephen

last: Naylor

first: G

middle: G

last: Konduri

first: Ru-Jeng


title: Targeting of the CD161 Inhibitory Receptor Enhances T cell-mediated Immunity Against Hematological Malignancies

status: Public on Jan 02 2024

city: Boston

first: Kai

middle: W

last: Wucherpfennig

state: MA

address: One Jimmy Fund Way

institute: Dana-Farber Cancer Institute

summary: The CD161 inhibitory receptor is highly upregulated by tumor-infiltrating T cells in a number of human solid tumor types, and its ligand CLEC2D is expressed by both tumor cells and infiltrating myeloid cells.  Here we assessed the role of the CD161 receptor in hematological malignancies.   Systematic analysis of CLEC2D expression using the Cancer Cell Line Encyclopedia (CCLE) revealed that CLEC2D mRNA was most abundant in hematological malignancies, including B cell and T cell lymphomas as well as lymphocytic and myelogenous leukemias.  CLEC2D protein was detected by flow cytometry on a panel of cell lines representing a diverse set of hematological malignancies.  We therefore used yeast display to generate a panel of high-affinity, fully human CD161 monoclonal antibodies (mAbs) that blocked CLEC2D binding.  These mAbs were specific for CD161 and had a similar affinity for human and non-human primate CD161, a property relevant for clinical translation.  A high-affinity CD161 mAb enhanced key aspects of T cell function, including cytotoxicity, cytokine production and proliferation, against cell lines originating from patients with AML, DLBCL and Burkitt lymphoma.  In humanized mouse models, this CD161 mAb enhanced T cell-mediated immunity resulting in a significant survival benefit.  ScRNA-seq data demonstrated that CD161 mAb treatment enhanced expression of cytotoxicity genes by CD4 T cells as well as a tissue-resident memory program by CD4 and CD8 T cells that is associated with favorable survival outcomes in multiple human cancer types.  These fully human monoclonal antibodies thus represent potential immunotherapy agents for hematological malignancies.

overall_design: NSG mice were injected I.V. with NY-ESO-1+ Raji cells, followed by I.V. injection of NY-ESO-1 TCR+ CD161+ cells. Mice were treated with either isotype control or CD161 blocking antibody (n=5 per treatment group). On day 13, cells were isolated from both hind limbs and FACS sorted to isolate live CD3+ T cells (human CD45+ human CD19neg, ZsGreenneg, mouse CD45neg).  Cells were analyzed by scRNA-seq using the 10X Genomics platform.

first: Francesca

last: Alvarez Calderon

first: Sasha

last: Kyrysyuk

first: Byong

last: Kang

first: Shiwei

last: Zheng

first: Nathan

last: Matthewson

last: Pyrdol

first: Guo-Cheng

last: Yuan

first: K

last: Wittrup


title: Donor regulatory T cells rapidly adapt to recipient tissues to control acute graft-versus-host disease [TCR-seq]

city: Regensburg

first: Michael

last: Rehli

address: F.-J.-Strauss-Allee 11

department: Internal Med III

institute: University Hospital Regensburg

summary: Adoptive transfer of donor regulatory T cells (Treg) is a promising treatment option for Graft-versus-Host disease (GvHD), but has not yet found its way into routine clinical practice. To map distinctive properties of protective Treg (generated by either polyclonal or allogeneic in vitro expansion), we followed their fate in recipient organs (spleen, liver, colon) in a prophylactic mouse model of MHC-mismatched bone-marrow transplantation (BMT). Using comprehensive gene expression and T cell receptor profiling, we show that both in vitro expansion protocols generated Treg products that preserved hallmark Treg properties,  ameliorated GvHD symptoms, retained their phenotypic plasticity and rapidly acquired organ-specific gene expression profiles after BMT, comparable to their tissue-resident counterparts. When co-transplanted with GvHD-inducing T cells, Treg enabled hallmark suppressive and cytotoxic features, most evidently in the colon. Dominant Treg T cell receptor clonotypes were evenly distributed between organs and across recipients, suggesting a major role of ubiquitous alloantigen-specific Treg in controlling GvHD. Effective protection inversely correlated with the relative abundance of organ-specific Treg, that were transcriptionally distinct, less “activated” and preferentially accumulated in the colon of recipients receiving polyclonally expanded Treg. In summary, we provide a detailed atlas of Treg selection and adaptation in the prophylactic therapy of GvHD.

overall_design: Bulk TCR repertoire data of TRA and TRB mRNAs, generated separately in technical replicates, of independent isolations of C57BL/6 Treg from spleen (fresh, d0, homeostatic state),liver and colon (homeostatic state), in vitro expanded Treg (allo, poly, d11-14), or re-isolated Treg (spleen, liver, colon) 7d post transplant in either BMT control or GvHD models

first: David

last: Dittmar


title: RNA sequencing of necrotic core and non-necrotic rim of tumors from 4T1 cell xenografts into rat

city: Seattle

first: Kevin

last: Cheung

state: WA

address: 1100 Fairview Ave N

department: Public Health Sciences

institute: Fred Hutchinson Cancer Center

summary: The goal of this experiment was to determine differences in RNA expression between the central necrotic tumor core and the non-necrotic rim region in orthotopic xenograft models of 4T1 cells transplanted into immunocompromised rat hosts. These samples are derived from tissues obtained from 4T1 spheroids transplanted orthotopically into the mammary fat pads of SRG OncoRats. 4T1 cells were cultured as spheroids in suspension for 24-hours prior to transplantation in 2% matrigel. 600,000 cells in a 1:1 mixture of matrigel and DMEM/F12 were transplanted orthotopically into the right T4 fat pads of SRG rats. At day 27 post transplanation, primary tumors and lungs were collected. The primary tumors were cut in half to expose the cross-section, and the tumors were dissected to isolate the necrotic core region and the non-necrotic rim region. Samples were snap frozen and sent to Genewiz-Azenta for RNA extraction, library prep, and next-gen sequencing.

sample_organism:
Mus musculus
Rattus norvegicus

platform_organism:
Mus musculus
Rattus norvegicus

overall_design: 4T1 cells were orthotopically transplanted into mammary fat pads of SRG hosts. Tumor and lungs were collected. Tumors were dissected into necrotic and non-necrotic region, and submitted separately for sequencing RNA sequencing. Rat mammary glands (no transplant) and 4T1 cell pellets were sequences as mono-species rat and mouse controls to evaluate the species disambiguation algorithm.

first: Ami

last: Yamamoto

first: Yin

last: Huang

first: Brad

last: Krajina


title: Donor regulatory T cells rapidly adapt to recipient tissues to control acute graft-versus-host disease

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

overall_design: Refer to individual Series


title: Nucleophosmin 1 regulates H3K79me2 levels and peri-nucleolar heterochromatin organization through interaction with the methyltransferase DOT1L [RNA-seq]

city: Freiburg

first: Tanja

last: Vogel

address: Albertstr. 23

institute: University of Freiburg

summary: The histone methyltransferase DOT1L catalyzes methylation of H3K79 and it is highly conserved in mammals. DOT1L plays a functional role in several biological processes including cell cycle regulation, DNA repair, RNA splicing and gene expression, suggesting a complex role in chromatin organization and regulation. Such a remarkable range of functions performed by DOT1L can be the result, at least partially, of its interaction with a plethora of proteins and presence in different complexes. Here, we characterized the cooperation of DOT1L with the nucleolar protein NPM1 and the impact of both proteins on peri-nucleolar heterochromatin activity. We show that i) DOT1L interacts preferentially with monomeric NPM1 in the nucleus; ii) DOT1L acts in concert with NPM1 to maintain each other’s protein homeostasis; iii) NPM1 depletion results in H3K79me2 upregulation at chromatin remodeling genes and transcriptional activation of Ezh2; iv) DOT1L and NPM1 preserved DNA satellite expression at peri-nucleolar heterochromatin via epigenetic mechanisms dependent on H3K27me3. Our findings give insights into molecular mechanisms employed by DOT1L and NPM1 to regulate heterochromatin activities around the nucleoli and shed light on one aspect of the complex role of both proteins in chromatin dynamics.

overall_design: To survey if changes in H3K79me2 levels impact the transcriptome of N2a cells upon reduced levels of NPM1, we generated long-read RNAseq data from N2a cells in control and NPM1 KD conditions using three biological replicates.

first: Annalisa

last: Izzo

first: Ipek

last: Akol

first: Alejandro

last: Villareal

first: Marta

last: Garcia-Miralles

first: Patrick

last: Bovio

first: Stefanie

last: Heidrich


title: Donor regulatory T cells rapidly adapt to recipient tissues to control acute graft-versus-host disease [bulk RNA-seq]

overall_design: total RNA expression profiles of independent isolations of C57BL/6 Treg from spleen (fresh, d0, homeostatic state),liver and colon (homeostatic state), in vitro expanded Treg (allo, poly, d11-14), or re-isolated Treg (spleen, liver, colon) 7d post transplant in either BMT control or GvHD models


title: Effect of depletion of NUDT21 on gene expression in HCT116

city: SHANGHAI

first: zhenyu

last: huang

state: SHANGHAI

address: No.1665 kongjiang RD. Yangpu District

institute: xinhua hospital affiliated to shanghai jiaotong university school of medcine

summary:
To investigate the  function of NUDT21 in the regulation of progression of colorectal cancer, we established HCT116 cell line in which each target gene has been knocked down by shRNA.
We then performed gene expression profiling analysis using data obtained from RNA-seq of 4 different cells at two time points.

overall_design: Comparative gene expression profiling analysis of RNA-seq data for HCT116 cells and its KD derivatives (shScramble , shNUDT21).

first: Xiaojian

last: Chen

first: Yun

last: Liu


title: Donor regulatory T cells rapidly adapt to recipient tissues to control acute graft-versus-host disease [scRNA-seq & scTCR-seq]

overall_design: Donor Treg were reisolated from recipient mice in a GvHD model and sequenced (singleCell RNASeq, TCRSeq)

first: Franziska

last: Pielmeier

first: Hanna

last: Stanewsky

first: Nicholas

last: Strieder


title: de novo methylation by Clr4

city: Charlestown

first: Richard

last: Joh

department: Center for Cancer Research

institute: Mass General Hospital

summary: To investigate the role of de novo methylation by Clr4, we performed the reintroduction of Clr4 and ecopic insertion of lnc230

overall_design: Flag-Clr4 ChIPseq on S pombe.

first: Jasbeer

last: Khanduja

first: Mo

last: Motamedi


title: Nucleophosmin 1 regulates H3K79me2 levels and peri-nucleolar heterochromatin organization through interaction with the methyltransferase DOT1L [ChIP-seq]

summary: The histone methyltransferase DOT1L catalyzes methylation of H3K79 and it is highly conserved in mammals. DOT1L plays a functional role in several biological processes including cell cycle regulation, DNA repair, RNA splicing and gene expression, suggesting a complex role in chromatin organization and regulation. Such a remarkable range of functions performed by DOT1l can be the result, at least partially, of its interaction with different proteins and presence in different complexes. Here, we characterized the interaction of DOT1L with the nucleolar protein NPM1 and the impact of the DOT1L/NPM1 complex on nucleoli organization and activity. We show that i) DOT1L interacts preferentially with monomeric NPM1 in the nucleus; ii) DOT1L interaction with NPM1 is key to maintain their protein homeostasis; iii) NPM1 depletion results in H3K79me2 upregulation at chromatin remodeling genes; iv) DOT1L/NPM1 complex preserved peri-nucleolar heterochromatin organization via epigenetic mechanisms. Our findings unravel the molecular mechanisms employed by the DOT1L/NPM1 complex to maintain heterochromatin organization around the nucleoli and shed light on one aspect of the complex role of DOT1L in chromatin dynamics.

overall_design: H3K79me2 ChIP-seq of N2A cells treated with control or Npm1 shRNA.


title: Nucleophosmin 1 regulates H3K79me2 levels and peri-nucleolar heterochromatin organization through interaction with the methyltransferase DOT1L


title: A549 cigarette smoke resistant cell line expression profiling

city: Giessen

first: Jochen

last: Wilhelm

state: Hessen

address: Aulweg 132

department: Institute for Lung Health

institute: Justus-Liebig University Giessen

summary: Chronic obstructive pulmonary disease (COPD) is currently the third cause of death worldwide with still increasing mortality and morbidity. Primary etiology of COPD is cigarette smoking. However, in clinic, not all smokers develop COPD. The underlying mechanism remains unclear. A549 cells, which are widely used in vitro as a model of alveolar type II pulmonary epithelium, were subjected to step-wise increasing cigarette smoke extract (CSE) treatments. Those cigarette smoke extract resistant (SER) cells were cultured and used for further experiments. The aim of this study is to investigate the differentially expressed genes in SER group with or without CSE treatment and identify potential genes or pathways which could play a role in COPD pathogenesis.

overall_design: 6 groups (2 genotypes x 3 time points), each with 4 biological replicates

first: Cheng-Yu

last: Wu

first: Norbert

last: Weissmann